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WO2014176284A1 - Selective drug delivery compositions and methods of use - Google Patents

Selective drug delivery compositions and methods of use Download PDF

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Publication number
WO2014176284A1
WO2014176284A1 PCT/US2014/035043 US2014035043W WO2014176284A1 WO 2014176284 A1 WO2014176284 A1 WO 2014176284A1 US 2014035043 W US2014035043 W US 2014035043W WO 2014176284 A1 WO2014176284 A1 WO 2014176284A1
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WO
WIPO (PCT)
Prior art keywords
sdm
amino acid
molecule
peg
peptide
Prior art date
Application number
PCT/US2014/035043
Other languages
French (fr)
Inventor
Jesus Gonzalez
Junjie Liu
Marcel MIAMPAMBA
Original Assignee
Avelas Biosciences, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Avelas Biosciences, Inc. filed Critical Avelas Biosciences, Inc.
Priority to US14/786,402 priority Critical patent/US20160082119A1/en
Priority to EP14788083.5A priority patent/EP2988786A4/en
Publication of WO2014176284A1 publication Critical patent/WO2014176284A1/en
Priority to HK16110317.7A priority patent/HK1222122A1/en
Priority to US16/450,836 priority patent/US20190374560A1/en
Priority to US16/988,398 priority patent/US20200376013A1/en

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
    • A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
    • A61K31/704—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin attached to a condensed carbocyclic ring system, e.g. sennosides, thiocolchicosides, escin, daunorubicin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/56—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
    • A61K31/57—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids substituted in position 17 beta by a chain of two carbon atoms, e.g. pregnane or progesterone
    • A61K31/573—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids substituted in position 17 beta by a chain of two carbon atoms, e.g. pregnane or progesterone substituted in position 21, e.g. cortisone, dexamethasone, prednisone or aldosterone
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
    • A61K47/59—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes
    • A61K47/60—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
    • A61K47/645—Polycationic or polyanionic oligopeptides, polypeptides or polyamino acids, e.g. polylysine, polyarginine, polyglutamic acid or peptide TAT
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A61K47/65—Peptidic linkers, binders or spacers, e.g. peptidic enzyme-labile linkers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • Targeted delivery of therapeutic agents, such as cytotoxic agents, to tumor cells is desirable to avoid killing normal cells following systemic administration of such agents.
  • Typical targeted drug delivery systems are composed of a cytotoxic agent conjugated to a tumor-specific antibody, forming an antibody-drug conjugate (ADC), also called an "immunoconjugate".
  • ADC antibody-drug conjugate
  • the tumor-specific antibody binds to a tumor biomarker (e.g. a tumor antigen) expressed on the surface of the tumor cells.
  • the ADC will selectively bind to tumor cells in the body, and thereby deliver the therapeutic agent intracellularly to the tumor cells, and not normal cells.
  • the cytotoxic agent is not active when conjugated to the antibody, but becomes active upon being cleaved from the antibody intracellularly.
  • ADCs include gemtuzumab ozogamicin (Mylotarg), brentuximab vendotin (Adcetris), trastuzumab emtasine (Kadcyla).
  • compositions for the delivery of therapeutic agents are described herein.
  • selective delivery molecule conjugates comprising: (a) a selective delivery molecule of Formula I, having the structure:
  • X is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids
  • C B is 0- 1 amino acid
  • D B is a therapeutic agent or an imaging agent
  • the carrier or targeting ligand is covalently bound to any amino acid of A.
  • the carrier or targeting ligand is covalently bound to any amino acid of B.
  • the targeting ligand is an antibody.
  • the selective delivery molecule is covalently bound to any amino acid on the targeting antibody.
  • the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody.
  • the targeting ligand binds to a tumor antigen or tumor-specific receptor.
  • the targeting antibody binds to CD3, CD19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 (NCAM), CS-125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or
  • the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203.
  • the targeting ligand binds to a tumor antigen or tumor-specific receptor.
  • the targeting ligand is an integrin or a lectin.
  • the carrier is a polyethylene glycol (PEG) polymer.
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the therapeutic agent is a chemotherapeutic agent, a steroid, an
  • the therapeutic agent is a cytotoxin.
  • the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, or auritstatin.
  • the therapeutic agent is cortisone.
  • a and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A.
  • A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines.
  • A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some
  • CB is selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CB is selected from a D amino acid, a L amino acid, an a-amino acid, a ⁇ -amino acid, or a ⁇ -amino acid. In some embodiments, CB is selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a
  • X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14.
  • X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQL L, and RLQLK(Ac).
  • the selective delivery molecule of Formula I is: SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM- 1 10, SDM-1 11, SDM-112, SDM-113, SDM-114, SDM-1 15, SDM-116, SDM-117, SDM-1 18, SDM-1 19, SDM-120, SDM-121 , SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM- 127, SDM-128, SDM-129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141 , SDM-142, SDM-143, SDM- 144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-101
  • selective delivery molecule conjugates comprising: (a) a selective delivery molecule of Formula II, having the structure:
  • X is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids
  • CB and CM each independently comprise 0-1 amino acid
  • ⁇ is a macromolecule
  • DB is a therapeutic agent or an imaging agent
  • the carrier or targeting ligand is covalently bound to any amino acid of A. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of B. In some embodiments, the targeting ligand is an antibody. In some embodiments, the selective delivery molecule is covalently bound to any amino acid on the targeting antibody. In some embodiments, the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody. In some embodiments, the targeting antibody binds to a tumor antigen or a tumor antigen or tumor-specific receptor.
  • the targeting antibody binds to CD3, CD19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 (NCAM), CS-125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or
  • the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203.
  • the targeting ligand binds to a tumor antigen or tumor-specific receptor.
  • the targeting ligand is an integrin or a lectin.
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the therapeutic agent is a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, or an anti-inflammatory agent.
  • the therapeutic agent is a cytotoxin.
  • the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, or auritstatin.
  • the therapeutic agent is cortisone.
  • a and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A.
  • A is a peptide comprising 5 or 9 consecutive glutamates.
  • B is a peptide comprising 8 or 9 consecutive arginines.
  • A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines.
  • A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines.
  • CB and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid.
  • CB and CM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a ⁇ -amino acid, or a y-amino acid.
  • CB and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CB and CM are each independently selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • C M is para-4-acetyl L-phenylalanine.
  • X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease.
  • X is cleavable by a matrix metalloproteinase.
  • X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14.
  • X comprises a peptide linkage.
  • X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPPvSFL, PPRSFL, RLQLKL, and RLQLK(Ac).
  • M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer.
  • M is selected from dextran, a polyethylene glycol (PEG) polymer, albumin, or a combination thereof.
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa),
  • the selective delivery molecule of Formula II is: SDM-101, SDM-102, SDM- 103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM-110, SDM-111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM-118, SDM-119, SDM- 120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM- 137, SDM-138, SDM-139, SDM-140, SDM-141, SDM-142, SDM-143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, S
  • selective delivery molecule conjugates comprising: (a) a selective delivery molecule of Formula V, having the structure:
  • X is a cleavable linker
  • ⁇ is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids
  • C B and C M each independently comprise 0-1 amino acid; M is a macromolecule;
  • D B is a therapeutic agent or an imaging agent
  • the carrier or targeting ligand is covalently bound to any amino acid of A. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of B. In some embodiments, the targeting ligand is an antibody. In some embodiments, the selective delivery molecule is covalently bound to any amino acid on the targeting antibody. In some embodiments, the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody. In some embodiments, the targeting antibody binds to a tumor antigen or a tumor antigen or tumor-specific receptor.
  • the targeting antibody binds to CD3, CD19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 ( CAM), CS-125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or
  • the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203.
  • the targeting ligand binds to a tumor antigen or tumor-specific receptor.
  • the targeting ligand is an integrin or a lectin.
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the therapeutic agent is a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, or an anti-inflammatory agent.
  • the therapeutic agent is a cytotoxin.
  • the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, or auritstatin.
  • the therapeutic agent is cortisone.
  • a and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A.
  • A is a peptide comprising 5 or 9 consecutive glutamates.
  • B is a peptide comprising 8 or 9 consecutive arginines.
  • A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines.
  • A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines.
  • CB and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid.
  • CB and CM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a ⁇ -amino acid, or a r-amino acid.
  • CB and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CB and CM are each independently selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CM is para-4-acetyl L-phenylalanine.
  • X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease.
  • X is cleavable by a matrix metalloproteinase.
  • X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14.
  • X comprises a peptide linkage.
  • X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac).
  • Y is cleavable by a protease. In some embodiments, Y is cleavable by an intracellular protease.
  • Y is cleavable by a lysosomal protease. In some embodiments, Y is cleavable by Cathepsin B. In some embodiments, Y comprises a self-immolative spacer. In some embodiments, Y comprises a PABC spacer or a derivative thereof. In some embodiments, M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a polyethylene glycol (PEG) polymer, albumin, or a combination thereof.
  • PEG polyethylene glycol
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa),
  • PEG 5kDa approximately 5kDa
  • PEG lOkDa approximately 12kDa
  • PEG 12kDa approximately 15kDa
  • 20kDa approximately 20kDa
  • PEG 30kDa approximately 30kDa
  • 40kDa PEG 40kDa
  • the selective delivery molecule of Formula V is: SDM-101, SDM-102, SDM-103, SDM-104, SDM-105, SDM- 106, SDM-107, SDM-108, SDM-109, SDM-110, SDM-111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM-118, SDM-119, SDM-120, SDM-121, SDM-122, SDM- 123, SDM-124, SDM-125, SDM-126, SDM-127, SDM- 128, SDM- 129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM-134, SDM-135, SDM- 136, SDM- 137, SDM-138, SDM-139, SDM- 140, SDM-141 , SDM-142, SDM-143, SDM-144, SDM- 145, SDM- 146, SDM-
  • X is a cleavable linker
  • Y is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids
  • C B and C M each independently comprise 0-1 amino acid
  • M is a macromolecule
  • D B is a therapeutic agent or an imaging agent
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In
  • C B and C M are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid.
  • C B and C M are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a ⁇ -amino acid, or a ⁇ -amino acid.
  • C B and C M are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CB and CM are each independently selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CM is para-4-acetyl L-phenylalanine.
  • X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease.
  • X is cleavable by a matrix metalloproteinase.
  • X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14.
  • X comprises a peptide linkage.
  • X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac).
  • Y is cleavable by a protease. In some embodiments, Y is cleavable by an intracellular protease.
  • Y is cleavable by a lysosomal protease. In some embodiments, Y is cleavable by Cathepsin B. In some embodiments, Y comprises a self-immolative spacer. In some embodiments, Y comprises a PABC spacer or a derivative thereof.
  • the therapeutic agent is a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, or an antiinflammatory agent. In some embodiments, the therapeutic agent is a cytotoxin. In some embodiments, the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, or auritstatin.
  • the therapeutic agent is cortisone.
  • M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer.
  • M is selected from dextran, a polyethylene glycol (PEG) polymer, albumin, or a combination thereof.
  • PEG polyethylene glycol
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa),
  • PEG 5kDa approximately 5kDa
  • PEG lOkDa approximately 12kDa
  • PEG 12kDa approximately 15kDa
  • 20kDa approximately 20kDa
  • PEG 30kDa approximately 30kDa
  • 40kDa PEG 40kDa
  • molecule is SDM-101, SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM- 109, SDM-110, SDM-111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM-118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM- 126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141, SDM-142, SDM- 143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM
  • the selective delivery molecule is covalently bound to a carrier or targeting ligand.
  • the carrier or targeting ligand is covalently bound to any amino acid of A.
  • the carrier or targeting ligand is covalently bound to any amino acid of B.
  • the targeting ligand is an antibody.
  • the selective delivery molecule is covalently bound to any amino acid on the targeting antibody.
  • the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody.
  • the targeting antibody binds to a tumor antigen or a tumor antigen or tumor-specific receptor.
  • the targeting antibody binds to CD3, CD19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 (NCAM), CS- 125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or SLC44A4 (AGS-5).
  • the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab,
  • the targeting ligand binds to a tumor antigen or tumor-specific receptor.
  • the targeting ligand is an integrin or a lectin.
  • the carrier or targeting ligand is covalently bound to any amino acid of A.
  • the carrier or targeting ligand is covalently bound to any amino acid of B.
  • the targeting ligand is an antibody.
  • the selective delivery molecule is covalently bound to any amino acid on the targeting antibody.
  • the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody.
  • the targeting antibody binds to a tumor antigen or a tumor antigen or tumor-specific receptor.
  • the targeting antibody binds to CD3, CD 19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 (NCAM), CS-125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or
  • the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203.
  • the targeting ligand binds to a tumor antigen or tumor-specific receptor.
  • the targeting ligand is an integrin or a lectin.
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylatmg agents, vinylsulfones, pyridyl disulfides, TNB-tbiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • compositions comprising a selective delivery molecule conjugate provided herein and one or more pharmaceutically acceptable carriers, glidants, diluents, or excipients.
  • the cancer is a breast cancer, colorectal cancer, ovarian cancer, lung cancer, esophageal cancer, pancreatic cancer, gastro-intestinal cancer, squamous cell carcinoma, prostate cancer, melanoma, or thyroid cancer.
  • the therapeutic agent is a
  • the methods further comprise administering an additional anti-cancer agent.
  • the inflammation is acute inflammation or chronic inflammation.
  • the inflammation is associated with rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, sepsis, erythema nodosum leprosum, multiple sclerosis, psoriasis, systemic lupus erythematosis, type I diabetes, atherosclerosis,
  • the therapeutic agent is an anti-inflammatory agent.
  • the therapeutic agent is a steroid.
  • the autoimmune disease is Celiac disease, diabetes mellitus type 1, Sarcoidosis, systemic lupus erythematosus (SLE), Sjogren's syndrome, Churg- Strauss Syndrome, Hashimoto's thyroiditis, Graves' disease, idiopathic thrombocytopenic purpura, Addison's Disease, rheumatoid arthritis (RA), Polymyositis (PM), or Dermatomyositis (DM).
  • the methods further comprise imaging the cancer.
  • the methods further comprise imaging the site of
  • Figure 1 Cleavage of SDM-145 by hMMP-9.
  • LC-MS confirmed that the molecular weight of the peak at ⁇ 9.4 min was consistent with the fragment generated by hMMP-9 cleavage at the expected cleavage site. The fragment's chemical structure was shown in the chromatogram.
  • Figure 2 Cleavage of SDM-145 by Cathepsin B.
  • LC- MS confirmed that the molecular weight of the peak at ⁇ 9.0 min was consistent with the freed doxorubicin.
  • Figure 3 illustrates schematics of exemplary protease activated antibody conjugates and protease cleavage steps.
  • Figure 4 illustrates a schematic of exemplary dual protease activated drug delivery conjugate and protease cleavage steps.
  • Figure 5 illustrates a higher resolution schematic of exemplary dual protease activated drug delivery conjugate.
  • Figure 6 illustrates an exemplary scheme for protease cleavage and uptake of a dual protease activated drug delivery conjugate: (a) extracellular proteases (e.g. matrix
  • metalloproteinases cleave conjugate near target cells
  • CPP cell penetrating peptide
  • lysosomal proteases e.g. Cathepsin B
  • active therapeutic cargo e.g. Doxorubicin
  • FIG. 7 illustrates a schematic of exemplary thiol-reactive drug delivery conjugates.
  • the SDMs react efficiently with albumin Cys(34)-SH in circulation albumin after injected into blood stream.
  • Albumin-SDM conjugates have improved pharmacokinetic profiles and efficient targeted cargo delivery.
  • Figure 8 illustrates MALDI-TOF Spectrum of SDM-147.
  • Figure 9 illustrates therapeutic activity of SDM-147 in a 4T1 breast cancer mouse model.
  • ranges and amounts can be expressed as “about” a particular value or range. About also includes the exact amount. Hence “about 40 mg” means “about 40 mg” and also “40 mg.” Generally, the term “about” includes an amount that would be expected to be within experimental error.
  • the terms "individual,” “patient,” or “subject” are used interchangeably. As used herein, they mean any mammal (i.e. species of any orders, families, and genus within the taxonomic classification animalia: chordata: vertebrata: mammalia). In some embodiments, the mammal is a human. None of the terms require or are limited to situation characterized by the supervision (e.g. constant or intermittent) of a health care worker (e.g. a doctor, a registered nurse, a nurse practitioner, a physician's assistant, an orderly, or a hospice worker).
  • a health care worker e.g. a doctor, a registered nurse, a nurse practitioner, a physician's assistant, an orderly, or a hospice worker.
  • alkyl refers to an aliphatic hydrocarbon group.
  • the alkyl moiety may be a saturated alkyl or an unsaturated alkyl.
  • an alkyl group can be a monoradical or a diradical (i.e., an alkylene group).
  • the "alkyl” moiety may have 1 to 10 carbon atoms (whenever it appears herein, a numerical range such as “1 to 10" refers to each integer in the given range; e.g., "1 to 10 carbon atoms” means that the alkyl group may consist of 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and including 10 carbon atoms, although the present definition also covers the occurrence of the term "alkyl” where no numerical range is designated).
  • the alkyl group could also be a "lower alkyl” having 1 to 6 carbon atoms.
  • the alkyl group of the compounds described herein may be designated as "C1-C4 alkyl" or similar designations.
  • C1-C4 alkyl indicates that there are one to four carbon atoms in the alkyl chain, i.e., the alkyl chain is selected from: methyl, ethyl, propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, and t-butyl.
  • Typical alkyl groups include, but are in no way limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tertiary butyl, pentyl, hexyl, ethenyl, propenyl, butenyl, and the like.
  • the linker comprises a ring structure (e.g., an aryl).
  • ring refers to any covalently closed structure. Rings include, for example, carbocycles (e.g., aryls and cycloalkyls), heterocycles (e.g., heteroaryls and non-aromatic heterocycles), aromatics (e.g. aryls and heteroaryls), and non-aromatics (e.g., cycloalkyls and non-aromatic heterocycles). Rings can be optionally substituted. Rings can be monocyclic or polycyclic.
  • aryl refers to an aromatic ring wherein each of the atoms forming the ring is a carbon atom.
  • Aryl rings can be formed by five, six, seven, eight, nine, or more than nine carbon atoms.
  • Aryl groups can be optionally substituted. Examples of aryl groups include, but are not limited to phenyl, naphthalenyl, phenanthrenyl, anthracenyl, fluorenyl, and indenyl.
  • an aryl group can be a monoradical or a diradical (i.e., an arylene group).
  • cycloalkyl refers to a monocyclic or polycyclic non-aromatic radical, wherein each of the atoms forming the ring (i.e. skeletal atoms) is a carbon atom. Cycloalkyls may be saturated, or partially unsaturated. Cycloalkyl groups include groups having from 3 to 10 ring atoms. Cycloalkyls include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.
  • the ring is a cycloalkane. In some embodiments, the ring is a cycloalkene.
  • the ring is an aromatic ring.
  • aromatic refers to a planar ring having a delocalized ⁇ -electron system containing 4n+2 ⁇ electrons, where n is an integer.
  • Aromatic rings can be formed from five, six, seven, eight, nine, or more than nine atoms.
  • Aromatics can be optionally substituted.
  • aromatic includes both carbocyclic aryl (e.g., phenyl) and heterocyclic aryl (or “heteroaryl” or “heteroaromatic”) groups (e.g., pyridine).
  • heterocyclic aryl or “heteroaryl” or “heteroaromatic” groups (e.g., pyridine).
  • the term includes monocyclic or fused-ring polycyclic (i.e., rings which share adjacent pairs of carbon atoms) groups.
  • the ring is a heterocycle.
  • heterocycle refers to heteroaromatic and heteroalicyclic groups containing one to four heteroatoms each selected from O, S and N, wherein each heterocyclic group has from 4 to 10 atoms in its ring system, and with the proviso that the ring of said group does not contain two adjacent O or S atoms.
  • Non-aromatic heterocyclic groups include groups having only 3 atoms in their ring system, but aromatic heterocyclic groups must have at least 5 atoms in their ring system.
  • the heterocyclic groups include benzo-fused ring systems.
  • An example of a 3-membered heterocyclic group is aziridinyl.
  • An example of a 4-membered heterocyclic group is azetidinyl (derived from azetidine).
  • An example of a 5-membered heterocyclic group is thiazolyl.
  • An example of a 6-membered heterocyclic group is pyridyl, and an example of a 10-membered heterocyclic group is quinolinyl.
  • non- aromatic heterocyclic groups are pyrrolidinyl, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, dihydropyranyl, tetrahydrothiopyranyl, piperidino, morpholino, thiomorpholino, thioxanyl, piperazinyl, azetidinyl, oxetanyl, thietanyl, homopiperidinyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl, thiazepinyl, 1,2,3,6-tetrahydropyridinyl, 2-pyrrolinyl, 3-pyrrolinyl, indolinyl, 2H-pyranyl, 4H-pyranyl, dioxanyl, 1,3-dioxolanyl, pyrazolinyl, dithianyl,
  • aromatic heterocyclic groups are pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinox
  • a group derived from pyrrole may be pyrrol- 1-yl (reattached) or pyrrol-3-yl (C-attached).
  • a group derived from imidazole may be imidazol-1- yl or imidazol-3-yl (both N-attached) or imidazol-2-yl, imidazol-4-yl or imidazol-5-yl (all C- attached).
  • a heterocycle group can be a monoradical or a diradical (i.e., a heterocyclene group).
  • the ring is fused.
  • fused refers to structures in which two or more rings share one or more bonds.
  • the ring is a dimer.
  • the ring is a trimer.
  • the ring is a substituted.
  • Carbocyclic or “carbocycle” refers to a ring wherein each of the atoms forming the ring is a carbon atom.
  • Carbocycle includes aryl and cycloalkyl. The term thus distinguishes carbocycle from heterocycle ("heterocyclic") in which the ring backbone contains at least one atom which is different from carbon (i.e., a heteroatom).
  • Heterocycle includes heteroaryl and
  • heterocycloalkyl Carbocycles and heterocycles can be optionally substituted.
  • the linker is substituted.
  • the term "optionally substituted” or “substituted” means that the referenced group may be substituted with one or more additional group(s) individually and independently selected from Ci-Cealkyl, C 3 -Cscycloalkyl, aryl, heteroaryl, C2-C6heteroalicyclic, hydroxy, Ci-Cealkoxy, aryloxy, Ci-C6alkylthio, arylthio, Ci- C 6 alkylsulfoxide, arylsulfoxide, Ci-C 6 alkylsulfone, arylsulfone, cyano, halo, C 2 -C 8 acyl, C 2 - Csacyloxy, nitro, Ci-Cehaloalkyl, Ci-Cefluoroalkyl, and amino, including Ci-Cealkylamino, and the protected derivatives thereof.
  • selective delivery molecules allow the targeted delivery of therapeutic agents and/or imaging agents to specific cells and/or tissues.
  • selective delivery molecules comprise (a) an acidic sequence (portion of A) which is effective to inhibit or prevent uptake of the molecule into cells or tissue retention, (b) a molecular transport or retention sequence (portion of B) (e.g., a cell penetrating peptide (CPP), (c) a cleavable linker X located between portion of A and portion of B, (d) at least one cargo moiety (portion D) bound to portion of B, and optionally (e) a macromolecular carrier bound to portion of A.
  • portion of A an acidic sequence
  • portion of B e.g., a cell penetrating peptide (CPP)
  • CPP cell penetrating peptide
  • cleavage of the X linker allows the separation of portion of A from portion of B, thereby promoting the uptake or retention of portion of B and the attached cargo into cells or tissue retention.
  • the therapeutic cargo is a chemotherapeutic agent.
  • the therapeutic cargo is a cytotoxin.
  • conjugating a selective delivery molecule disclosed herein to a targeting ligand allows the SDM to be targeted to specific cells having a specific cell surface marker.
  • a targeting ligand such as an antibody
  • the cell specific delivery of an existing targeting ligand-drug conjugate is improved by attachment of a selective delivery molecule described herein to the targeting ligand.
  • the targeting antibody binds to a tumor antigen or a receptor that is upregulated in tumor cells. Accordingly, provided herein are improved antibody-drug conjugates for delivery of therapeutic agents to target cells and tissues, such as cancer cells and other diseased cells.
  • the targeting ligand-conjugated SDMs described herein provide significant advantages over existing targeting ligand-drug conjugates (e.g., antibody-drug conjugates). In some embodiments, the targeting ligand-conjugated SDMs described herein provide improved tumor penetration and retention over existing targeting ligand -drug conjugates. In some embodiments, the targeting ligand -conjugated SDMs described herein provide dual targeting specificity. In some embodiments, the dual targeting mechanism comprises 1) ligand (e.g., antibody) targeting of cell specific markers on diseased cells and 2) pathological protease activity targeting of increased extracellular protease activity at physiological location of the diseased cell.
  • ligand e.g., antibody
  • the targeting ligand-conjugated SDMs described herein provide different options for conjugating a variety of therapeutic cargo molecules for delivery to the diseased cell.
  • a variety of configurations is available for the attachment of the therapeutic cargo to the SDM or the ligand (e.g., antibody) itself.
  • Figure 3 illustrates exemplary configurations of targeting ligand-conjugated SDMs, where the targeting ligand is an antibody.
  • the therapeutic cargo is internalized with the targeting ligand.
  • the therapeutic cargo is released from the targeting ligand-conjugated SDM prior to uptake by the cell.
  • the modular design of the targeting ligand-conjugated SDMs described herein enables easy modification of the conjugates to change protease recognition sites and therapeutic cargo molecules.
  • increased retention and penetration of the cells achieved by the SDM conjugates provided herein increase the efficacy of the therapeutic cargo.
  • a lower dosage of the therapeutic cargo is employed compared to existing targeting ligand-drug conjugates that lack an SDM.
  • a less toxic therapeutic cargo is needed to treat the target cell.
  • increased retention and penetration of the cells achieved by the conjugates provided herein allows for less toxic therapeutic cargo molecules to be employed compared to existing targeting ligand-drug conjugates that lack an SDM.
  • SDMs that provide single protease targeting. For example, in some embodiments, cleavage of the X linker located between portion of A and portion of B of the SDM by a protease located near the target cell allows the separation of portion of A from portion of B, thereby promoting the uptake or retention of portion of B and the attached cargo into cells or tissue retention.
  • the protease exhibits higher expression in the extracellular area surrounding the target cell (e.g., a cancer cell) as compared to a non-target cell (e.g., non-diseased/non-cancerous cell).
  • the protease is a matrix metalloproteinase (MMP).
  • MMP matrix metalloproteinase
  • the therapeutic cargo is a
  • the therapeutic cargo is a cytotoxin.
  • the therapeutic cargo is doxorubicin, calicheamicin, maytansinoid, or auritstatin.
  • the therapeutic cargo is an anti-inflammatory agent.
  • the therapeutic cargo is a steroid.
  • the therapeutic cargo is cortisone or a derivative thereof.
  • the SDM is conjugated to a targeting ligand.
  • the SDM comprises an X linker located between portion of A and portion of B and a second linker Y located between portion of B and the therapeutic cargo molecule (portion D).
  • the X linker comprises an extracellular protease cleavage site and the Y linker comprises an intracellular protease cleavage site.
  • cleavage of the X linker located between portion of A and portion of B of the selective delivery molecule by a protease located near the target cell allows the separation of portion of A from portion of B, thereby promoting the uptake or retention of portion of B and the attached cargo into cells or tissue retention, and then cleavage of the Y linker by an intracellular protease allows the therapeutic cargo to be released into the cell after uptake.
  • such configurations allow therapeutic cargos to be maintained in an inactive state until the therapeutic cargos are taken up by the cell following cleavage of the X linker.
  • the extracellular protease that cleaves the X linker exhibits a higher expression in the extracellular area surrounding the target cell (e.g., a cancer cell) as compared to a non-target cell (e.g., non-diseased/non-cancerous cell).
  • the extracellular protease is a matrix metalloproteinase (MMP).
  • MMP matrix metalloproteinase
  • the intracellular protease that cleaves the Y linker is a lysosomal protease.
  • the protease is one that is activated at low pH. In some embodiments, the protease is one that is activated at low pH and is expressed in an endosome. In some embodiments, the protease is a cathepsin. In some embodiments, the cathepsin is cathepsin B or cathepsin D. In some embodiments, the cathepsin is cathepsin B. In some embodiments, the Y linker additionally comprises a self-immolative cleavage site located between the intracellular protease cleavage site and the attached therapeutic cargo.
  • the self-immolative cleavage site is a PABC (p-aminobenzylcarbonyl) spacer and analogs thereof.
  • the self-immolative cleavage site is a thiazole containing linker.
  • the selective delivery molecule comprising an X linker located between portion of A and portion of B and a second linker Y located between portion of B and the therapeutic cargo molecule (portion D) is conjugated to a targeting ligand, such as a targeting antibody.
  • the targeting ligand binds to a tumor antigen or a receptor that is upregulated in tumor cells.
  • the SDM is conjugated to a targeting ligand.
  • Figures 4 and 5 illustrate schematics of exemplary dual protease drug delivery conjugates.
  • Figure 6 illustrates delivery of an antibody-SDM conjugate with dual protease targeting.
  • the SDMs provided herein are conjugated to albumin and/or contain a free thiol reactive group for interacting with albumin in vivo.
  • Albumin is a carrier for tumor targeting because it accumulates in solid tumors due to the pathophysiology of tumor tissue, characterized by a high metabolic turnover, angiogenesis, hypervasculature, a defective vascular architecture and an impaired lymphatic drainage.
  • Albumin-drug conjugates show improved the pharmacokinetic profiles.
  • albumin conjugates have limited tumor penetration and distribution due to their big molecular size and the tumor tissue's microenvironment, such as increased interstitial fluid pressure and dense extracellular matrix.
  • thiol-reactive SDMs provided herein form albumin conjugates in vivo.
  • the conjugates increase the drug's tumor penetration.
  • the conjugates increase improve drug's distribution and activity.
  • thiol-reactive SDMs react with the free Cys34 thiol of the circulating albumin. The albumin-SDM conjugate is then transported and accumulated in the tumor tissues.
  • the up-regulated MMPs in tumor tissues cleave the MMPs sensitive linker in ACPP part and release the poly-Arg-drug fragment.
  • poly-Arg has excellent cell penetrating capability
  • the poly-Arg-drug fragment is able to efficiently bind to the tumor cell and get internalized.
  • drug is regenerated after an enzymatic cleavage by intracellular proteases, such as Cathepsin B in the lysosome.
  • the thiol reactive group of the SDM is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • Figure 7 illustrates exemplary schematics of exemplary thiol- reactive SDMs.
  • SDMs Selective Delivery Molecules
  • a carrier modulates plasma half-life of a selective delivery molecule disclosed herein. In some embodiments, a carrier modulates solubility of a selective delivery molecule disclosed herein. In some embodiments, a carrier modulates bio-distribution of a selective delivery molecule disclosed herein.
  • a carrier decreases uptake of a selective delivery molecule by non- target cells or tissues. In some embodiments, a carrier decreases uptake of a selective delivery molecule into cartilage. In some embodiments, a carrier decreases uptake of a selective delivery molecule into joints relative to target tissue.
  • a carrier increases uptake of a selective delivery molecule by target cells or tissues. In some embodiments, a carrier decreases uptake of a selective delivery molecule into the liver relative to target tissue. In some embodiments, a carrier decreases uptake of a selective delivery molecule into kidneys. In some embodiments, a carrier enhances uptake into cancer tissue. In some embodiments, a carrier enhances uptake into lymphatic channels and/or lymph nodes.
  • a carrier increases plasma half-life by reducing glomerular filtration. In some embodiments, a carrier modulates plasma half-life by increasing or decreases metabolism or protease degradation. In some embodiments, a carrier increases tumor uptake due to enhanced permeability and retention (EPR) of tumor vasculature. In some embodiments, a carrier increases the aqueous solubility of selective delivery molecule.
  • EPR enhanced permeability and retention
  • any carrier is independently directly or indirectly (e.g., via CM) bound to A, B, or X.
  • any carrier is independently bound to A at the n- terminal poly glutamate.
  • any carrier is independently bound to A (or, the n- terminal poly glutamate) by a covalent linkage.
  • any carrier is independently bound to B at the c-terminal polyarginine.
  • any carrier is independently bound to B (or, the c-terminal polyarginine) by a covalent linkage.
  • any carrier is independently directly or indirectly bound to linkers between X and A, X and B, B and C N terminus, and A and C/N terminus.
  • the covalent linkage comprises an ether bond, thioether bond, amine bond, amide bond, oxime bond, carbon-carbon bond, carbon- nitrogen bond, carbon-oxygen bond, or carbon-sulfur bond.
  • carrier is selected from a macromolecule such as a protein, a synthetic or natural polymer, or a dendrimer.
  • carrier is selected from dextran, a PEG polymer (e.g., a PEG polymer having an average molecular weight of
  • PEG 0.5kDa approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa),
  • carrier is a PEG polymer.
  • the size of carrier is between about 50kDa and about 70kDa.
  • the selective delivery molecule is conjugated to albumin.
  • albumin is excluded from the glomerular filtrate under normal physiological conditions.
  • the selective delivery molecule comprises a reactive group such as maleimide that can form a covalent conjugate with albumin.
  • a selective delivery molecule comprising albumin results in enhanced accumulation of cleaved selective delivery molecules in tumors in a cleavage dependent manner.
  • albumin conjugates have good pharmacokinetic properties.
  • the selective delivery molecule is conjugated to PEG polymers. In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately IkDa (PEG IkDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 2kDa (PEG 2kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 3kDa (PEG 3kDa).
  • the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 4kDa (PEG 4kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 5kDa (PEG 5kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately lOkDa (PEG lOkDa). In some embodiments, the selective delivery molecule is conjugated PEG polymers having an average molecular weight of approximately 12 kDa ( PEG 12kDa).
  • the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 15kDa (PEG 15kDa). In some embodiments, selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 20 kDa (PEG 20kDa). In some embodiments, selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 30 kDa (PEG 30kDa). In some embodiments, selective delivery molecules conjugated to PEG30kDa had a longer half-life as compared to free peptides. In some
  • selective delivery molecules are conjugated to PEG polymers having an average molecular weight of between about 20 to about 40kDa which have hepatic and renal clearance.
  • the selective delivery molecule is conjugated to a dextran.
  • the selective delivery molecule is conjugated to a dextran having an average molecular weight of approximately 70kDa.
  • dextran conjugates being a mixture of molecular weights, are difficult to synthesize and purify reproducibly.
  • the selective delivery molecule is conjugated to streptavidin.
  • the selective delivery molecule is conjugated to a fifth generation PAMAM dendrimer.
  • a carrier is capped.
  • capping a carrier improves the pharmacokinetics and reduces cytotoxicity of a carrier by adding hydrophilicity.
  • the cap is selected from: Acetyl, succinyl, 3-hydroxypropionyl, 2-sulfobenzoyl, glycidyl, PEG-2, PEG-4, PEG-8 and PEG-12.
  • a therapeutic cargo and an SDM are conjugated to a targeting ligand.
  • the SDM comprises a therapeutic cargo and the SDM comprising the therapeutic cargo is conjugated to a targeting ligand.
  • the therapeutic cargo and the SDM are conjugated to the same site on the targeting ligand.
  • the therapeutic cargo is first conjugated to an SDM, and then the SDM comprising the therapeutic cargo is attached to the targeting ligand.
  • the therapeutic cargo and the SDM are conjugated to two different sites on the targeting ligand.
  • the SDM is conjugated to an existing ligand- drug conjugate.
  • an SDM is conjugated to a targeting ligand, and then a therapeutic cargo is conjugated to the targeting ligand -SDM conjugate.
  • a therapeutic cargo is conjugated to a targeting ligand, and then an SDM is conjugated to the targeting ligand-therapeutic cargo conjugate.
  • the targeting ligand is conjugated to the acidic sequence (portion of A) of an SDM.
  • the targeting ligand is conjugated to molecular transport or retention sequence (portion of B) of an SDM.
  • any of a variety of known methods for conjugation of molecules to polypeptides such as targeting ligands are employed for the conjugation of the therapeutic cargo and/or SDMs provided herein.
  • a therapeutic cargo and an SDM are conjugated to a targeting antibody.
  • the SDM comprises a therapeutic cargo and the SDM comprising the therapeutic cargo is conjugated to a targeting antibody.
  • the therapeutic cargo and the SDM are conjugated to the same site on the targeting antibody (e.g., Form 1 and Form 2 of Fig. 3).
  • the therapeutic cargo is first conjugated to an SDM, and then the SDM comprising the therapeutic cargo is attached to the targeting antibody.
  • the therapeutic cargo and the SDM are conjugated to two different sites on the targeting antibody (e.g., Form 3 of Fig. 3).
  • the SDM is conjugated to an existing antibody-drug conjugate.
  • an SDM is conjugated to a targeting antibody, and then a therapeutic cargo is conjugated to the antibody-SDM conjugate.
  • a therapeutic cargo is conjugated to a targeting antibody, and then an SDM is conjugated to the antibody-therapeutic cargo conjugate.
  • the targeting antibody is conjugated to the acidic sequence (portion of A) of an SDM (Form 1, Fig. 3).
  • the targeting antibody is conjugated to molecular transport or retention sequence (portion of B) of an SDM (Form 2 and Form 3, Fig. 3).
  • any of a variety of known methods for conjugation of molecules to antibodies are employed for the conjugation of the therapeutic cargo and/or SDMs provided herein.
  • the targeting ligand is a molecule that binds to a cell surface molecule expressed on the surface of a target cell. In some embodiments, the targeting ligand binds to a receptor expressed on the surface of a target cell. In some embodiments, the targeting ligand binds to a cell surface antigen expressed on the surface of a target cell. In some embodiments, the targeting ligand binds to a carbohydrate, a polypeptide or glycoprotein expressed on the surface of a target cell. In some embodiments, the targeting ligand is a lectin or an integrin. In some embodiments, the targeting ligand is an antibody. In some embodiments, the targeting ligand is a targeting non-antibody. In some embodiments, the targeting ligand is a co-stimulatory molecule.
  • the targeting ligand is a ligand that binds to a cell surface molecule of a diseased cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule that is specific to the diseased cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the diseased cell compared to non- diseased cells. In some embodiments, the targeting ligand is an antibody. In some embodiments, the diseased cell is a cancer cell.
  • the targeting ligand is a ligand that binds to a cell surface molecule expressed by a hematopoietic cell.
  • the targeting ligand targets an inflammatory cell (e.g., neutrophil, macrophage, monocyte, eosinophil, basophil).
  • the targeting ligand targets a cell involved in an autoimmune disease (e.g. a lymphocyte, such as a B lymphocyte or a T lymphocyte).
  • the targeting ligand is a ligand that binds to a cell surface molecule of a cancer cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule that is specific to the cancer cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the cancer cell compared to non-cancer cells. In some embodiments, targeting ligand is a ligand that binds to a cell surface molecule that that is expressed by the cancer cell but not a non-cancer cell.
  • the targeting ligand is a ligand that binds to a tumor antigen. In some embodiments, the targeting ligand is a ligand that binds to a cell surface receptor that is upregulated in the tumor cell (i.e., has a higher expression) compared to a non-tumor cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface receptor that that is expressed by the tumor cell but not a non-tumor cell. In some embodiments, the targeting ligand is an antibody.
  • the targeting antibody used in the compositions and methods provided herein is an antibody that binds to a cell surface molecule on the targeted cell.
  • the targeting antibody is an antibody that binds to a cell surface molecule that is specific for the targeted cell.
  • the targeting antibody is an antibody that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the target cell compared to non-targeted cells.
  • the targeting antibody is an antibody that binds to a cell surface molecule of a diseased cell.
  • the targeting antibody is an antibody that binds to a cell surface molecule that is specific to the diseased cell.
  • the targeting antibody is an antibody that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the diseased cell compared to non- diseased cells.
  • the targeting antibody is an antibody that binds to a cell surface molecule of a cancer cell. In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule that is specific to the cancer cell. In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the cancer cell compared to non-cancer cells. In some embodiments, the targeting antibody binds to a cell surface molecule that that is expressed by the cancer cell but not a non- cancer cell. In some embodiments, the antibody binds to a tumor antigen.
  • the targeting antibody binds to a cell surface receptor that is upregulated in the tumor cell (i.e., has a higher expression) compared to a non-tumor cell. In some embodiments, the targeting antibody binds to a cell surface receptor that that is expressed by the tumor cell but not a non-tumor cell.
  • the targeting antibody is a tumor specific antibody.
  • the targeting antibody binds to CD3, CD19, CD20, CD22, CD25, CD30, CD33, CD52, interleukin-2 receptor (IL-2), HLA-DR10p, tenascin, CEA, MUC1, TAG72, EBBB2 receptor (HER2), CD56 (NCAM), CS-125, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), SLC44A4 (AGS-5), folate receptor, an integrin, such as ⁇ 3 -integrin, transferrin receptor, granulocyte-macrophage colony-stimulating factor (GM- CSF) receptor, aminopeptidase N (CD 13), galactosamine receptor, leutenizing hormone releasing hormone (LHRH) receptor, vascular endothelial growth factor (VEGF) receptor (FLK1), ROR1, mesothelin,
  • IL-2 interleukin-2 receptor
  • the targeting antibody is gemtuzumab, inotuumab, trastuzumab (Herceptin), HD37, M195, LMB2, lyml, 81C6, HMFG1, CC49, rituximab, epratuzumab, lorvotuzumab, 2C3, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN-75, PSMA ADC, ASG-5ME or mdx-1203.
  • the targeting antibody is a variant of gemtuzumab, inotuumab, trastuzumab (Herceptin), HD37, M195, LMB2, lyml, 81C6, HMFG1, CC49, rituximab, epratuzumab, lorvotuzumab, 2C3, imgn388, SAR3419, ⁇ 1 ⁇ 062, brentixumab, glembatumumab, SGN-75, PSMA ADC, ASG-5ME or mdx-1203.
  • the comprises an antigen-binding fragment of gemtuzumab, inotuumab, trastuzumab (Herceptin), HD37, M195, LMB2, lyml, 81C6, HMFG1, CC49, rituximab, epratuzumab, lorvotuzumab, 2C3, imgn388, SAR3419, ⁇ 1 ⁇ 062, brentixumab, glembatumumab, SGN-75, PSMA ADC, ASG-5ME or mdx-1203.
  • the targeting ligand is a non-antibody ligand that binds to a receptor.
  • the targeting ligand binds to folate receptor, avp3-integrin, transferrin receptor, GM-CSF receptor, aminopeptidase N (CD 13), galactosamine receptor and LHRH receptor.
  • the targeting ligand comprises RGD, NGR, folate, transferrin, GM-CSF, or galactosamine.
  • the targeting antibody is natural antibody. In some embodiments, the targeting antibody is a synthetic antibody. In some embodiments, the targeting antibody is a recombinant antibody. In some embodiments, the targeting antibody is an antibody fragment containing at least a portion of the variable region of the immunoglobulin molecule that retains the binding specificity ability of the full-length immunoglobulin. In some embodiments, the targeting antibody is any protein having a binding domain that is homologous or substantially homologous to an immunoglobulin antigen-binding domain (antibody combining site). In some embodiments, the targeting antibody is a multispecific antibodies (e.g., bispecific antibodies). In some embodiments, the targeting antibody is a human antibody or non-human antibody.
  • the targeting antibody is a humanized antibody. In some embodiments, the targeting antibody is a chimeric antibody. In some embodiments, the targeting antibody is an intrabody. In some embodiments, the targeting antibody is an antibody fragment, such as, but not limited to, Fab fragment, Fab' fragment, F(ab') 2 fragment, Fv fragment, disulfide-linked Fv (dsFv), Fd fragment, Fd' fragment, single-chain Fv (scFv), single-chain Fab (scFab), diabody, anti-idiotypic (anti-Id) antibody, or antigen-binding fragments of any of the above In some embodiments, the targeting antibody is any member of any immunoglobulin type (e.g., IgG, IgM, IgD, IgE, IgA and IgY), any class (e.g. IgGl , IgG2, IgG3, IgG4, IgAl and IgA2) or subclass (e.g., IgG
  • the targeting antibody is a monoclonal antibody. In some embodiments, the targeting antibody is an IgG antibody. In some embodiments, the targeting antibody is a monovalent antibody. In some embodiments, the targeting antibody is multivalent antibody. In some embodiments, the targeting antibody is a bivalent antibody. In some
  • the targeting antibody is an antibody fragment, such as a single-chain variable fragment (scFv). In some embodiments, the targeting antibody is a humanized antibody. In some embodiments, the targeting antibody is a variant of a known tumor specific antibody. In some embodiments, the targeting antibody is an antigen-binding fragment of a known tumor specific antibody. [0081] In some embodiments, the SDM and/or therapeutic cargo is conjugated to a portion of the antibody such that conjugation does not interfere with antibody-antigen binding. In some embodiments, the SDM and/or therapeutic cargo is conjugated to the Fc portion of the antibody. Selective delivery molecules
  • SDMs selective delivery molecules
  • the SDMs described herein are conjugated to a targeting ligand.
  • the targeting ligand is an antibody.
  • the SDMs described herein are conjugated to an existing antibody-drug conjugate.
  • the SDMs described herein comprise a therapeutic cargo.
  • the SDMs described herein comprising a therapeutic cargo are conjugated to a targeting ligand.
  • the targeting ligand is an antibody.
  • the SDM is an SDM of Formula I, having the structure:
  • X is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids
  • C B comprises 0- 1 amino acid
  • D b is a therapeutic agent
  • a and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A.
  • A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines.
  • C B is selected from a naturally-occurring amino acid or a non- naturally-occurring amino acid. In some embodiments, C B is selected from a D amino acid, a L amino acid, an a-amino acid, a ⁇ -amino acid, or a ⁇ -amino acid. In some embodiments, c B is selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxyl amine or hydrazine group.
  • CB is selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some
  • X comprises a peptide linkage.
  • X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQL (Ac).
  • X comprises the amino acid sequence PLGLAG.
  • X comprises the amino acid sequence PLG-C(me)-AG.
  • X comprises the amino acid sequence RPLALWRS.
  • X comprises the amino acid sequence DPRSFL.
  • X comprises the amino acid sequence RLQLKL.
  • X comprises the amino acid sequence RLQLK(Ac).
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the SDM is an SDM comprising the structure of Formula I.
  • the selective delivery molecule of Formula I is: SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM- 110, SDM-1 1 1 , SDM- 1 12, SDM-1 13, SDM-114, SDM-115, SDM-116, SDM-1 17, SDM- 118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM- 129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-
  • the SDM is an SDM of Formula II, having the structure:
  • X is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids; c B and c M each independently comprise 0-1 amino acid;
  • M is a macromolecule
  • DB is a therapeutic agent
  • [CM-M] is bound to at any position of A or X, and [CB-D b ] is bound to any amino acid of B.
  • the therapeutic agent is cortisone.
  • a and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A.
  • A is a peptide comprising 5 or 9 consecutive glutamates.
  • B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines.
  • A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines.
  • CB and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid.
  • CB and c M are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a fi- amino acid, or a -amino acid.
  • CB and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxyl amine or hydrazine group.
  • CB and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CM is para-4-acetyl L-phenylalanine.
  • X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage.
  • X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALW S, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQL (Ac).
  • X comprises the amino acid sequence PLGLAG.
  • X comprises the amino acid sequence PLG-C(me)-AG.
  • X comprises the amino acid sequence RPLALWRS.
  • X comprises the amino acid sequence
  • X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac).
  • M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a PEG polymer, albumin, or a combination thereof. In some embodiments, M is a PEG.
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa),
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the SDM is an SDM comprising the structure of Formula II.
  • the selective delivery molecule of Formula II is: SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM- 110, SDM-1 1 1 , SDM- 1 12, SDM-1 13, SDM-114, SDM-115, SDM-116, SDM-1 17, SDM- 118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM- 129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141 , SDM-142, SDM-143, SDM-144, SDM-145, SDM- 146, SDM-147, SDM
  • the SDM does not comprise a cargo molecule.
  • an SDM without a cargo molecule is conjugated to an existing antibody-drug conjugate (e.g. Form 3, Fig. 3).
  • the SDM is an SDM of Formula III or IV, having the structure:
  • X is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids;
  • c M comprises 0-1 amino acid;
  • M is a macromolecule
  • a and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A.
  • A is a peptide comprising 5 or 9 consecutive glutamates.
  • B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines.
  • C M is selected from a naturally-occurring amino acid or a non- naturally-occurring amino acid. In some embodiments, C M is selected from a D amino acid, a L amino acid, an a-amino acid, a ⁇ -amino acid, or a r-amino acid. In some embodiments, C M is selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxyl amine or hydrazine group.
  • C M is selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • C M is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • C M is para-4-acetyl L-phenylalanine.
  • X is cleavable by a protease. In some embodiments, X is cleavable by a matrix metalloproteinase.
  • X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)- AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac). In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence RPLALWRS.
  • X comprises the amino acid sequence DPRSFL. In some embodiments, X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac).
  • M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a PEG polymer, albumin, or a combination thereof. In some embodiments, M is a PEG.
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)).
  • PEG 0.5kDa approximately 0.5kDa
  • PEG lkDa approximately 2kDa
  • PEG 3kDa approximately 4kDa
  • PEG 4kDa approximately 5kDa
  • PEG 5kDa approximately lOkDa
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the SDM is an SDM comprising the structure of Formula III or IV.
  • the selective delivery molecule of Formula III or IV is: SDM- 101, SDM- 102, SDM- 103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM- 110, SDM-111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM-118, SDM-119, SDM- 120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM- 137, SDM-138, SDM-139, SDM-140, SDM-141, SDM-142, SDM-143, SDM-101, SDM-
  • an SDM that comprises more than one protease cleavage site.
  • the SDM comprises a cleavage site for an
  • the SDM comprises a cleavable linker X and a cleavable linker Y, where linker X comprises a cleavage site for an extracellular protease and linker Y comprises a cleavage site for an intracellular protease.
  • linker X is located between portion of A and portion of B, and linker Y is located between portion of B and the therapeutic cargo.
  • the intracellular protease that cleaves the Y linker is a lysosomal protease.
  • the protease is one that is activated at low pH.
  • the protease is one that is activated at low pH and is expressed in an endosome.
  • the protease is a cathepsin.
  • the cathepsin is cathepsin B.
  • the Y linker additionally comprises a self-immolative cleavage site located between the intracellular protease cleavage site and the attached therapeutic cargo.
  • the self-immolative cleavage site is a PABC (p-aminobenzylcarbonyl) spacer and its analogs thereof.
  • the self- immolative cleavage site is a thiazole containing linker.
  • the SDM is an SDM of Formula V, having the structure:
  • X is a cleavable linker
  • Y is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids
  • C B and C M each independently comprise 0-1 amino acid
  • M is a macromolecule
  • D B is a therapeutic agent
  • [C M -M] is bound to at any position of A or X, and [C B -D b ] is bound to any amino acid of B.
  • a and B do not have an equal number of acidic and basic amino acids.
  • the number of basic amino acids in B is greater than the number of acidic amino acids in A.
  • A is a peptide comprising 5 or 9 consecutive glutamates.
  • B is a peptide comprising 8 or 9 consecutive arginines.
  • A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some
  • C B , and C M are each independently a 0-1 amino acid. In some embodiments, C B and C M are each independently selected from a naturally-occurring amino acid or a non-naturally- occurring amino acid. In some embodiments, C B and C M are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a ⁇ -amino acid, or a ⁇ -amino acid. In some
  • C B and C M are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • C B and C M are each independently selected from D-cysteine, D- glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • C B is any amino acid having a free thiol group.
  • C B is D-cysteine.
  • C M is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, C M is para-4-acetyl L-phenylalanine.
  • X is cleavable by a protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage.
  • X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)- AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac).
  • X comprises the amino acid sequence PLGLAG.
  • X comprises the amino acid sequence PLG-C(me)-AG.
  • X comprises the amino acid sequence RPLALWRS.
  • X comprises the amino acid sequence DPRSFL.
  • X comprises the amino acid sequence RLQLKL.
  • X comprises the amino acid sequence RLQLK(Ac).
  • M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a PEG polymer, albumin, or a combination thereof. In some embodiments, M is a PEG.
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa),
  • Y is cleavable by a protease. In some embodiments, Y is cleavable by an intracellular protease. In some embodiments, Y comprises an amino acid sequence that is cleavable by Cathepsin B. In some embodiments, Y comprises the amino acid sequence Phe-Lys or Val-Cit (L-citrulline). In some embodiments, Y comprises a site for self-immolative cleavage.
  • Y comprises a PABC self-immolative spacer.
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the SDM is an SDM comprising the structure of Formula V.
  • the selective delivery molecule of Formula V is: SDM-101, SDM-102, SDM-103, SDM-104, SDM-105, SDM- 106, SDM-107, SDM-108, SDM-109, SDM-110, SDM-1 11, SDM-112, SDM-113, SDM-1 14, SDM-1 15, SDM-116, SDM-117, SDM-118, SDM-1 19, SDM-120, SDM-121 , SDM-122, SDM- 123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM
  • the SDM is an SDM comprising an imaging agent. In certain embodiments, the SDM is an SDM comprising an imaging agent and a therapeutic agent. In certain embodiments, the SDM is an SDM comprising two or more imaging agents or two or more therapeutic agents.
  • the SDM is an SDM comprising two or more imaging agents for Forster resonance energy transfer (FRET) imaging, where one imaging agent is conjugated to the A portion of the SDM and one imaging agent is conjugated to the B portion of the SDM.
  • FRET Forster resonance energy transfer
  • the SDM is an SDM of Formula VI, having the structure:
  • X is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids
  • C A , C B , and C M each independently comprise 0-1 amino acid
  • M is a macromolecule
  • D A and D B are each independently selected from an imaging agent and a therapeutic; and wherein [C M -M] is bound to at any position of A or X, [D A -C A ] is bound to any amino acid of A, and [C B -D b ] is bound to any amino acid of B.
  • a and B do not have an equal number of acidic and basic amino acids.
  • the number of basic amino acids in B is greater than the number of acidic amino acids in A.
  • A is a peptide comprising 5 or 9 consecutive glutamates.
  • B is a peptide comprising 8 or 9 consecutive arginines.
  • A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines.
  • C A , C B , and C M are each independently a 0-1 amino acid. In some embodiments, C A , C B , and C M are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, C A , C B , and C M are each
  • C A , C B , and C M are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • C A , C B , and C M are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • CB is any amino acid having a free thiol group. In some embodiments, CB is D- cysteine. In some embodiments, CA is any amino acid having a -terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CA is lysine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L- phenylalanine. In some embodiments, X is cleavable by a protease.
  • X is cleavable by a matrix metalloproteinase.
  • X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14.
  • X comprises a peptide linkage.
  • X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac).
  • X comprises the amino acid sequence PLGLAG.
  • X comprises the amino acid sequence PLG-C(me)-AG.
  • X comprises the amino acid sequence RPLALWRS. In some embodiments, X comprises the amino acid sequence DPRSFL. In some embodiments, X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac).
  • M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a PEG polymer, albumin, or a combination thereof. In some embodiments, M is a PEG.
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa),
  • DA and DB are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other.
  • DA and D B are Cy5 and Cy7.
  • DA and D B are Cy5 and IRDye750.
  • DA and DB are Cy5 and IRDye800.
  • DA and D B are Cy5 and ICG.
  • DA and D B are a fluorescent moiety and a fluorescence-quenching moiety.
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the SDM is an SDM comprising the structure of Formula VI.
  • the molecule of Formula I is: SDM- 14, SDM- 15, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-32, or SDM-35.
  • the SDM is an SDM of Formula VI, having the structure:
  • X is a cleavable linker
  • A is a peptide with a sequence comprising 5 to 9 acidic amino acids
  • B is a peptide with a sequence comprising 7 to 9 basic amino acids
  • CA, C b , and c M each independently comprise 0-1 amino acid
  • M is a polyethylene glycol (PEG) polymer
  • DA and D B are each independently an imaging agent
  • [CM -M] is bound to at any position of A or X
  • [DA-CA] is bound to any amino acid of A
  • [c B -D B ] is bound to any amino acid of B.
  • a and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A.
  • A is a peptide comprising 5 or 9 consecutive glutamates.
  • B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines.
  • A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines.
  • CA, C b , and CM are each independently a 0-1 amino acid.
  • CA, C b , and c M are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid.
  • CA, C b , and CM are each
  • CA, CB, and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CA, CB, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • CB is any amino acid having a free thiol group. In some embodiments, CB is D- cysteine. In some embodiments, CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CA is lysine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L- phenylalanine. In some embodiments, X is cleavable by a protease.
  • X is cleavable by a matrix metalloproteinase.
  • X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14.
  • X comprises a peptide linkage.
  • X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQL L, and RLQLK(Ac).
  • X comprises the amino acid sequence PLGLAG.
  • X comprises the amino acid sequence PLG-C(me)-AG.
  • X comprises the amino acid sequence RPLALWRS. In some embodiments, X comprises the amino acid sequence DPRSFL. In some embodiments, X comprises the amino acid sequence PPRSFL. In some embodiments, X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac).
  • DA and DB are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other. In some embodiments, DA and DB are Cy5 and Cy7. In some embodiments, DA and D B are Cy5 and IRDye750. In some embodiments, DA and D B are Cy5 and IRDye800.
  • D A and D B are Cy5 and ICG.
  • D A and DB are a fluorescent moiety and a fluorescence-quenching moiety.
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo.
  • the SDM is an SDM comprising the structure of Formula VI.
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), and approximately 40kDa (PEG 40kDa)).
  • the molecule of Formula VI is: SDM-14, SDM-15, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-32; or SDM-35.
  • the SDM is an SDM of Formula VI, having the structure:
  • X is a peptide linker cleavable by a matrix metalloproteinase
  • A is a peptide with a sequence comprising 5 or 9 consecutive glutamates
  • B is a peptide with a sequence comprising 8 or 9 consecutive arginines
  • CA, CB, and c M each independently comprise 0-1 amino acid
  • M is a polyethylene glycol (PEG) polymer
  • DA and DB are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other;
  • [CM -M] is bound to at any position of A or X
  • [DA-CA] is bound to any amino acid of A
  • [CB -DB] is bound to any amino acid of B.
  • a and B do not have an equal number of acidic and basic amino acids.
  • CA, C b , and c M are each
  • CA, C b , and CM are each independently a 0-1 amino acid.
  • CA, CB, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L- phenylalanine.
  • CB is any amino acid having a free thiol group.
  • CB is D-cysteine.
  • CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, D A and D B are Cy5 and Cy7. In some embodiments, DA and D B are Cy5 and Cy7.
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)).
  • the SDM is an SDM comprising the structure of Formula VI.
  • the SDM is an SDM of Formula VI, having the structure:
  • X is a peptide linker cleavable by a matrix metalloproteinase
  • A is a peptide with a sequence comprising 5 consecutive glutamates
  • B is a peptide with a sequence comprising 8 consecutive arginines
  • CA, C b , and c M each independently comprise 0-1 amino acid
  • M is a polyethylene glycol (PEG) polymer
  • DA and DB are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other;
  • CA, C b , and c M are each independently a 0-1 amino acid.
  • CA, c B , and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CA, C b , and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • c B is any amino acid having a free thiol group.
  • c B is D- cysteine.
  • CA is any amino acid having a N-terminal amine group.
  • CA is D-glutamate.
  • CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CM is para-4-acetyl L-phenylalanine.
  • X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence
  • DA and D B are Cy5 and Cy7. In some embodiments, DA and D B are Cy5 and IRDye750. In some embodiments, DA and D B are Cy5 and IRDye800. In some embodiments, DA and D B are Cy5 and ICG.
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)).
  • PEG 0.5kDa approximately IkDa
  • PEG 2kDa approximately 2kDa
  • PEG 3kDa approximately 4kDa
  • PEG 4kDa approximately 5kDa
  • PEG 5kDa approximately lOkDa
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the SDM is an SDM comprising the structure of Formula VI.
  • the SDM is an SDM of Formula VI, having the structure:
  • X is a peptide linker cleavable by a matrix metalloproteinase
  • A is a peptide with a sequence comprising 9 consecutive glutamates
  • B is a peptide with a sequence comprising 9 consecutive arginines
  • CA, CB, and CM each independently comprise 0-1 amino acid
  • M is a polyethylene glycol (PEG) polymer
  • DA and D B are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other;
  • CA, C b , and c M are each independently a 0-1 amino acid.
  • CA, CB, and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CA, CB, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • CB is any amino acid having a free thiol group.
  • CB is D- cysteine.
  • CA is any amino acid having a N-terminal amine group.
  • CA is D-glutamate.
  • CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CM is para-4-acetyl L-phenylalanine.
  • X comprises the amino acid sequence PLGLAG.
  • X comprises the amino acid sequence PLG-C(me)-AG.
  • X comprises the amino acid sequence
  • DA and D B are Cy5 and Cy7.
  • DA and DB are Cy5 and IRDye750.
  • DA and DB are Cy5 and IRDye800.
  • DA and DB are Cy5 and ICG.
  • M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa),
  • A comprises a thiol reactive group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • the SDM is an SDM comprising the structure of Formula VI.
  • the SDM comprises a structure selected from: SDM-1 , SDM-2, SDM-3, SDM-4, SDM-5, SDM-6, SDM-7, SDM-8, SDM-9, SDM-10, SDM-11, SDM-12, SDM- 13, SDM-14, SDM-15, SDM-16, SDM-17, SDM-18, SDM-19, SDM-20, SDM-21 , SDM-22, SDM- 23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-28, SDM-29, SDM-30, SDM-31 , SDM-32, SDM- 33, SDM-34, SDM-35, SDM-36, SDM-37, SDM-38, SDM-39, and SDM-40 (see International PCT Pub.
  • the selective delivery molecule comprises a structure selected from: SDM-14, SDM-15, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-32, or SDM-35.
  • the selective delivery molecule comprises Peptide P-3 (see International PCT Pub. No. WO2013/019681).
  • A is a peptide with a sequence comprising 2 to 20 acidic amino acids. In some embodiments, peptide portion of A comprises between about 2 to about 20 acidic amino acids. In some embodiments, peptide portion of A comprises between about 5 to about 20 acidic amino acids. In some embodiments, A has a sequence comprising 5 to 9 acidic amino acids. In some embodiments, A has a sequence comprising 5 to 8 acidic amino acids. In some
  • A has a sequence comprising 5 to 7 acidic amino acids. In some embodiments, A has a sequence comprising 5 acidic amino acids. In some embodiments, A has a sequence comprising 6 acidic amino acids. In some embodiments, A has a sequence comprising 7 acidic amino acids. In some embodiments, A has a sequence comprising 8 acidic amino acids. In some embodiments, A has a sequence comprising 9 acidic amino acids.
  • peptide portion of A comprises between about 2 to about 20 consecutive acidic amino acids. In some embodiments, peptide portion of A comprises between about 5 to about 20 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 5 to 9 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 5 to 8 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 5 to 7 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 5 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 6 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 7 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 8 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 9 consecutive acidic amino acids.
  • peptide portion of A comprises between about 2 to about 20 acidic amino acids selected from, aspartates and glutamates. In some embodiments, peptide portion of A comprises between about 5 to about 20 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 9 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 8 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 7 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 acidic amino acids selected from, aspartates and glutamates.
  • A has a sequence comprising 6 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 7 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 8 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 9 acidic amino acids selected from, aspartates and glutamates.
  • peptide portion of A comprises between about 2 to about 20 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, peptide portion of A comprises between about 5 to about 20 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 9 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 8 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 7 consecutive acidic amino acids selected from, aspartates and glutamates.
  • A has a sequence comprising 5 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 6 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 7 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 8 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 9 consecutive acidic amino acids selected from, aspartates and glutamates.
  • peptide portion of A comprises between about 2 to about 20 glutamates. In some embodiments, peptide portion of A comprises between about 5 to about 20 glutamates. In some embodiments, A has a sequence comprising 5 to 9 glutamates. In some embodiments, A has a sequence comprising 5 to 8 glutamates. In some embodiments, A has a sequence comprising 5 to 7 glutamates. In some embodiments, A has a sequence comprising 5 glutamates. In some embodiments, A has a sequence comprising 6 glutamates. In some
  • A has a sequence comprising 7 glutamates. In some embodiments, A has a sequence comprising 8 glutamates. In some embodiments, A has a sequence comprising 9 glutamates.
  • peptide portion of A comprises between about 2 to about 20 consecutive glutamates. In some embodiments, peptide portion of A comprises between about 5 to about 20 consecutive glutamates. In some embodiments, A has a sequence comprising 5 to 9 consecutive glutamates. In some embodiments, A has a sequence comprising 5 to 8 consecutive glutamates. In some embodiments, A has a sequence comprising 5 to 7 consecutive glutamates. In some embodiments, A has a sequence comprising 5 consecutive glutamates. In some embodiments, A has a sequence comprising 6 consecutive glutamates. In some embodiments, A has a sequence comprising 7 consecutive glutamates. In some embodiments, A has a sequence comprising 8 consecutive glutamates. In some embodiments, A has a sequence comprising 9 consecutive glutamates.
  • portion of A comprises 5 consecutive glutamates (i.e., EEEEE or eeeee). In some embodiments, portion of A comprises 9 consecutive glutamates (i.e., EEEEEEEEE or eeeeeeee).
  • An acidic portion of A may include amino acids that are not acidic. Acidic portion of A may comprise other moieties, such as negatively charged moieties. In embodiments of a selective delivery molecule disclosed herein, an acidic portion of A may be a negatively charged portion, preferably having about 2 to about 20 negative charges at physiological pH that does not include an amino acid. [00106] In some embodiments, the amount of negative charge in portion of A is approximately the same as the amount of positive charge in portion of B. In some embodiments, the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B.
  • improved tissue uptake is seen in a selective delivery molecule wherein the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B. In some embodiments, improved solubility is observed in a selective delivery molecule wherein the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B. In some embodiments, faster tissue uptake is seen in a selective delivery molecule wherein the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B. In some embodiments, greater tissue uptake is seen in a selective delivery molecule wherein the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B.
  • Portion of A is either L-amino acids or D-amino acids.
  • D-amino acids are preferred in order to minimize immunogenicity and nonspecific cleavage by background peptidases or proteases.
  • Cellular uptake of oligo-D-arginine sequences is known to be as good as or better than that of oligo-L-arginines.
  • portion of A comprises a thiol reactive group.
  • the thiol group is an N-terminal thiol group.
  • the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents.
  • the thiol reactive group covalently binds to a carrier protein.
  • the carrier protein is albumin.
  • the thiol reactive group covalently binds to Cysteine 34 of albumin.
  • the thiol reactive group covalently binds to albumin in vivo.
  • portion of A may include non-standard amino acids, such as, for example, hydroxylysine, desmosine, isodesmosine, or other non-standard amino acids.
  • Portion of A may include modified amino acids, including post-translationally modified amino acids such as, for example, methylated amino acids (e.g., methyl histidine, methylated forms of lysine, etc.), acetylated Amino acids, amidated amino acids, formylated amino acids, hydroxylated amino acids, phosphorylated amino acids, or other modified amino acids.
  • Portion of A may also include peptide mimetic moieties, including portions linked by non-peptide bonds and amino acids linked by or to non-amino acid portions.
  • B is a peptide with a sequence comprising 5 to 15 basic amino acids. In some embodiments, peptide portion of B comprises between about 5 to about 20 basic amino acids. In some embodiments, peptide portion of B comprises between about 5 to about 12 basic amino acids. In some embodiments, peptide portion of B comprises between about 7 to about 9 basic amino acids. In some embodiments, peptide portion of B comprises between about 7 to about 8 basic amino acids. In some embodiments, peptide portion of B comprises 9 basic amino acids. In some embodiments, peptide portion of B comprises 8 basic amino acids. In some embodiments, peptide portion of B comprises 7 basic amino acids.
  • peptide portion of B comprises between about 5 to about 20 consecutive basic amino acids. In some embodiments, peptide portion of B comprises between about 5 to about 12 consecutive basic amino acids. In some embodiments, peptide portion of B comprises between about 7 to about 9 consecutive basic amino acids. In some embodiments, peptide portion of B comprises between about 7 to about 8 consecutive basic amino acids. In some embodiments, peptide portion of B comprises 9 consecutive basic amino acids. In some embodiments, peptide portion of B comprises 8 consecutive basic amino acids. In some embodiments, peptide portion of B comprises 7 consecutive basic amino acids.
  • peptide portion of B comprises between about 5 to about 20 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 5 to about 12 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 7 to about 9 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 7 to about 8 basic amino acids selected from arginines, histidines, and lysines.
  • peptide portion of B comprises 9 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises 8 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises 7 basic amino acids selected from arginines, histidines, and lysines.
  • peptide portion of B comprises between about 5 to about 20 consecutive basic amino acids selected from arginines, histidines, and lysines.
  • peptide portion of B comprises between about 5 to about 12 consecutive basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 7 to about 9 consecutive basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 7 to about 8 consecutive basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises 9 consecutive basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises 8 consecutive basic amino acids selected from arginines, histidines, and lysines. In some
  • peptide portion of B comprises between about 5 to about 20 arginines. In some embodiments, peptide portion of B comprises between about 5 to about 12 arginines. In some embodiments, peptide portion of B comprises between about 7 to about 9 arginines. In some embodiments, peptide portion of B comprises between about 7 to about 8 arginines. In some embodiments, peptide portion of B comprises 9 arginines. In some
  • peptide portion of B comprises 8 arginines. In some embodiments, peptide portion of B comprises 7 arginines.
  • peptide portion of B comprises between about 5 to about 20 consecutive arginines. In some embodiments, peptide portion of B comprises between about 5 to about 12 consecutive arginines. In some embodiments, peptide portion of B comprises between about 7 to about 9 consecutive arginines. In some embodiments, peptide portion of B comprises between about 7 to about 8 consecutive arginines. In some embodiments, peptide portion of B comprises 9 consecutive arginines. In some embodiments, peptide portion of B comprises 8 consecutive arginines. In some embodiments, peptide portion of B comprises 7 consecutive arginines.
  • a basic portion of B may include amino acids that are not basic.
  • Basic portion of B may comprise other moieties, such as positively charged moieties.
  • a basic portion of B may be a positively charged portion, preferably having between about 5 and about 20 positive charges at physiological pH, that does not include an amino acid.
  • the amount of negative charge in portion of A is approximately the same as the amount of positive charge in portion of B. In some embodiments, the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B.
  • Portion of B is either L-amino acids or D-amino acids.
  • D-amino acids are preferred in order to minimize immunogenicity and nonspecific cleavage by background peptidases or proteases.
  • Cellular uptake of oligo-D-arginine sequences is known to be as good as or better than that of oligo-L-arginines.
  • portion of B may include non-standard amino acids, such as, for example, hydroxylysine, desmosine, isodesmosine, or other non-standard amino acids.
  • Portion of B may include modified amino acids, including post-translationally modified amino acids such as, for example, methylated amino acids (e.g., methyl histidine, methylated forms of lysine, etc.), acetylated amino acids, amidated amino acids, formylated amino acids, hydroxylated amino acids, phosphorylated amino acids, or other modified amino acids.
  • Portion of B may also include peptide mimetic moieties, including portions linked by non-peptide bonds and amino acids linked by or to non-amino acid portions.
  • X is a peptide cleavable by a protease
  • the cargo e.g., D A and D B
  • the macromolecule carriers (M) are attached indirectly to A-X-B.
  • the cargo (e.g., D A and D B ) and the macromolecule carriers (M) are attached indirectly to A-X-B by a conjugation group (C A , C B , and C M ).
  • the cargo (e.g., D A and D B ) and the macromolecule carriers (M) are attached indirectly to A-X-B by a reactive conjugation group (C A , C B , and C M ).
  • the cargo (e.g., D A and D B ) and the macromolecule carriers (M) are attached indirectly to A-X-B by an orthogonally reactive conjugation group (C A , C b , and c M ).
  • C A , C b , and c M each independently comprise an amino acid. In some embodiments, C A , C B , and C M each independently comprise 0-10 amino acids. In some embodiments, C A , C B , and C M each independently comprise 1 amino acid. In some embodiments, C A , C b , and c M each independently comprise 2 amino acids. In some embodiments, C A , C b , and c M each independently comprise 3 amino acids. In some embodiments, CA, CB, and CM each independently comprise 4 amino acids. In some embodiments, CA, CB, and CM each independently comprise 5 amino acids. In some embodiments, C A , C B , and C M each independently comprise an amino acid. In some embodiments, C A , C B , and C M each independently comprise 0-10 amino acids. In some embodiments, C A , C B , and C M each independently comprise 1 amino acid. In some embodiments, C A , C b , and c M each independently comprise 2 amino acids.
  • C A , C b , and c M each independently comprise 7 amino acids. In some embodiments, C A , C B , and C M each independently comprise 8 amino acids. In some embodiments, C A , C B , and C M each independently comprise 9 amino acids. In some embodiments, C A , C B , and C M each independently comprise 10 amino acids.
  • C A , C B , and C M each independently comprise a derivatized amino acid.
  • multiple cargos (D) are attached to a derivatized amino acid conjugation group.
  • the conjugation group comprises a receptor ligand.
  • C A , C B , and C M each independently comprise a naturally-occurring amino acid or a non-naturally-occurring amino acid.
  • C A , C B , and C M each independently comprise from a D amino acid, a L amino acid, an a-amino acid, a ⁇ -amino acid, or a T-amino acid.
  • C A , C B , and C M each independently comprise any amino acid having a free thiol group, any amino acid containing a free amine group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CA, CB, and CM each independently comprise D-cysteine, D-glutamate, lysine, and para-4-acetyl L- phenylalanine.
  • CB comprises any amino acid having a free thiol group.
  • CB comprises D-cysteine.
  • CA comprises any amino acid having a N-terminal amine group. In some embodiments, CA comprises D-glutamate. In some embodiments, CA comprises lysine. In some embodiments, CM comprises any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM comprises para-4-acetyl L-phenylalanine.
  • CA, C b , and c M are each independently selected from a naturally- occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CA, CB, and c M are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a ⁇ - amino acid, or a ⁇ -amino acid.
  • CA, C b , and c M are each independently any amino acid having a free thiol group, any amino acid containing a free amine group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CA, CB, and CM are each independently selected from: D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • CB is any amino acid having a free thiol group.
  • c B is D-cysteine.
  • CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CA is lysine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine.
  • a selective delivery molecule disclosed herein for delivering a therapeutic agent to a tissue or a plurality of cells.
  • the therapeutic agent is an anti-inflammatory agent.
  • the therapeutic agent is an anti-cancer agent.
  • the selective delivery molecule is used to treat colorectal cancer.
  • a D moiety is independently a therapeutic agent.
  • a D moiety comprises two or more therapeutic agents.
  • the two or more therapeutic agents are the same therapeutic agent.
  • the two or more therapeutic agents are different therapeutic agents.
  • a D moiety comprises 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 or more therapeutic agents.
  • the therapeutic agent is selected from: a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, an anti-inflammatory agent, or a combination thereof.
  • the therapeutic agent is a radiotherapeutic agent.
  • the therapeutic agent is a cytotoxin.
  • the therapeutic agent is a B cell receptor pathway inhibitor.
  • the therapeutic agent is a CD79A inhibitor, a CD79B inhibitor, a CD 19 inhibitor, a Lyn inhibitor, a Syk inhibitor, a PI3K inhibitor, a Blnk inhibitor, a PLGy inhibitor, a PKCP inhibitor, or a combination thereof.
  • the therapeutic agent is an antibody, B cell receptor signaling inhibitor, a PI3K inhibitor, an IAP inhibitor, an mTOR inhibitor, a radioimmunotherapeutic, a DNA damaging agent, a proteosome inhibitor, a histone deacytlase inhibitor, a protein kinase inhibitor, a hedgehog inhibitor, an Hsp90 inhibitor, a telomerase inhibitor, a Jakl/2 inhibitor, a protease inhibitor, a PKC inhibitor, a PARP inhibitor, or a combination thereof.
  • the therapeutic agent is selected from: chlorambucil, ifosphamide, doxorubicin, mesalazine, thalidomide, lenalidomide, temsirolimus, everolimus, fludarabine, fostamatinib, paclitaxel, docetaxel, ofatumumab, rituximab, dexamethasone, prednisone, CAL-101 , ibritumomab, tositumomab, bortezomib, pentostatin, endostatin,
  • afelimomab certolizumab pegol, etanercept, golimumab, infliximab, anakinra, basiliximab, canakinumab, daclizumab, mepolizumab, rilonacept, tocilizumab, ustekinumab, ciclosporin, tacrolimus, azathioprine, lenalidomide, methotrexate, thalidomide, adalimumab, alemtuzumab, bevacizumab, cetuximab, certolizumab pegol, , eculizumab, efalizumab, gemtuzumab, ibritumomab tiuxetan, muromonab-CD3, natalizumab, panitumumab, ranibizumab, rituximab, tositumomab, trastuzumab, cat
  • ramacurimab ranibizumab, siplizumab, sonepcizumab, tanezumab, tositumomab, trastuzumab, tremelimumab, tucotuzumab celmoleukin, veltuzumab, visilizumab, volociximab, zalutumumab, a syk inhibitor (e.g., R788), enzastaurin, dasatinib, erlotinib, everolimus, gefitinib, imatinib, lapatinib, nilotinib, pazonanib, sorafenib, sunitinib, temsirolimus, an angiogenesis inhibitor (e.g., GT-111, JI-101, R1530), a kinase inhibitors (e.g., AC220, AC480, ACE-041, AMG 900
  • interleukin II interleukin II, or rlL2
  • interferon alfa-2a interferon alfa -2b
  • interferon alfa -nl interferon alfa -n3, interferon beta-1 a, interferon gamma-1 b
  • iproplatin irinotecan hydrochloride, lanreotide acetate, letrozole, leuprolide acetate, liarozole hydrochloride, lometrexol sodium, lomustine, losoxantrone hydrochloride, masoprocol
  • hydrochloride puromycin, puromycin hydrochloride, pyrazofurin, riboprine, rogletimide, safingol, safingol hydrochloride, semustine, pumprazene, sparfosate sodium, sparsomycin, spirogermanium hydrochloride, spiromustine, spiroplatin, streptonigrin, streptozocin, sulofenur, talisomycin, tecogalan sodium, tegafur, teloxantrone hydrochloride, temoporfin, teniposide, teroxirone, testolactone, thiamiprine, thioguanine, thiotepa, tiazofurin, tirapazamine, toremifene citrate, trestolone acetate, triciribine phosphate, trimetrexate, trimetrexate glucuronate, triptorelin, tubulozole
  • the therapeutic agent is selected from: 20-epi-l, 25 dihydroxyvitamin D3, 5-ethynyluracil, abiraterone, aclarubicin, acylfulvene, adecypenol, adozelesin, aldesleukin, ALL-TK antagonists, altretamine, ambamustine, amidox, amifostine, aminolevulinic acid, amrubicin, amsacrine, anagrelide, anastrozole, andrographolide, angiogenesis inhibitors, antagonist D, antagonist G, antarelix, anti-dorsalizing morphogenetic protein- 1, antiandrogen, prostatic carcinoma, antiestrogen, antineoplaston, antisense oligonucleotides, aphidicolin glycinate, apoptosis gene modulators, apoptosis regulators, apurinic acid, ara-CDP-DL-PTBA, arg
  • combretastatin analogue conagenin, crambescidin 816, crisnatol, cryptophycin 8, cryptophycin A derivatives, curacin A, cyclopentanthraquinones, cycloplatam, cypemycin, cytarabine ocfosfate, cytolytic factor, cytostatin, dacliximab, decitabine, dehydrodidemnin B, deslorelin, dexamethasone, dexifosfamide, dexrazoxane, dexverapamil, diaziquone, didemnin B, didox, diethylnorspermine, dihydro-5-azacytidine, 9- dioxamycin, diphenyl spiromustine, docosanol, dolasetron, doxifiuridine, droloxifene, dronabinol, duocarmycin SA, ebsele
  • fluorodaunorunicin hydrochloride forfenimex, formestane, fostriecin, fotemustine, gadolinium texaphyrin, gallium nitrate, galocitabine, ganirelix, gelatinase inhibitors, gemcitabine, glutathione inhibitors, hepsulfam, heregulin, hexamethylene bisacetamide, hypericin, ibandronic acid, idarubicin, idoxifene, idramantone, ilmofosine, ilomastat, imidazoacridones, imiquimod, immunostimulant peptides, insulin-such as for example growth factor- 1 receptor inhibitor, interferon agonists, interferons, interleukins, iobenguane, iododoxorubicin, ipomeanol, 4-, iroplact, irsogladine, isobengazole, iso
  • oligonucleotides onapristone, ondansetron, ondansetron, oracin, oral cytokine inducer, ormaplatin, osaterone, oxaliplatin, oxaunomycin, palauamine, palmitoylrhizoxin, pamidronic acid, panaxytriol, panomifene, parabactin, pazelliptine, pegaspargase, peldesine, pentosan polysulfate sodium, pentostatin, pentrozole, perflubron, perfosfamide, perillyl alcohol, phenazinomycin, phenylacetate, phosphatase inhibitors, picibanil, pilocarpine hydrochloride, pirarubicin, piritrexim, placetin A, placetin B, plasminogen activator inhibitor, platinum complex, platinum compounds, platinum- triamine complex, porfimer sodium, porfiromycin, prednisone, prop
  • mercaptopurine thioguanine, pentostatin, mechloroethamine, cyclophosphamide, chlorambucil, meiphalan, ethylenimine, methylmelamine, hexamethlymelamine, thiotepa, busulfan, carmustine, lomusitne, semustine, streptozocin, decarbazine, fluorouracil, floxouridine, cytarabine,
  • mercaptopurine thioguanine
  • pentostatin erbulozole
  • Dolastatin 10 also known as DLS-10 and NSC-376128
  • Mivobulin isethionate also known as CI-980
  • Vincristine NSC-639829
  • Discodermolide also known as NVP-XX-A-296
  • ABT-751 Abbott, also known as E-7010
  • Altorhyrtins such as Altorhyrtin A and Altorhyrtin C
  • Spongistatins such as
  • the therapeutic agent is an anti-inflammatory agent.
  • the therapeutic agent is an anti-TNF agent, an IL-1 receptor antagonist, an IL-2 receptor antagonist, a cytotoxic agent, an immunomodulatory agent, an antibiotic, a T-cell co- stimulatory blocker, a B cell depleting agent, an immunosuppressive agent, an alkylating agent, an anti-metabolite, a plant alkaloid, a terpenoids, a topoisomerase inhibitor, an antitumour antibiotic, an antibody, a hormonal therapy, an anti-diabetes agent, a leukotriene inhibitor, or combinations thereof.
  • the therapeutic agent is selected from: alefacept, efalizumab, methotrexate, acitretin, isotretinoin, hydroxyurea, mycophenolate mofetil, sulfasalazine, 6- Thioguanine, Dovonex, Taclonex, betamethasone, tazarotene, hydroxychloroquine, etanercept, adalimumab, infliximab, abatacept, rituximab, tratuzumab, Anti-CD45 monoclonal antibody AHN- 12 (NCI), Iodine-131 Anti-Bl Antibody (Corixa Corp.), anti-CD66 monoclonal antibody BW 250/183 (NCI, Victoria General Hospital), anti-CD45 monoclonal antibody (NCI, Baylor College of Medicine), antibody anti-anb3 integrin (NCI), BIW-8962 (BioWa Inc.), Antibody BC8 (
  • etoposide etoposide phosphate
  • teniposide dactinomycin
  • doxorubicin daunorubicin
  • valrubicine idarubicine
  • epirubicin bleomycin
  • plicamycin mitomycin
  • finasteride goserelin
  • the therapeutic agent contains a radioactive moiety, for example a radioactive isotope such as 211 At, 1 1 I, 125 I, 90 Y, 186 Re, 188 Re, 153 Sm, 212 Bi, 32 P, 64 Cu radioactive isotopes of Lu, and others.
  • a radioactive isotope such as 211 At, 1 1 I, 125 I, 90 Y, 186 Re, 188 Re, 153 Sm, 212 Bi, 32 P, 64 Cu radioactive isotopes of Lu, and others.
  • Polymers are characterized by a distribution of molecular weights, and, as such, the molecular weight, presented herein for polymers, is only an approximate average molecular weight of a distribution of molecular weights of individual polymers. Unless stated otherwise, the molecular weight of a polymeric component will have a typical (i.e., as known in the art) error and standard deviation.
  • a carrier modulates plasma half-life of a selective delivery molecule disclosed herein. In some embodiments, a carrier modulates solubility of a selective delivery molecule disclosed herein. In some embodiments, a carrier modulates bio-distribution of a selective delivery molecule disclosed herein.
  • a carrier decreases uptake of a selective delivery molecule by non- target cells or tissues. In some embodiments, a carrier decreases uptake of a selective delivery molecule into cartilage. In some embodiments, a carrier decreases uptake of a selective delivery molecule into joints relative to target tissue.
  • a carrier increases uptake of a selective delivery molecule by target cells or tissues. In some embodiments, a carrier decreases uptake of a selective delivery molecule into the liver relative to target tissue. In some embodiments, a carrier decreases uptake of a selective delivery molecule into kidneys. In some embodiments, a carrier enhances uptake into cancer tissue. In some embodiments, a carrier enhances uptake into lymphatic channels and/or lymph nodes.
  • a carrier increases plasma half-life by reducing glomerular filtration. In some embodiments, a carrier modulates plasma half-life by increasing or decreases metabolism or protease degradation. In some embodiments, a carrier increases tumor uptake due to enhanced permeability and retention (EPR) of tumor vasculature. In some embodiments, a carrier increases the aqueous solubility of selective delivery molecule.
  • EPR enhanced permeability and retention
  • any M is independently directly or indirectly (e.g., via CM) bound to A, B, or X.
  • any M is independently bound to A at the n-terminal poly glutamate.
  • any M is independently bound to A (or, the n-terminal poly glutamate) by a covalent linkage.
  • any M is independently bound to B at the c-terminal polyarginine.
  • any M is independently bound to B (or, the c- terminal polyarginine) by a covalent linkage.
  • any M is independently directly or indirectly bound to linkers between X and A, X and B, B and C N terminus, and A and C/N terminus.
  • the covalent linkage comprises an ether bond, thioether bond, amine bond, amide bond, oxime bond, carbon-carbon bond, carbon-nitrogen bond, carbon-oxygen bond, or carbon-sulfur bond.
  • M is selected from a protein, a synthetic or natural polymer, or a dendrimer.
  • M is selected from dextran, a PEG polymer (e.g., a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (
  • M is a PEG polymer.
  • the size of M is between about 50kDa and about 70kDa.
  • the selective delivery molecule is conjugated to albumin.
  • albumin is excluded from the glomerular filtrate under normal physiological conditions.
  • the selective delivery molecule comprises a reactive group such as maleimide that can form a covalent conjugate with albumin.
  • a selective delivery molecule comprising albumin results in enhanced accumulation of cleaved selective delivery molecules in tumors in a cleavage dependent manner.
  • albumin conjugates have good pharmacokinetic properties.
  • the selective delivery molecule is conjugated to PEG polymers.
  • the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa).
  • the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately lkDa (PEG lkDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 2kDa (PEG 2kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 3kDa (PEG 3kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 4kDa (PEG 4kDa).
  • the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 5kDa (PEG 5kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately lOkDa (PEG lOkDa). In some embodiments, the selective delivery molecule is conjugated PEG polymers having an average molecular weight of approximately 12 kDa ( PEG 12kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 15kDa (PEG 15kDa).
  • selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 20 kDa (PEG 20kDa). In some embodiments, selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 30 kDa (PEG 30kDa). In some embodiments, selective delivery molecules conjugated to PEG30kDa had a longer half-life as compared to free peptides. In some
  • selective delivery molecules are conjugated to PEG polymers having an average molecular weight of between about 20 to about 40kDa which have hepatic and renal clearance.
  • the PEG groups are polydisperse and have a distribution of molecular weights. Thus, any characterization of a PEG group should be interpreted in light of the polydispersity of PEG, unless otherwise stated.
  • the selective delivery molecule is conjugated to a dextran.
  • the selective delivery molecule is conjugated to a dextran having a molecular weight of approximately 70kDa.
  • dextran conjugates being a mixture of molecular weights, are difficult to synthesize and purify reproducibly.
  • the selective delivery molecule is conjugated to streptavidin.
  • the selective delivery molecule is conjugated to a fifth generation PAMAM dendrimer.
  • a carrier is capped.
  • capping a carrier improves the pharmacokinetics and reduces cytotoxicity of a carrier by adding hydrophilicity.
  • the cap is selected from: Acetyl, succinyl, 3-hydroxypropionyl, 2-sulfobenzoyl, glycidyl, PEG-2, PEG-4, PEG-8 and PEG-12.
  • X is a linker consisting of one or more amino acids is used to join peptide sequence A (i.e., the sequence designed to inhibit the delivery action of peptide B) and peptide sequence B.
  • peptide linker will have no specific biological activity other than to join the molecules or to preserve some minimum distance or other spatial relationship between them.
  • the constituent amino acids of the linker may be selected to influence some property of the molecule such as the folding, net charge, or hydrophobicity.
  • an intact selective delivery molecule disclosed herein may not be able to enter the cell because of the presence of portion of A.
  • a strictly intracellular process for cleaving X would be ineffective to cleave X in healthy cells since portion of A, preventing uptake into cells, would not be effectively cleaved by intracellular enzymes in healthy cells since it would not be taken up and would not gain access to such intracellular enzymes.
  • a cell is injured or diseased (e.g., cancerous cells, hypoxic cells, ischemic cells, apoptotic cells, necrotic cells) such intracellular enzymes leak out of the cell and cleavage of A would occur, allowing entry of portion of B and/or cargo into the cell, effecting targeted delivery of portion of B and/or cargo D to neighboring cells.
  • X is cleaved in the extracellular space.
  • the fact that capillaries are often leaky around tumors and other trauma sites enhances the ability of high molecular weight molecules (e.g., molecular weight of about 30 kDa or more) to reach the interstitial compartment.
  • cells that do not express the relevant protease but that are immediately adjacent to expressing cells pick up cargo from a selective delivery molecule because linkage of a X linker is typically extracellular.
  • such bystander targeting is beneficial in the treatment of tumors because of the heterogeneity of cell phenotypes and the wish to eliminate as high a percentage of suspicious cells as possible.
  • X is a cleavable linker
  • the X linker is flexible. In some embodiments, the linker is rigid.
  • the X linker comprises a linear structure. In some embodiments, the X linker comprises a non-linear structure. In some embodiments, the X linker comprises a branched structure. In some embodiments, the X linker comprises a cyclic structure. [00153] In some embodiments, X is about 5 to about 30 atoms in length. In some embodiments, X is about 6 atoms in length. In some embodiments, X is about 8 atoms in length. In some
  • X is about 10 atoms in length. In some embodiments, X is about 12 atoms in length. In some embodiments, X is about 14 atoms in length. In some embodiments, X is about 16 atoms in length. In some embodiments, X is about 18 atoms in length. In some embodiments, X is about 20 atoms in length. In some embodiments, X is about 25 atoms in length. In some embodiments, X is about 30 atoms in length.
  • the linker binds peptide portion of A (i.e., the peptide sequence which prevents cellular uptake) to peptide portion of B (i.e., the delivery sequence) by a covalent linkage.
  • the covalent linkage comprises an ether bond, thioether bond, amine bond, amide bond, oxime bond, hydrazone bond, carbon-carbon bond, carbon-nitrogen bond, carbon-oxygen bond, or carbon-sulfur bond.
  • X comprises a peptide linkage.
  • the peptide linkage comprises L- amino acids and/or D-amino acids.
  • D-amino acids are preferred in order to minimize immunogenicity and nonspecific cleavage by background peptidases or proteases.
  • Cellular uptake of oligo-D-arginine sequences is known to be as good as or better than that of oligo-L-arginines.
  • a X linker is designed for cleavage in the presence of particular conditions or in a particular environment.
  • a X linker is cleavable under physiological conditions. Cleavage of such a X linker may, for example, be enhanced or may be affected by particular pathological signals or a particular environment related to cells in which cargo delivery is desired.
  • the design of a X linker for cleavage by specific conditions such as by a specific enzyme, allows the targeting of cellular uptake to a specific location where such conditions obtain.
  • one important way that selective delivery molecules provide specific targeting of cellular uptake to desired cells, tissues, or regions is by the design of the linker portion X to be cleaved by conditions near such targeted cells, tissues, or regions.
  • X is a pH-sensitive linker. In some embodiments, X is cleaved under basic pH conditions. In some embodiments, X is cleaved under acidic pH conditions. In some embodiments, X is cleaved by a protease, a matrix metalloproteinase, or a combination thereof. In some embodiments, X is cleaved by a reducing agent.
  • X is cleaved by an MMP.
  • MMPs matrix metalloproteinases
  • uptake of molecules having features of the invention are able to direct cellular uptake of cargo (at least one D moiety) to specific cells, tissues, or regions having active MMPs in the extracellular environment.
  • a X linker that includes the amino-acid sequences PLG-C(Me)-AG (SEQ ID NO: 1), PLGLAG (SEQ ID NO: 2) which are cleaved by the metalloproteinase enzymes MMP-2, MMP-9, or MMP-7 (MMPs involved in cancer and inflammation).
  • X is cleaved by proteolytic enzymes or reducing environment, as may be found near cancerous cells. Such an environment, or such enzymes, are typically not found near normal cells.
  • X is cleaved by serine proteases including but not limited to thrombin.
  • X is cleaved in or near tissues suffering from hypoxia.
  • cleavage in or near hypoxic tissues enables targeting of cancer cells and cancerous tissues, infarct regions, and other hypoxic regions.
  • X comprises a disulfide bond.
  • a linker comprising a disulfide bond is preferentially cleaved in hypoxic regions and so targets cargo delivery to cells in such a region.
  • Hypoxia is thought to cause cancer cells to become more resistant to radiation and chemotherapy, and also to initiate angiogenesis.
  • X is cleaved in a necrotic environment. Necrosis often leads to the release of enzymes or other cell contents that may be used to trigger cleavage of a X linker. In some embodiments, cleavage of X by necrotic enzymes (e.g., by calpains) allows cargo to be taken up by diseased cells and by neighboring cells that had not yet become fully leaky.
  • necrotic enzymes e.g., by calpains
  • X is an acid-labile linker.
  • X comprises an acetal or vinyl ether linkage. Acidosis is observed in sites of damaged or hypoxic tissue, due to the Warburg shift from oxidative phosphorylation to anaerobic glycolysis and lactic acid production. In some embodiments, acidosis is used as a trigger of cargo uptake by replacing some of the arginines within B by histidines, which only become cationic below pH 7.
  • a linker X disclosed herein may include non-standard amino acids, such as, for example, hydroxylysine, desmosine, isodesmosine, or other non-standard amino acids.
  • a linker disclosed herein may include modified amino acids, including post-translationally modified amino acids such as, for example, methylated amino acids (e.g., methyl histidine, methylated forms of lysine, etc.), acetylated amino acids, amidated amino acids, formylated amino acids, hydroxylated amino acids, phosphorylated amino acids, or other modified amino acids.
  • a linker disclosed herein may also include peptide mimetic moieties, including portions linked by non-peptide bonds and amino acids linked by or to non-amino acid portions.
  • the linker X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac).
  • the linker X comprises the amino acid sequence PLGLAG.
  • the linker X comprises the amino acid sequence PLG-C(me)-AG.
  • the linker X comprises the amino acid sequence PLGxAG, wherein x is any amino acid (naturally-occuring or non-naturally occurring).
  • the linker X comprises the amino acid sequence RPLALWRS.
  • the linker X comprises the amino acid sequence ESPAYYTA. In some embodiments, the linker X comprises the amino acid sequence DPRSFL. In some embodiments, the linker X comprises the amino acid sequence PPRSFL. In some embodiments, the linker X comprises the amino acid sequence RLQLKL. In some embodiments, the linker X comprises the amino acid sequence RLQLK(Ac).
  • the linker X comprises a peptide selected from: PR(S/T)(L/I)(S/T), where the letters in parentheses indicate that either one of the indicated amino acids may be at that position in the sequence); GGAANLVRGG; SGRIGFLRTA; SGRSA; GFLG; ALAL; FK;
  • GGPRGLPG HSSKLQ; LVLA-SSSFGY; GVSQNY-PIVG; GVVQA-SCRLA; f(Pip)R-S, where "f ' indicates D-phenylalanine and "Pip” indicates piperidine-2-carboxylic acid (pipecolinic acid, a proline analog having a six-membered ring); DEVD; GWEHDG; RPLALWRS, or a combination thereof.
  • X is cleaved under hypoxic conditions.
  • X comprises a disulfide linkage.
  • X comprises a quinine.
  • X is cleaved under necrotic conditions. In some embodiments, X comprises a molecule cleavable by a calpain.
  • X comprises 6-aminohexanoyl, 5-(amino)-3-oxapentanoyl, or a combination thereof. In some embodiments, X comprises a disulfide linkage. [00170] In some embodiments, the linker is an alkyl. In some embodiments, the linker is heteroalkyl.
  • the linker is an alkylene. In some embodiments, the linker is an alkenylene. In some embodiments, the linker is an alkynylene. In some embodiments, the linker is a heteroalkylene.
  • a selective delivery molecules disclosed herein comprises a single of linker. Use of a single mechanism to mediate uptake of both imaging and therapeutic cargoes is particularly valuable, because imaging with noninjurious tracer quantities can be used to test whether a subsequent therapeutic dose is likely to concentrate correctly in the target tissue.
  • a selective delivery molecules disclosed herein comprises a plurality of linkers. Where a selective delivery molecule disclosed herein includes multiple X linkages, separation of portion of A from the other portions of the molecule requires cleavage of all X linkages. Cleavage of multiple X linkers may be simultaneous or sequential. Multiple X linkages may include X linkages having different specificities, so that separation of portion of A from the other portions of the molecule requires that more than one condition or environment ("extracellular signals") be encountered by the molecule. Cleavage of multiple X linkers thus serves as a detector of combinations of such extracellular signals.
  • a selective delivery molecule may include two linker portions Xa and Xb connecting basic portion of B with acidic portion of A. Both X linkers a and Xb must be cleaved before acidic portion of A is separated from basic portion of B allowing entry of portion of B and cargo moiety C (if any) to enter a cell. It will be understood that a linker region may link to either a basic portion of B or a cargo moiety C independently of another linker that may be present, and that, where desired, more than two linker regions X may be included.
  • Combinations of two or more X linkers may be used to further modulate the targeting and delivery of molecules to desired cells, tissue or regions. Combinations of extracellular signals are used to widen or narrow the specificity of the cleavage of X linkers if desired. Where multiple X linkers are linked in parallel, the specificity of cleavage is narrowed, since each X linker must be cleaved before portion of A may separate from the remainder of the molecule. Where multiple X linkers are linked in series, the specificity of cleavage is broadened, since cleavage of any one X linker allows separation of portion of A from the remainder of the molecule.
  • a X linker is designed to place the protease- sensitive and reduction-sensitive sites in tandem, so that cleavage of either would suffice to allow separation of the acidic portion of A.
  • a X linker is designed to place the protease sensitive site between at least one pair of cysteines that are disulfide- bonded to each other. In that case, both protease cleavage and disulfide reduction are required in order to allow separation of portion of A.
  • Y is a linker consisting of one or more amino acids is used to join Cargo (D) to the remainder of the SDM. In some embodiments, Y is a linker consisting of one or more amino acids is used to join Cargo (D) to portion B.
  • the peptide linker will have no specific biological activity other than to join the molecules or to preserve some minimum distance or other spatial relationship between them. However, the constituent amino acids of the linker may be selected to influence some property of the molecule such as the folding, net charge, or hydrophobicity.
  • the linker binds cargo portion of D to peptide portion of B (i.e., the delivery sequence) by a covalent linkage.
  • the covalent linkage comprises an ether bond, thioether bond, amine bond, amide bond, oxime bond, hydrazone bond, carbon-carbon bond, carbon-nitrogen bond, carbon-oxygen bond, or carbon-sulfur bond.
  • the Y linker is flexible. In some embodiments, the Y linker is rigid. In some embodiments, the Y linker comprises a linear structure. In some embodiments, the Y linker comprises a non-linear structure. In some embodiments, the Y linker comprises a branched structure. In some embodiments, the linker comprises a cyclic structure.
  • Y linker comprises a peptide linkage.
  • the peptide linkage comprises L-amino acids and/or D-amino acids.
  • D-amino acids are preferred in order to minimize immunogenicity and nonspecific cleavage by background peptidases or proteases.
  • Cellular uptake of oligo-D-arginine sequences is known to be as good as or better than that of oligo-L-arginines.
  • a Y linker is designed for cleavage in the presence of particular conditions or in a particular environment.
  • a Y linker is cleavable by an intracellular protease.
  • Y is cleavable by an intracellular protease.
  • a Y linker is cleavable by a lysosomal protease.
  • the intracellular protease is a cysteine protease.
  • the intracellular protease is an aspartyl protease.
  • the intracellular protease is a serine protease.
  • the cysteine protease is a caspase, a cathepsin, calpain, papain or a legumain.
  • the intracellular protease is an initiator caspase. In some embodiments, the intracellular protease is an effector caspase.
  • the Y linker is cleavable by a protease selected from among cathepsin B, cathepsin L, cathepsin H, cathepsin K, cathepsin W, cathepsin C, cathepsin F, cathepsin V, cathepsin X, cathepsin S, cathepsin D, cathepsin G, HCP-1, HCP-2, dipeptidyl-peptidase I, MEROPS C 13, CED-3 peptidase, caspase 2, caspase 3, caspase 6, caspase 7, caspase 8, caspase 9, caspase 10, caspase 11; caspase 12, caspase 13, and caspase 14.
  • a protease selected from among cathepsin B, cathepsin L, cathepsin H, cathepsin K, cathepsin W, cathepsin C, cathepsin F, cathepsin V, ca
  • the Y linker is cleavable by a protease selected from among cathepsin B, cathepsin L, caspase 3, caspase 7, caspase 8, and caspase 9.
  • a Y linker is cleavable by Cathepsin B a dipeptidyl carboxypeptidase.
  • the linker has a lysine, citrulline, or arginine residue at the PI position and a large hydrophobic residue at the PI ' position.
  • the Y linker comprises an acid sensitive chemical linker.
  • acid sensitive chemical linker is hydrazone or a derivative thereof.
  • a Y linker comprises a self-immolative spacer.
  • the self- immolative spacer is of sufficient length to prevent the occurrence of steric hindrance between the B portion of the SDM and the therapeutic cargo.
  • Y comprises a p- aminobenzyl alcohol (PABOH) spacer or a derivative thereof.
  • Y comprises a p-aminobenzyl carbonyl (PABC) spacer or a derivative thereof.
  • Y comprises a branched bis(hydroxymethyl)styrene (BHMS) spacer or a derivative thereof.
  • BHMS branched bis(hydroxymethyl)styrene
  • Y comprises a 2-aminoimidazol-5 -methanol derivative or an ortho or para- aminobenzylacetal spacer.
  • Y comprises 2,6-bishydroxymethyl-p-cresol or hemithioaminal derivatives.
  • the Y linker comprises the lysosomally cleavable peptide. In some embodiments, the Y linker comprises the lysosomally cleavable dipeptide Phe-Arg. In some embodiments, the Y linker comprises the lysosomally cleavable dipeptide Phe-Lys. In some embodiments, the Y linker comprises the lysosomally cleavable dipeptide Val-Cit (1-citrulline). In some embodiments, the Y linker comprises the lysosomally cleavable tetrapeptide Gly-Phe-Leu- Gly. In some embodiments, the Y linker comprises the lysosomally cleavable tetrapeptide Ala-Leu- Ala-Leu.
  • the Y linker comprises the lysosomally cleavable peptide and a self-immolative spacer.
  • Y is a pH-sensitive linker. In some embodiments, Y is cleaved under acidic pH conditions. In some embodiments, Y is cleaved under acidic pH conditions of the lysosome.
  • a Y linker disclosed herein may include non-standard amino acids, such as, for example, hydroxylysine, desmosine, isodesmosine, or other non-standard amino acids.
  • a linker disclosed herein may include modified amino acids, including post-translationally modified amino acids such as, for example, methylated amino acids (e.g., methyl histidine, methylated forms of lysine, etc.), acetylated amino acids, amidated amino acids, formylated amino acids, hydroxylated amino acids, phosphorylated amino acids, or other modified amino acids.
  • a linker disclosed herein may also include peptide mimetic moieties, including portions linked by non-peptide bonds and amino acids linked by or to non-amino acid portions.
  • an SDM provided herein is conjugated to an imaging agent.
  • the imaging agent is conjugated to portion of A, portion of B or both portions A and B.
  • the imaging agent is conjugated to the target ligand.
  • an imaging agent is a dye.
  • an imaging agent is a fluorescent moiety.
  • a fluorescent moiety is selected from: a fluorescent protein, a fluorescent peptide, a fluorescent dye, a fluorescent material or a combination thereof.
  • fluorescent moieties are encompassed within the term "fluorescent moiety.” Specific examples of fluorescent moieties given herein are illustrative and are not meant to limit the fluorescent moieties for use with the targeting molecules disclosed herein.
  • fluorescent dyes include, but are not limited to, xanthenes (e.g., rhodamines, rhodols and fluoresceins, and their derivatives); bimanes; coumarins and their derivatives (e.g., umbelliferone and aminomethyl coumarins); aromatic amines (e.g., dansyl; squarate dyes);
  • benzofurans fluorescent cyanines; indocarbocyanines; carbazoles; dicyanomethylene pyranes; polymethine; oxabenzanthrane; xanthene; pyrylium; carbostyl; perylene; acridone; quinacridone; rubrene; anthracene; coronene; phenanthrecene; pyrene; butadiene; stilbene; porphyrin;
  • pthalocyanine lanthanide metal chelate complexes
  • rare-earth metal chelate complexes and derivatives of such dyes.
  • fluorescein dyes include, but are not limited to, 5-carboxyfluorescein, fluorescein-5-isothiocyanate, fiuorescein-6-isothiocyanate and 6-carboxyfluorescein.
  • rhodamine dyes include, but are not limited to, tetramethylrhodamine-6- isothiocyanate, 5-carboxytetramethylrhodamine, 5-carboxy rhodol derivatives, tetramethyl and tetraethyl rhodamine, diphenyldimethyl and diphenyldiethyl rhodamine, dinaphthyl rhodamine, rhodamine 101 sulfonyl chloride (sold under the trade name of TEXAS RED®).
  • cyanine dyes include, but are not limited to, Cy3, Cy3B, Cy3.5, Cy5, Cy5.5, Cy7, IRDYE680, Alexa Fluor 750, IRDye800CW, ICG.
  • fluorescent peptides include GFP (Green Fluorescent Protein) or derivatives of GFP (e.g., EBFP, EBFP2, Azurite, mKalamal, ECFP, Cerulean, CyPet, YFP, Citrine, Venus, YPet).
  • GFP Green Fluorescent Protein
  • derivatives of GFP e.g., EBFP, EBFP2, Azurite, mKalamal, ECFP, Cerulean, CyPet, YFP, Citrine, Venus, YPet.
  • Fluorescent labels are detected by any suitable method.
  • a fluorescent label may be detected by exciting the fluorochrome with the appropriate wavelength of light and detecting the resulting fluorescence, e.g., by microscopy, visual inspection, via photographic film, by the use of electronic detectors such as charge coupled devices (CCDs), photomultipliers, etc.
  • CCDs charge coupled devices
  • the imaging agent is labeled with a positron-emitting isotope (e.g., 18 F) for positron emission tomography (PET), gamma-ray isotope (e.g., 99m Tc) for single photon emission computed tomography (SPECT), or a paramagnetic molecule or nanoparticle (e.g.,Gd 3+ chelate or coated magnetite nanoparticle) for magnetic resonance imaging (MRI).
  • a positron-emitting isotope e.g., 18 F
  • PET positron emission tomography
  • gamma-ray isotope e.g., 99m Tc
  • SPECT single photon emission computed tomography
  • MRI magnetic resonance imaging
  • the imaging agent is labeled with: a gadolinium chelate, an iron oxide particle, a super paramagnetic iron oxide particle, an ultra small paramagnetic particle, a manganese chelate or gallium containing agent.
  • gadolinium chelates include, but are not limited to diethylene triamine pentaacetic acid (DTPA), l ,4,7,10-tetraazacyclododecane-l ,4,7,10-tetraacetic acid (DOTA), and l,4,7-triazacyclononane-N,N',N"-triacetic acid (NOT A).
  • the imaging agent is a near-infrared fluorophore for near-infra red (near-IR) imaging, a luciferase (firefly, bacterial, or coelenterate) or other luminescent molecule for bioluminescence imaging, or a perfluorocarbon- filled vesicle for ultrasound.
  • the imaging agent is a nuclear probe.
  • the imaging agent is a SPECT or PET radionuclide probe.
  • the radionuclide probe is selected from: a technetium chelate, a copper chelate, a radioactive fluorine, a radioactive iodine, a indiuim chelate.
  • Tc chelates include, but are not limited to HYNIC, DTPA, and DOTA.
  • the imaging agent contains a radioactive moiety, for example a radioactive isotope such as 211 At, 1 1 I, 125 1, 90 Y, 186 Re, 18 Re, 153 Sm, 212 Bi, 32 P, 64 Cu radioactive isotopes of Lu, and others.
  • a radioactive isotope such as 211 At, 1 1 I, 125 1, 90 Y, 186 Re, 18 Re, 153 Sm, 212 Bi, 32 P, 64 Cu radioactive isotopes of Lu, and others.
  • a selective delivery molecule according to Formulas I-VI comprising an imaging agent is employed in guided surgery.
  • the selective delivery molecule preferentially localized to cancerous, or other undesirable tissues (i.e. necrotic tissues).
  • a selective delivery molecule according to Formula I comprising an imaging agent is employed in a guided surgery to remove colorectal cancer.
  • guided surgery employing the selective delivery molecule allows a surgeon to excise as little healthy (i.e., non-cancerous) tissue as possible.
  • guided surgery employing the selective delivery molecule allows a surgeon to visualize and excise more cancerous tissue than the surgeon would have been able to excise without the presence of the selective delivery molecule.
  • the surgery is fluorescence-guided surgery.
  • the selective delivery molecule comprises a structure selected from SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM-110, SDM-111 , SDM-112, SDM-1 13, SDM-114, SDM-115, SDM-1 16, SDM- 1 17, SDM-1 18, SDM-119, SDM-120, SDM-121 , SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM- 134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM- 141 , SDM-142, SDM-143, SDM-144, SDM-145, SDM-146, SDM-147
  • the selective delivery molecule is a derivative of SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM- 108, SDM-109, SDM-110, SDM-111 , SDM-112, SDM-1 13, SDM-114, SDM-115, SDM-1 16, SDM-1 17, SDM-118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM- 125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141 , SDM- 142, SDM-143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148
  • the selective delivery molecule comprises a structure selected from: SDM-1 , SDM-2, SDM-3, SDM-4, SDM-5, SDM-6, SDM-7, SDM-8, SDM-9, SDM-10, SDM-1 1 , SDM-12, SDM-13, SDM-14, SDM-15, SDM-16, SDM-17, SDM-18, SDM-19, SDM-20, SDM-21 , SDM-22, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-28, SDM-29, SDM-30, SDM-31 , SDM-32, SDM-33, SDM-34, SDM-35, SDM-36, SDM-37, SDM-38, SDM-39, SDM-40, SDM-41 , SDM-42, SDM-43, SDM-44, SDM-45, SDM-46, SDM-47, SDM-48, SDM-49, SDM-50, SDM- 1
  • the selective delivery molecule comprises a structure selected from: SDM-14, SDM-15, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-32, or SDM-35.
  • the selective delivery molecule is derived from Peptide P-l, P-2, P-3, P-4, P-5, P-6, P-7, P-8, P-9, P-10, P-l 1, P- 12, P-13, P-14, P-15, P-16, P-17, P-18, P-19, P-20, P-21 , P-21 , or P-3.
  • the antibody-conjugated SDMs described herein are optionally conjugated to high molecular weight molecules that increase the multivalency and avidity of labeling.
  • the high molecular weight molecules are water-soluble polymers.
  • suitable water-soluble polymers include, but are not limited to, peptides, saccharides, poly(vinyls), poly(ethers), poly(amines), poly(carboxylic acids) and the like.
  • the water-soluble polymer is dextran, polyethylene glycol (PEG), polyoxyalkylene, polysialic acid, starch, or hydroxyethyl starch. Any suitable method is used to conjugate peptides to water-soluble polymers ( see Hermanson G., Bioconjugate Techniques 2 nd Ed., Academic Press, Inc. 2008).
  • compositions comprising any of SDMs as disclosed herein.
  • the pharmaceutical compositions comprising an SDM comprises an SDM of any of Formulas I-VI and a pharmaceutically acceptable carrier.
  • compositions comprising any of the antibody-conjugated SDMs as disclosed herein.
  • the pharmaceutical compositions comprising any of the antibody-conjugated SDMs as disclosed herein.
  • compositions herein are formulated using one or more physiologically acceptable carriers including excipients and auxiliaries which facilitate processing of the active agents into preparations which are used pharmaceutically. Proper formulation is dependent upon the route of administration chosen.
  • a summary of pharmaceutical compositions is found, for example, in Remington: The Science and Practice of Pharmacy, Nineteenth Ed (Easton, Pa.: Mack Publishing Company, 1995); Hoover, John E., Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania 1975; Liberman, H.A. and Lachman, L., Eds.,
  • a pharmaceutical composition disclosed herein further comprises a pharmaceutically acceptable diluent(s), excipient(s), or carrier(s).
  • the pharmaceutical compositions includes other medicinal or pharmaceutical agents, carriers, adjuvants, such as preserving, stabilizing, wetting or emulsifying agents, solution promoters, salts for regulating the osmotic pressure, and/or buffers.
  • the pharmaceutical compositions also contain other therapeutically valuable substances. WSGR Docket No. 39088-711.601
  • a pharmaceutical composition disclosed herein is administered to a subject by any suitable administration route, including but not limited to, parenteral (intravenous, subcutaneous, intraperitoneal, intramuscular, intravascular, intrathecal, intravitreal, infusion, or local) administration.
  • parenteral intravenous, subcutaneous, intraperitoneal, intramuscular, intravascular, intrathecal, intravitreal, infusion, or local
  • Formulations suitable for intramuscular, subcutaneous, peritumoral, or intravenous injection include physiologically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions.
  • suitable aqueous and non-aqueous carriers, diluents, solvents, or vehicles including water, ethanol, polyols (propyleneglycol, polyethylene-glycol, glycerol, cremophor and the like), suitable mixtures thereof, vegetable oils (such as olive oil) and injectable organic esters such as ethyl oleate.
  • Formulations suitable for subcutaneous injection also contain optional additives such as preserving, wetting, emulsifying, and dispensing agents.
  • an active agent is optionally formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hank's solution, Ringer's solution, or physiological saline buffer.
  • Parenteral injections optionally involve bolus injection or continuous infusion.
  • Formulations for injection are optionally presented in unit dosage form, e.g., in ampoules or in multi dose containers, with an added preservative.
  • the pharmaceutical composition described herein are in a form suitable for parenteral injection as a sterile suspensions, solutions or emulsions in oily or aqueous vehicles, and contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
  • Pharmaceutical formulations for parenteral administration include aqueous solutions of an active agent in water soluble form. Additionally, suspensions are optionally prepared as appropriate oily injection suspensions.
  • the pharmaceutical composition described herein is in unit dosage forms suitable for single administration of precise dosages.
  • the formulation is divided into unit doses containing appropriate quantities of an active agent disclosed herein.
  • the unit dosage is in the form of a package containing discrete quantities of the formulation.
  • Non-limiting examples are packaged tablets or capsules, and powders in vials or ampoules.
  • aqueous suspension compositions are packaged in single-dose non-reclosable containers.
  • multiple-dose reclosable containers are used, in which case it is typical to include a preservative in the composition.
  • formulations for parenteral injection are presented in unit dosage form, which include, but are not limited to ampoules, or in multi dose containers, with an added preservative.
  • the SDMs of Formulas I-VI and carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, allow the targeted delivery of therapeutic agents and/or imaging agents to specific cells and/or tissues.
  • the molecules comprise a basic peptide sequence (B) which is designed to be transported across a cellular membrane, an acidic peptide sequence (A) which inhibits uptake of peptide B into cells, a linker X which is cleavable under specific conditions, cargo moieties (at least DA and DB) bound to peptides A and B, or X and a macromolecular carrier.
  • cleavage of the linker X linker frees peptide B from peptide A and allows the transport of peptide B (and any cargo attached thereto) across a cellular membrane.
  • the selective delivery molecules of Formulas I-IV enable targeted delivery of one or more cargos (e.g., therapeutic agents or imaging agents) to a cell tissue.
  • delivering cargo to a tissue of interest comprising contacting the tissue of interest with an antibody-conjugated SDM comprising a targeting antibody conjugated to an SDM of any of Formulas I-VI.
  • the tissue of interest is cancerous tissue (or, cancer).
  • the cancerous tissue is: breast cancer tissue, colon cancer tissue, squamous cell carcinoma tissue, prostate cancer tissue, melanoma tissue, or thyroid cancer tissue.
  • the cancerous tissue is breast cancer tissue.
  • the cancerous tissue is colon cancer tissue.
  • the tissue of interest is an inflamed tissue.
  • the inflamed tissue is the result if acute or chronic inflammation.
  • the inflamed tissue is caused by an inflammatory disease is or is associated with an inflammatory disease.
  • the inflamed tissue is caused by an inflammatory disease is or is associated with rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, sepsis, erythema nodosum leprosum, multiple sclerosis, psoriasis, systemic lupus erythematosis, type I diabetes, atherosclerosis, encephalomyelitis, Alzheimer's disease, stroke, traumatic brain injury, Parkinson's disease or septic shock.
  • an inflammatory disease is or is associated with rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, sepsis, erythema nodosum leprosum, multiple sclerosis, psoriasis, systemic lupus erythematosis, type I diabetes, atherosclerosis, encephalomyelitis, Alzheimer's disease, stroke, traumatic brain injury, Parkinson's
  • the SDMs of Formulas I-VI and carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, allow the targeted delivery of therapeutic agents to specific cells and/or tissues (e.g., cancerous tissues).
  • the molecules comprise a basic peptide sequence (B) which is designed to be transported across a cellular membrane, an acidic peptide sequence (A) which inhibits uptake of peptide B into cells, a linker X which is cleavable under specific conditions, therapeutic agents bound to peptides A and B, or X and a macromolecular carrier.
  • the SDMs of Formulas I-VI and carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, enable targeted delivery of one or more therapeutic agents to a cell or tissue.
  • targeted delivery of a therapeutic agent to a cell or tissue enables a medical professional to treat a specific tissue.
  • targeted delivery of a therapeutic agent to a cell or tissue enables a medical professional to treat a specific tissue (e.g., cancerous tissue).
  • targeted delivery of a therapeutic agent to a cell or tissue decreases the dosage of the therapeutic agent.
  • targeted delivery of a therapeutic agent to a cell or tissue decreases contact of the therapeutic agent with healthy tissue.
  • targeted delivery of a therapeutic agent to a cell or tissue decreases unwanted side-effects arising from use of high concentrations of a therapeutic agent or contact.
  • targeted delivery of a therapeutic agent to a cell or tissue decreases unwanted side-effects arising from contact between the therapeutic agent and healthy tissue.
  • an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, is employed for the treatment of cancer.
  • the cancer is AIDS-related cancers (e.g., AIDS-related lymphoma), anal cancer, basal cell carcinoma, bile duct cancer (e.g., extrahepatic), bladder cancer, bone cancer, (osteosarcoma and malignant fibrous histiocytoma), breast cancer, cervical cancer, colon cancer, colorectal cancer, endometrial cancer (e.g., uterine cancer), ependymoma, esophageal cancer, eye cancer (e.g., intraocular melanoma and retinoblastoma), gastric (stomach) cancer, germ cell tumor, (e.g., extracranial, extragonadal, ovarian), head and neck cancer, leukemia, lip and oral cavity cancer, liver cancer, lung cancer (e.g., small cell lung cancer, non-small cell lung cancer, adenocarcinoma of the lung, and squamous carcinoma of the lung),
  • AIDS-related cancers
  • the cancer is a lymphoid cancer (e.g., lymphoma).
  • the cancer is a B-cell cancer.
  • the cancer is precursor B-cell cancers (e.g., precursor B-lymphoblastic leukemia/lymphoma) and peripheral B- cell cancers (e.g., B-cell chronic lymphocytic leukemia/pro lymphocytic leukemia/small lymphocytic lymphoma (small lymphocytic (SL) NHL), lymphoplasmacytoid
  • lymphoma/immunocytoma mantel cell lymphoma, follicle center lymphoma, follicular lymphoma (e.g., cytologic grades: I (small cell), II (mixed small and large cell), III (large cell) and/or subtype: diffuse and predominantly small cell type), low grade/follicular non-Hodgkin's lymphoma (NHL), intermediate grade/follicular NHL, marginal zone B-cell lymphoma (e.g., extranodal (e.g., MALT- type +/- monocytoid B cells) and/or Nodal (e.g., +/- monocytoid B cells)), splenic marginal zone lymphoma (e.g., +/- villous lymphocytes), Hairy cell leukemia, plasmacytoma/plasma cell myeloma (e.g., myeloma and multiple myeloma), diffuse large B-cell lymphoma (e
  • the cancer is a T-cell and/or putative NK-cell cancer.
  • the cancer is precursor T-cell cancer (precursor T-lymphoblastic
  • T-cell chronic lymphocytic leukemia/prolymphocytic leukemia and large granular lymphocyte leukemia (LGL) (e.g., T-cell type and/or NK-cell type), cutaneous T-cell lymphoma (e.g., mycosis fungoides/Sezary syndrome), primary T-cell lymphomas unspecified (e.g., cytological categories (e.g., medium-sized cell, mixed medium and large cell), large cell, lymphoepitheloid cell, subtype hepatosplenic ⁇ T-cell
  • cytological categories e.g., medium-sized cell, mixed medium and large cell
  • large cell lymphoepitheloid cell
  • angioimmunoblastic T-cell lymphoma e.g., angiocentric lymphoma
  • intestinal T-cell lymphoma e.g., +/- enteropathy associated
  • adult T-cell lymphoma/leukemia ATL
  • anaplastic large cell lymphoma e.g., CD30+, T- and null-cell types
  • anaplastic large-cell lymphoma e.g., CD30+, T- and null-cell types
  • Hodgkin's like e.gkin's like.
  • the cancer is Hodgkin's disease.
  • the cancer is leukemia.
  • the cancer is chronic myelocytic I (granulocytic) leukemia, chronic myelogenous, and chronic lymphocytic leukemia (CLL), acute lymphoblastic leukemia (ALL), acute myeloid leukemia, acute lymphocytic leukemia, and acute myelocytic leukemia (e.g., myeloblastic, promyelocytic, myelomonocytic, monocytic, and erythroleukemia).
  • CLL chronic lymphocytic leukemia
  • ALL acute lymphoblastic leukemia
  • acute myeloid leukemia acute lymphocytic leukemia
  • acute myelocytic leukemia e.g., myeloblastic, promyelocytic, myelomonocytic, monocytic, and erythroleukemia.
  • the cancer is a liquid tumor or plasmacytoma. In some embodiments, the cancer is a liquid tumor or plasmacytoma.
  • the cancer is extramedullary plasmacytoma, a solitary myeloma, and multiple myeloma.
  • the plasmacytoma is multiple myeloma.
  • the cancer is lung cancer.
  • the cancer is prostate cancer.
  • the prostate cancer is an adenocarcinoma.
  • the prostate cancer is a sarcoma
  • the prostate cancer is stage A prostate cancer (the cancer cannot be felt during a rectal exam).
  • the prostate cancer is stage B prostate cancer (i.e., the tumor involves more tissue within the prostate, it can be felt during a rectal exam, or it is found with a biopsy that is done because of a high PSA level).
  • the prostate cancer is stage C prostate cancer (i.e., the cancer has spread outside the prostate to nearby tissues).
  • the prostate cancer is stage D prostate cancer.
  • the prostate cancer is androgen independent prostate cancer (AIPC).
  • the prostate cancer is androgen dependent prostate cancer.
  • the prostate cancer is refractory to hormone therapy. In some embodiments, the prostate cancer is substantially refractory to hormone therapy. In some embodiments, the prostate cancer is refractory to chemotherapy. In some embodiments, the prostate cancer is metastatic prostate cancer. In some embodiments, the individual is a human who has a gene, genetic mutation, or polymorphism associated with prostate cancer (e.g.,
  • RNASEL/HPC 1 RNASEL/HPC 1 , ELAC2/HPC2, SR-A/MSRl , CHEK2, BRCA2, PO 1 , OGG1 , MIC-1 , TLR4, and PTEN
  • the prostate cancer is HER2 positive. In some embodiments, the prostate cancer is HER2 negative.
  • the cancer has metastasized and is characterized by circulating tumor cells.
  • an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody is employed for the treatment of inflammation or an inflammatory disease.
  • the inflammation is chronic inflammation.
  • the inflammation is acute inflammation.
  • inflammation or inflammatory disease is or is associated with rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, sepsis, erythema nodosum leprosum, multiple sclerosis, psoriasis, systemic lupus erythematosis, type I diabetes, atherosclerosis,
  • an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody is employed for the treatment of an autoimmune disease.
  • the autoimmune disease is Celiac disease, diabetes mellitus type 1, Sarcoidosis, systemic lupus erythematosus (SLE), Sjogren's syndrome, Churg-Strauss Syndrome, Hashimoto's thyroiditis, Graves' disease, idiopathic thrombocytopenic purpura, Addison's Disease, rheumatoid arthritis (RA), Polymyositis (PM), or Dermatomyositis (DM).
  • a therapeutic agent is selected from: a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, an anti-inflammatory agent, or a combination thereof.
  • a therapeutic agent is a CD79A inhibitor, a CD79B inhibitor, a CD 19 inhibitor, a Lyn inhibitor, a Syk inhibitor, a PI3K inhibitor, a Blnk inhibitor, a PLCy inhibitor, a PKCp inhibitor, or a combination thereof.
  • a therapeutic agent is an antibody, B cell receptor signaling inhibitor, a PI3K inhibitor, an IAP inhibitor, an mTOR inhibitor, a radioimmunotherapeutic, a DNA damaging agent, a proteosome inhibitor, a histone deacytlase inhibitor, a protein kinase inhibitor, a hedgehog inhibitor, an Hsp90 inhibitor, a telomerase inhibitor, a Jakl/2 inhibitor, a protease inhibitor, a PKC inhibitor, a PARP inhibitor, or a combination thereof.
  • a therapeutic agent is a B cell receptor pathway inhibitor.
  • a therapeutic agent is selected from: chlorambucil, ifosphamide, doxorubicin, mesalazine, thalidomide, lenalidomide, temsirolimus, everolimus, fludarabine, fostamatinib, paclitaxel, docetaxel, ofatumumab, rituximab, dexamethasone, prednisone, CAL-101, ibritumomab, tositumomab, bortezomib, pentostatin, endostatin, bendamustine, chlorambucil, chlormethine, cyclophosphamide, ifosfamide, melphalan, prednimustine, trofosfamide, busulfan, mannosulfan, treosulfan, carboquone, thiotepa, triaziquone, carmustine, fotemustine, lo
  • liarozole hydrochloride lometrexol sodium, lomustine, losoxantrone hydrochloride, masoprocol, maytansine, mechlorethamine hydrochloride, megestrol acetate, melengestrol acetate, melphalan, menogaril, mercaptopurine, methotrexate, methotrexate sodium, metoprine, meturedepa, mitindomide, mitocarcin, mitocromin, mitogillin, mitomalcin, mitomycin, mitosper, mitotane, mitoxantrone hydrochloride, mycophenolic acid, nocodazoie, nogalamycin, ormaplatin, oxisuran, pegaspargase, peliomycin, pentamustine, peplomycin sulfate, perfosfamide, pipobroman, piposulfan, piroxantrone hydrochloride, plicamycin, plomestan
  • a therapeutic agent is selected from: 20-epi-l, 25 dihydroxyvitamin D3, 5-ethynyluracil, abiraterone, aclarubicin, acylfulvene, adecypenol, adozelesin, aldesleukin, ALL-TK antagonists, altretamine, ambamustine, amidox, amifostine, aminolevulinic acid, amrubicin, amsacrine, anagrelide, anastrozole, andrographolide, angiogenesis inhibitors, antagonist D, antagonist G, antarelix, anti-dorsalizing morphogenetic protein- 1, antiandrogen, prostatic carcinoma, antiestrogen, antineoplaston, antisense oligonucleotides, aphidicolin glycinate, apoptosis gene modulators, apoptosis regulators, apurinic acid, ara-CDP-DL-PTBA,
  • fluorodaunorunicin hydrochloride forfenimex, formestane, fostriecin, fotemustine, gadolinium texaphyrin, gallium nitrate, galocitabine, ganirelix, gelatinase inhibitors, gemcitabine, glutathione inhibitors, hepsulfam, heregulin, hexamethylene bisacetamide, hypericin, ibandronic acid, idarubicin, idoxifene, idramantone, ilmofosine, ilomastat, imidazoacridones, imiquimod, immunostimulant peptides, insulin-such as for example growth factor- 1 receptor inhibitor, interferon agonists, interferons, interleukins, iobenguane, iododoxorubicin, ipomeanol, 4-, iroplact, irsogladine, isobengazole, iso
  • oligonucleotides onapristone, ondansetron, ondansetron, oracin, oral cytokine inducer, ormaplatin, osaterone, oxaliplatin, oxaunomycin, palauamine, palmitoylrhizoxin, pamidronic acid, panaxytriol, panomifene, parabactin, pazelliptine, pegaspargase, peldesine, pentosan polysulfate sodium, WSGR Docket No. 39088-711.601
  • pentostatin pentrozole, perfiubron, perfosfamide, perillyl alcohol, phenazinomycin, phenylacetate, phosphatase inhibitors, picibanil, pilocarpine hydrochloride, pirarubicin, piritrexim, placetin A, placetin B, plasminogen activator inhibitor, platinum complex, platinum compounds, platinum- triamine complex, porfimer sodium, porfiromycin, prednisone, propyl bis-acridone, prostaglandin J2, proteasome inhibitors, protein A-based immune modulator, protein kinase C inhibitor, protein kinase C inhibitors, microalgal, protein tyrosine phosphatase inhibitors, purine nucleoside phosphorylase inhibitors, purpurins, pyrazoloacridine, pyridoxylated hemoglobin polyoxyethylerie conjugate, raf antagonists, raltitrexe
  • mercaptopurine thioguanine, pentostatin, mechloroethamine, cyclophosphamide, chlorambucil, meiphalan, ethylenimine, methylmelamine, hexamethlymelamine, thiotepa, busulfan, carmustine, lomusitne, semustine, streptozocin, decarbazine, fluorouracil, floxouridine, cytarabine,
  • mercaptopurine thioguanine
  • pentostatin erbulozole
  • Dolastatin 10 also known as DLS-10 and NSC-376128
  • Mivobulin isethionate also known as CI-980
  • Vincristine NSC-639829
  • Discodermolide also known as NVP-XX-A-296
  • ABT-751 Abbott, also known as WSGR Docket No. 39088-711.601
  • Altorhyrtins such as Altorhyrtin A and Altorhyrtin C
  • Spongistatins such as Altorhyrtin A and Altorhyrtin C
  • a therapeutic agent is an anti-inflammatory agent.
  • a therapeutic agent is an anti-TNF agent, an IL-1 receptor antagonist, an IL-2 receptor antagonist, a cytotoxic agent, an immunomodulatory agent, an antibiotic, a T-cell co- stimulatory blocker, a B cell depleting agent, an immunosuppressive agent, an alkylating agent, an anti-metabolite, a plant alkaloid, a terpenoids, a topoisomerase inhibitor, an antitumour antibiotic, an antibody, a hormonal therapy, an anti-diabetes agent, a leukotriene inhibitor, or combinations thereof.
  • a therapeutic agent is selected from: alefacept, efalizumab, methotrexate, acitretin, isotretinoin, hydroxyurea, mycophenolate mofetil, sulfasalazine, 6- Thioguanine, Dovonex, Taclonex, betamethasone, tazarotene, hydroxychloroquine, etanercept, adalimumab, infliximab, abatacept, rituximab, tratuzumab, Anti-CD45 monoclonal antibody AHN- 12 (NCI), Iodine-131 Anti-Bl Antibody (Corixa Corp.), anti-CD66 monoclonal antibody BW 250/183 (NCI,shire General Hospital), anti-CD45 monoclonal antibody (NCI, Baylor College of Medicine), antibody anti-anb3 integrin (NCI), BIW-8962 (BioWa Inc.), Antibody BC8
  • etoposide etoposide phosphate
  • teniposide dactinomycin
  • doxorubicin daunorubicin
  • valrubicine WSGR Docket No. 39088-711.601
  • idarubicine idarubicine
  • epirubicin bleomycin
  • plicamycin mitomycin
  • finasteride goserelin
  • an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody is administered with one or more additional therapeutic agents.
  • the additional therapeutic agent is selected from among the therapeutic agents listed herein.
  • the additional therapeutic agent is administered prior to, following, or simultaneously (i.e., concurrently) with an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, provided herein.
  • the SDMs of Formulas I-VI and carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, allow the targeted delivery of imaging agents to specific cells and/or tissues (e.g., cancerous tissues).
  • the SDMs comprise a basic peptide sequence (B) which is designed to be transported across a cellular membrane or retained by tissue, an acidic peptide sequence (A) which inhibits uptake and retention of peptide B into cells, a linker X which is cleavable under specific conditions, imaging moieties bound to peptides A and B, or X and a macromolecular carrier.
  • cleavage of the linker X linker frees peptide B from peptide A and allows the transport of peptide B (and any imaging moieties attached thereto) across a cellular membrane or retention of B to tissue.
  • the SDMs enable targeted delivery of one or more WSGR Docket No. 39088-711.601
  • imaging agents to a cell or tissue.
  • targeted delivery of an imaging agent to a cell or tissue enables a medical professional to visualize/image a specific tissue.
  • targeted delivery of an imaging agent to a cell or tissue enables a medical professional to visualize/image a specific tissue (e.g., cancerous tissue).
  • targeted delivery of an imaging agent to a cell or tissue enables a medical professional to remove (or, surgically excise) the tissue of interest (e.g., cancerous tissue).
  • targeted delivery of an imaging agent to a cell or tissue enables a medical professional to remove (or, surgically excise) the tissue of interest (e.g., cancerous tissue) with a decrease in surgical margins.
  • targeted delivery of an imaging agent to a cell or tissue enables a medical professional to remove (or, surgically excise) a tumor/cancerous tissue and decreases the chance that some of the tumor/cancerous tissue will not be removed.
  • targeted delivery of an imaging agent to a cell or tissue enables a medical professional to maximally debulk a tumor/cancerous tissue.
  • targeted delivery of an imaging agent to cancerous breast tissue decreases the chances of an unnecessary operations and re-operations.
  • targeted delivery of an imaging agent to a cell or tissue enables a medical professional to more accurately sample (e.g., biopsy (e.g., excision biopsy, incision, biopsy, aspiration biopsy, or needle biopsy)) tissue of interest (e.g., cancerous tissue).
  • tissue of interest e.g., cancerous tissue.
  • targeted delivery of an imaging agent to a cell or tissue enables a medical professional to visualize/image a specific tissue (e.g., cancerous tissue) within an excised tissue containing healthy tissue.
  • Enabling identification of target tissue can guide the pathologist on where to section of pathological evaluation and decreases the chances of a pathologist missing unhealthy tissue (e.g., cancerous tissue) and sampling healthy tissue which may produce a false negative.
  • tissue e.g., cancerous tissue
  • tissue removed following use of a compound of Formula I is used to prepare a pathology section or slide.
  • cancerous tissue removed following use of a compound of Formula I is used to prepare a pathology section or slide which is used to diagnose a tissue as malignant or benign.
  • targeted delivery of an imaging agent to cancerous breast tissue enables a medical professional to accurately stage cancer enabling medical treatment decisions.
  • targeted delivery of an imaging agent to cancerous tissue enables a medical professional to observe the size of a tumor (cancerous tissue) or the spread (e.g., metastatic lesions) of cancerous tissue.
  • targeted delivery of an imaging agent to a cell or tissue enables a medical professional to design an efficacious treatment regimen.
  • a selective delivery molecule according to Formula I comprising an imaging agent is employed in guided surgery.
  • the selective delivery molecule preferentially localized to cancerous, or other pathological tissues with up-regulated protease activity (e.g. tissues undergoing inflammatory response).
  • a selective delivery molecule according to Formula I comprising an imaging agent is employed in a guided surgery to remove colorectal cancer.
  • guided surgery employing the selective delivery molecule allows a surgeon to excise as little healthy (i.e., non-cancerous) tissue as possible.
  • guided surgery employing the selective delivery molecule allows a surgeon to visualize and excise more cancerous tissue than the surgeon would have been able to excise without the presence of the selective delivery molecule.
  • the surgery is fluorescence-guided surgery.
  • an imaging agent is a dye.
  • an imaging agent is a fluorescent moiety.
  • a fluorescent moiety is selected from: a fluorescent protein, a fluorescent peptide, a fluorescent dye, a fluorescent material or a combination thereof.
  • fluorescent moieties are encompassed within the term "fluorescent moiety.” Specific examples of fluorescent moieties given herein are illustrative and are not meant to limit the fluorescent moieties for use with the targeting molecules disclosed herein.
  • fluorescent dyes include, but are not limited to, xanthenes (e.g., rhodamines, rhodols and fluoresceins, and their derivatives); bimanes; coumarins and their derivatives (e.g., umbelliferone and aminomethyl coumarins); aromatic amines (e.g., dansyl; squarate dyes);
  • benzofurans fluorescent cyanines; indocarbocyanines; carbazoles; dicyanomethylene pyranes; polymethine; oxabenzanthrane; xanthene; pyrylium; carbostyl; perylene; acridone; quinacridone; rubrene; anthracene; coronene; phenanthrecene; pyrene; butadiene; stilbene; porphyrin;
  • pthalocyanine lanthanide metal chelate complexes
  • rare-earth metal chelate complexes and derivatives of such dyes.
  • fluorescein dyes include, but are not limited to, 5-carboxyfluorescein, fluorescein-5-isothiocyanate, fluorescein-6-isothiocyanate and 6-carboxyfluorescein.
  • rhodamine dyes include, but are not limited to, tetramethylrhodamine-6- isothiocyanate, 5-carboxytetramethylrhodamine, 5-carboxy rhodol derivatives, tetramethyl and tetraethyl rhodamine, diphenyldimethyl and diphenyldiethyl rhodamine, dinaphthyl rhodamine, rhodamine 101 sulfonyl chloride (sold under the tradename of TEXAS RED®).
  • cyanine dyes include, but are not limited to, Cy3, Cy3B, Cy3.5, Cy5, Cy5.5,
  • fluorescent peptides include GFP (Green Fluorescent Protein) or derivatives of GFP (e.g., EBFP, EBFP2, Azurite, mKalamal, ECFP, Cerulean, CyPet, YFP, Citrine, Venus, YPet).
  • GFP Green Fluorescent Protein
  • derivatives of GFP e.g., EBFP, EBFP2, Azurite, mKalamal, ECFP, Cerulean, CyPet, YFP, Citrine, Venus, YPet.
  • Fluorescent labels are detected by any suitable method.
  • a fluorescent label may be detected by exciting the fluorochrome with the appropriate wavelength of light and detecting the resulting fluorescence, e.g., by microscopy, visual inspection, via photographic film, by the use of electronic detectors such as charge coupled devices (CCDs), photomultipliers, etc.
  • CCDs charge coupled devices
  • the imaging agent is labeled with a positron-emitting isotope (e.g., 18 F) for positron emission tomography (PET), gamma-ray isotope (e.g., 99m Tc) for single photon emission computed tomography (SPECT), or a paramagnetic molecule or nanoparticle
  • a positron-emitting isotope e.g., 18 F
  • PET positron emission tomography
  • gamma-ray isotope e.g., 99m Tc
  • SPECT single photon emission computed tomography
  • SPECT single photon emission computed tomography
  • MRI magnetic resonance imaging
  • the imaging agent is labeled with: a gadolinium chelate, an iron oxide particle, a super paramagnetic iron oxide particle, an ultra small paramagnetic particle, a manganese chelate or gallium containing agent.
  • gadolinium chelates include, but are not limited to diethylene triamine pentaacetic acid (DTPA), l ,4,7,10-tetraazacyclododecane-l ,4,7,10-tetraacetic acid (DOTA), and l,4,7-triazacyclononane-N,N',N"-triacetic acid (NOT A).
  • the imaging agent is a near-infrared fluorophore for near-infra red (near-IR) imaging, a luciferase (firefly, bacterial, or coelenterate) or other luminescent molecule for bioluminescence imaging, or a perfluorocarbon- filled vesicle for ultrasound.
  • the imaging agent is a nuclear probe.
  • the imaging agent is a SPECT or PET radionuclide probe.
  • the radionuclide probe is selected from: a technetium chelate, a copper chelate, a radioactive fluorine, a radioactive iodine, a indiuim chelate.
  • Tc chelates include, but are not limited to HYNIC, DTPA, and DOTA.
  • the imaging agent contains a radioactive moiety, for example a radioactive isotope such as 211 At, 131 1, 125 1, 90 Y, 186 Re, 188 Re, 153 Sm, 212 Bi, 32 P, 64 Cu radioactive isotopes of Lu, and others.
  • a radioactive isotope such as 211 At, 131 1, 125 1, 90 Y, 186 Re, 188 Re, 153 Sm, 212 Bi, 32 P, 64 Cu radioactive isotopes of Lu, and others.
  • Xi is a cleavable linker
  • Ai is a peptide with a sequence comprising 5 to 9 acidic amino acids and having a first reactive amino acid moiety CA;
  • Bi is a peptide with a sequence comprising 7 to 9 basic amino acids and having a second reactive amino acid moiety CB;
  • Ai-Xi-Bi has a third reactive amino acid moiety CM of Ai or Xi;
  • CA is capable of reacting with a first cargo moiety comprising DA
  • CB is capable of reacting with a second cargo moiety comprising DB
  • CM is capable of reacting with a macromolecular carrier comprising M to form a molecule of Formula I.
  • the CA, CB, and CM have functional groups that are orthogonally reactive.
  • CA, CB, and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid.
  • CA, CB, and CM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a B-amino acid, or a y-amino acid.
  • CA, CB, and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
  • CA, CB, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
  • CB is any amino acid having a free thiol group.
  • c B is D-cysteine.
  • CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CA is lysine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine.
  • orthogonally reactive means a plurality of groups can be attached to a molecule via a sequence of reactions that do not cross react enabling specific attachment of each group in the presence of the others.
  • the three groups (DA, D b , and D M ) are able to be attached to Ai-Xi-Bi via CA, CB, and CM using a sequence of 3 independent reactions that do not cross react so that each group is attached to only one site of Ai-Xi-Bi.
  • the molecule further comprises a polyethylene glycol (PEG) polymer.
  • PEG polyethylene glycol
  • the PEG polymer is covalently linked to the molecule at the F(4- WSGR Docket No. 39088-711.601
  • the molecule comprises groups that can be orthogonally reacted. In some embodiments, the groups that can be orthogonally reacted are chosen from: an amine, thiol and an acetyl phenylalanine. In some embodiments, the molecule comprises an amine, a thiol, and an acetyl phenylalanine.
  • the PEG polymer has an average molecular weight of 500 daltons. In some embodiments, the PEG polymer has an average molecular weight of 2,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 3,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 4,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 5,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 10,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 12,000 daltons.
  • the PEG polymer has an average molecular weight of 15,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 20,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 30,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 40,000 daltons.
  • the molecule further comprises a fluorescent moiety.
  • a fluorescent moiety Disclosed herein, in certain embodiments, is the use of the molecule in the synthesis of a molecule according to Formulas I- VI.
  • a-Mercaptoethyl-co-methoxy, poly-oxyethylene (average molecular weight around 2,000, 5,000, 20,000 and 40,000 daltons) [mPEG(2 )-SH, mPEG(5 )-SH, mPEG(20 )-SH, mPEG(40K)-SH] and a-aminoxyl-co-methoxy, polyoxyethylene (average molecular weight around 2,000, 5,000, 20,000 and 40,000) [mPEG(2K)-ONH 2 , mPEG(5 )-ONH 2 , mPEG(20K)-ONH 2 , mPEG(40K)-ONH 2 ] were purchased from NOF America Corporation (Irvine, CA).
  • Compound 1 was supplied by GL Biochem Ltd. (Shanghai, China).
  • Doxorubicin was purchased from NuB locks LLC (Oceanside, CA).
  • Lyophilized peptide P1-P18 was supplied by Polypeptide Group (San Diego, CA).
  • 3-Maleimidopropionic acid pentafluorophenyl ester 7 was purchased from Molecular Biosciences (Boulder, CO).
  • Compound 17 was purchased from MedChem Express (Princeton, CO).
  • LC-MS analysis was carried out on an Agilent 1200 SL series in combination with AB SCIEX API 3200, equipped with CTC PAL autosampler operating at 4°C, a vacuum degasser, binary pump, UV-VIS detector, associated Analyst 1.5 analytical software and a Phenomenex column (Kinetex 2.6 ⁇ C18 100A, 100 x 2.1 mm) or a Waters 2695 separation module equipped with a Waters 2487 dual ⁇ absorbance detector in combination with Finnigan LCQ Deca XP mass spectrometer. The equipment is associated with Xcalibur analytical software and Peeke Scientific columns (Titan 200 5 ⁇ , C18-MC, 50/100 x 2.1 mm).
  • Preparation HPLC were carried out on an Agilent system (Agilent 1200 series) and a Thermo Scientific column (Hypersil Gold C I 8, 5 ⁇ , 250 x 10 mm), or a Waters Delta Prep preparative HPLC System and a Varian column (F75L, C I 8, 15 ⁇ , 1200g), or a Waters PrepLC System equipped with a Waters 2487 dual ⁇ absorbance detector, Fraction Collector III, Masslynx software and a Thermo Scientific column (Hypersil Gold C18, 5 ⁇ , 250 x 10 mm) or a Phenomenex column (luna, C 18(2), 5 ⁇ , 100A AX 150 x 30 mm).
  • the mobile phase consisted of a water (0.05% TFA)(solvent A)/acetonitrile (0.05% TFA)(solvent B) gradient unless otherwise specified.
  • Conjugate SDM-145 (3.0 mg) was dissolved in water (135 ⁇ ) to make a stock solution (0.5 mM).
  • a TCNB buffer 50 mM tris, 10 mM CaCl 2 , 150 mM NaCl, 0.05% Brij35, pH 7.5, 480 iV> in a HPLC sample vial was added SDM-145 stock solution (10 ⁇ ) and hMMP-9 (10 ⁇ , 100 nM) purchased from EMD Millipore (Billerica, MA). The resulting solution was gently mixed well and incubated 37 °C.
  • Conjugate SDM-145 (3.0 mg) was dissolved in water (135 ⁇ ) to make a stock solution (0.5 mM).
  • a sodium acetate buffer 25 mM NaAc, 1 mM EDTA, pH 5.0, 480 ⁇
  • conjugate SDM-145 stock solution (10 ⁇ ) and Cathepsin B, human liver (10 ⁇ L, 100 nM) purchased from EMD Millipore (Billerica, MA). The resulting solution was gently mixed well and incubated 37 °C.
  • Cathespin B Labile AC PP -Cortisone conjugate was synthesized as follows:
  • Example 11 Breast cancer Mouse Therapeutic model and Assay
  • mice Female BALB/c mice (8-10 weeks old) purchased from Harlan (Indianapolis, IN, 46259) or Charles River (Wilmington, MA, 01887) were used after 4-7 day of acclimatization period. All studies were conducted at research facility under the Institutional Animal Care and Use Committee (IACUC) approved protocol # EBl 1-002-009. On the first day of study, animals were weighed and assessed for health status. Only animals with no sign of disease were selected for the study. Each involved animal was lightly anesthetized with a mixture of ketamine/xylazine administered intraperitoneally to subdue voluntary movement.
  • IACUC Institutional Animal Care and Use Committee
  • Highly metastatic 4T1 tumor cells (ATCC® Number CRL-2539TM) suspended in DPBS/MatrigelTM (1 : 1 vol) were then injected subcutaneously (4xl0 5 tumor cells/50 into the right upper mammary fat pad of the lightly anesthetized animal. Each involved animal was then allowed to recover from anesthesia, housed back in the vivarium and kept under controlled environmental conditions.
  • tumor volume width x length/2.
  • each involved tumor-bearing mouse was restrained using the tail rotating tail injector (Cat.# RTI, Braintree Scientific, Inc., Braintree, MA 02185) and dosed with vehicle or test compound

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Abstract

Described herein are methods and compositions for intracellular delivery of therapeutic molecules. Disclosed herein are selective delivery conjugate comprising a targeting ligand conjugated to a selective delivery molecule (a) an acidic sequence (portion of A) which is effective to inhibit or prevent the uptake into cells or tissue retention, (b) a molecular transport or retention sequence (portion of B), and (c) a linker between portion of A and portion of B, and (d) at least one cargo moiety. Also, described are selective delivery molecules comprising a second linker comprising an intracellular cleavage site and optionally a self-immolative cleavage site.

Description

SELECTIVE DRUG DELIVERY COMPOSITIONS AND METHODS OF USE
RELATED APPLICATION
[0001] This application claims the benefit of and right of priority to U.S. Provisional Application No. 61/814,771, filed April 22, 2013, which is incorporated herein by reference in its entirety
BACKGROUND OF THE INVENTION
[0002] Targeted delivery of therapeutic agents, such as cytotoxic agents, to tumor cells is desirable to avoid killing normal cells following systemic administration of such agents. Typical targeted drug delivery systems are composed of a cytotoxic agent conjugated to a tumor-specific antibody, forming an antibody-drug conjugate (ADC), also called an "immunoconjugate". The tumor-specific antibody binds to a tumor biomarker (e.g. a tumor antigen) expressed on the surface of the tumor cells. When systemically administered, the ADC will selectively bind to tumor cells in the body, and thereby deliver the therapeutic agent intracellularly to the tumor cells, and not normal cells. The cytotoxic agent is not active when conjugated to the antibody, but becomes active upon being cleaved from the antibody intracellularly. Examples of ADCs include gemtuzumab ozogamicin (Mylotarg), brentuximab vendotin (Adcetris), trastuzumab emtasine (Kadcyla).
SUMMARY OF THE INVENTION
[0003] Described herein are compositions for the delivery of therapeutic agents.
[0004] Described herein, in certain embodiments, are selective delivery molecule conjugates comprising: (a) a selective delivery molecule of Formula I, having the structure:
Figure imgf000003_0001
Formula I
wherein,
X is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids;
CB, is 0- 1 amino acid;
DB is a therapeutic agent or an imaging agent;
wherein [cB-DB] is bound to any amino acid on B; and
(b) a carrier or targeting ligand, wherein the carrier or targeting ligand covalently bound to the selective delivery molecule. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of A. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of B. In some embodiments, the targeting ligand is an antibody. In some embodiments, the selective delivery molecule is covalently bound to any amino acid on the targeting antibody. In some embodiments, the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody. In some embodiments, the targeting ligand binds to a tumor antigen or tumor-specific receptor. In some embodiments, the targeting antibody binds to CD3, CD19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 (NCAM), CS-125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or
SLC44A4 (AGS-5). In some embodiments, the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203. In some embodiments, the targeting ligand binds to a tumor antigen or tumor-specific receptor. In some embodiments, the targeting ligand is an integrin or a lectin. In some embodiments, the carrier is a polyethylene glycol (PEG) polymer. In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In some embodiments, the therapeutic agent is a chemotherapeutic agent, a steroid, an
immunotherapeutic agent, a targeted therapy, or an anti-inflammatory agent. In some embodiments, the therapeutic agent is a cytotoxin. In some embodiments, the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, or auritstatin. In some embodiments, the therapeutic agent is cortisone. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some
embodiments, CB is selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CB is selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a τ-amino acid. In some embodiments, CB is selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a
hydroxylamine or hydrazine group. In some embodiments, CB is selected from D-cysteine, D- glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQL L, and RLQLK(Ac). In some embodiments, the selective delivery molecule of Formula I is: SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM- 1 10, SDM-1 11, SDM-112, SDM-113, SDM-114, SDM-1 15, SDM-116, SDM-117, SDM-1 18, SDM-1 19, SDM-120, SDM-121 , SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM- 127, SDM-128, SDM-129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141 , SDM-142, SDM-143, SDM- 144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151 , SDM-152, and SDM-153.
[0005] Described herein, in certain embodiments, are selective delivery molecule conjugates comprising: (a) a selective delivery molecule of Formula II, having the structure:
Figure imgf000005_0001
Formula II
wherein,
X is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids;
CB and CM each independently comprise 0-1 amino acid;
Μ is a macromolecule;
DB is a therapeutic agent or an imaging agent,
wherein [CM -M] is bound to at any position on A or X, [CB-Db] is bound to any amino acid on B; and
(b) a carrier or targeting ligand, wherein the carrier or targeting ligand is covalently bound to the selective delivery molecule.
In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of A. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of B. In some embodiments, the targeting ligand is an antibody. In some embodiments, the selective delivery molecule is covalently bound to any amino acid on the targeting antibody. In some embodiments, the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody. In some embodiments, the targeting antibody binds to a tumor antigen or a tumor antigen or tumor-specific receptor. In some embodiments, the targeting antibody binds to CD3, CD19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 (NCAM), CS-125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or
SLC44A4 (AGS-5). In some embodiments, the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203. In some embodiments, the targeting ligand binds to a tumor antigen or tumor-specific receptor. In some embodiments, the targeting ligand is an integrin or a lectin. In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In some embodiments, the therapeutic agent is a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, or an anti-inflammatory agent. In some embodiments, the therapeutic agent is a cytotoxin. In some embodiments, the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, or auritstatin. In some embodiments, the therapeutic agent is cortisone. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some
embodiments, CB and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CB and CM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a y-amino acid. In some embodiments, CB and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CB and CM are each independently selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPPvSFL, PPRSFL, RLQLKL, and RLQLK(Ac). In some embodiments, M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a polyethylene glycol (PEG) polymer, albumin, or a combination thereof. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa),
approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)). In some embodiments, the selective delivery molecule of Formula II is: SDM-101, SDM-102, SDM- 103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM-110, SDM-111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM-118, SDM-119, SDM- 120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM- 137, SDM-138, SDM-139, SDM-140, SDM-141, SDM-142, SDM-143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151, SDM-152, and SDM-153.
[0006] Described herein, in certain embodiments, are selective delivery molecule conjugates comprising: (a) a selective delivery molecule of Formula V, having the structure:
A-[CM-M]-X-B-Y-[CB-DB]
Formula V
wherein,
X is a cleavable linker;
Υ is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids;
CB and CM each independently comprise 0-1 amino acid; M is a macromolecule;
DB is a therapeutic agent or an imaging agent,
wherein [CM-M] is bound to at any position on A or X, and [CB-DB] is bound to any amino acid on B; and
(b) a carrier or targeting ligand, wherein the carrier or targeting ligand covalently bound to the selective delivery molecule.
In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of A. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of B. In some embodiments, the targeting ligand is an antibody. In some embodiments, the selective delivery molecule is covalently bound to any amino acid on the targeting antibody. In some embodiments, the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody. In some embodiments, the targeting antibody binds to a tumor antigen or a tumor antigen or tumor-specific receptor. In some embodiments, the targeting antibody binds to CD3, CD19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 ( CAM), CS-125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or
SLC44A4 (AGS-5). In some embodiments, the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203. In some embodiments, the targeting ligand binds to a tumor antigen or tumor-specific receptor. In some embodiments, the targeting ligand is an integrin or a lectin. In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In some embodiments, the therapeutic agent is a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, or an anti-inflammatory agent. In some embodiments, the therapeutic agent is a cytotoxin. In some embodiments, the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, or auritstatin. In some embodiments, the therapeutic agent is cortisone. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some
embodiments, CB and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CB and CM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a r-amino acid. In some embodiments, CB and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CB and CM are each independently selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac). In some embodiments, Y is cleavable by a protease. In some embodiments, Y is cleavable by an intracellular protease. In some embodiments, Y is cleavable by a lysosomal protease. In some embodiments, Y is cleavable by Cathepsin B. In some embodiments, Y comprises a self-immolative spacer. In some embodiments, Y comprises a PABC spacer or a derivative thereof. In some embodiments, M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a polyethylene glycol (PEG) polymer, albumin, or a combination thereof. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa),
approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)). In some embodiments, the selective delivery molecule of Formula V is: SDM-101, SDM-102, SDM-103, SDM-104, SDM-105, SDM- 106, SDM-107, SDM-108, SDM-109, SDM-110, SDM-111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM-118, SDM-119, SDM-120, SDM-121, SDM-122, SDM- 123, SDM-124, SDM-125, SDM-126, SDM-127, SDM- 128, SDM- 129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM-134, SDM-135, SDM- 136, SDM- 137, SDM-138, SDM-139, SDM- 140, SDM-141 , SDM-142, SDM-143, SDM-144, SDM- 145, SDM- 146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151 , SDM-152, and SDM- 153.
[0007] Described herein, in certain embodiments, are selective delivery molecules of Formula V, having the structure:
A-[CM-M]-X-B-Y-[CB-DB]
Formula V
wherein,
X is a cleavable linker;
Y is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids;
CB and CM each independently comprise 0-1 amino acid;
M is a macromolecule;
DB is a therapeutic agent or an imaging agent; and
wherein [CM-M] is bound to at any position of A or X, and [CB-Db] is bound to any amino acid of B. In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some
embodiments, CB and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CB and CM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a τ-amino acid. In some embodiments, CB and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CB and CM are each independently selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac). In some embodiments, Y is cleavable by a protease. In some embodiments, Y is cleavable by an intracellular protease. In some embodiments, Y is cleavable by a lysosomal protease. In some embodiments, Y is cleavable by Cathepsin B. In some embodiments, Y comprises a self-immolative spacer. In some embodiments, Y comprises a PABC spacer or a derivative thereof. In some embodiments, the therapeutic agent is a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, or an antiinflammatory agent. In some embodiments, the therapeutic agent is a cytotoxin. In some embodiments, the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, or auritstatin. In some embodiments, the therapeutic agent is cortisone. In some embodiments, M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a polyethylene glycol (PEG) polymer, albumin, or a combination thereof. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa),
approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)). In some embodiments, molecule is SDM-101, SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM- 109, SDM-110, SDM-111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM-118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM- 126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141, SDM-142, SDM- 143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151 , SDM-152, and SDM-153. In some embodiments, the selective delivery molecule is covalently bound to a carrier or targeting ligand. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of A. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of B. In some embodiments, the targeting ligand is an antibody. In some embodiments, the selective delivery molecule is covalently bound to any amino acid on the targeting antibody. In some embodiments, the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody. In some embodiments, the targeting antibody binds to a tumor antigen or a tumor antigen or tumor-specific receptor. In some embodiments, the targeting antibody binds to CD3, CD19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 (NCAM), CS- 125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or SLC44A4 (AGS-5). In some embodiments, the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab,
glembatumumab, SGN-75, PSMA ADC, ASG-5ME or mdx-1203. In some embodiments, the targeting ligand binds to a tumor antigen or tumor-specific receptor. In some embodiments, the targeting ligand is an integrin or a lectin.
[0008] Described herein, in certain embodiments, are selective delivery molecule conjugates comprising: a carrier or targeting ligand; and any selective delivery molecule provided herein. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of A. In some embodiments, the carrier or targeting ligand is covalently bound to any amino acid of B. In some embodiments, the targeting ligand is an antibody. In some embodiments, the selective delivery molecule is covalently bound to any amino acid on the targeting antibody. In some embodiments, the selective delivery molecule is covalently bound to an amino acid in the Fc portion of the antibody. In some embodiments, the targeting antibody binds to a tumor antigen or a tumor antigen or tumor-specific receptor. In some embodiments, the targeting antibody binds to CD3, CD 19, CD22, CD30, CD33, CD52, HER2 (ErB2), CD56 (NCAM), CS-125, Integrin, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), or
SLC44A4 (AGS-5). In some embodiments, the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203. In some embodiments, the targeting ligand binds to a tumor antigen or tumor-specific receptor. In some embodiments, the targeting ligand is an integrin or a lectin. In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylatmg agents, vinylsulfones, pyridyl disulfides, TNB-tbiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo.
[0009] Described herein, in certain embodiments, are selective delivery molecule according to SDM-101, SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM- 109, SDM-1 10, SDM-111, SDM-112, SDM-113, SDM-1 14, SDM-115, SDM-116, SDM-1 17, SDM-1 18, SDM-119, SDM-120, SDM-121 , SDM-122, SDM-123, SDM-124, SDM-125, SDM- 126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141 , SDM-142, SDM- 143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151 , or SDM-152
[0010] Described herein, in certain embodiments, are pharmaceutical compositions comprising a selective delivery molecule conjugate provided herein and one or more pharmaceutically acceptable carriers, glidants, diluents, or excipients.
[0011] Described herein, in certain embodiments, are methods for treating cancer in a subject in need thereof, comprising administering to a subject having cancer a therapeutically effective amount of a selective delivery molecule conjugate provided herein, thereby treating the cancer. In some embodiments, the cancer is a breast cancer, colorectal cancer, ovarian cancer, lung cancer, esophageal cancer, pancreatic cancer, gastro-intestinal cancer, squamous cell carcinoma, prostate cancer, melanoma, or thyroid cancer. In some embodiments, the therapeutic agent is a
chemotherapeutic agent. In some embodiments, the therapeutic agent is a cytotoxin. In some embodiments, the methods further comprise administering an additional anti-cancer agent.
[0012] Described herein, in certain embodiments, are methods for treatment of inflammation in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a selective delivery molecule conjugate provided herein, thereby treating the inflammation. In some embodiments, the inflammation is acute inflammation or chronic inflammation. In some embodiments, the inflammation is associated with rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, sepsis, erythema nodosum leprosum, multiple sclerosis, psoriasis, systemic lupus erythematosis, type I diabetes, atherosclerosis,
encephalomyelitis, Alzheimer's disease, stroke, traumatic brain injury, Parkinson's disease or septic shock. In some embodiments, the therapeutic agent is an anti-inflammatory agent. In some embodiments, the therapeutic agent is a steroid. [0013] Described herein, in certain embodiments, are methods for treatment of an autoimmune disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a selective delivery molecule conjugate provided herein, thereby treating the autoimmune disease. In some embodiments, the autoimmune disease is Celiac disease, diabetes mellitus type 1, Sarcoidosis, systemic lupus erythematosus (SLE), Sjogren's syndrome, Churg- Strauss Syndrome, Hashimoto's thyroiditis, Graves' disease, idiopathic thrombocytopenic purpura, Addison's Disease, rheumatoid arthritis (RA), Polymyositis (PM), or Dermatomyositis (DM).
[0014] Described herein, in certain embodiments, are methods for delivering a therapeutic agent or an imaging agent to a cancer cell in a subject, comprising administering to the subject having cancer a selective delivery molecule conjugate provided herein or a selective delivery molecule provided herein, thereby delivering a therapeutic agent or an imaging agent to a cancer cell. In some embodiments, the methods further comprise imaging the cancer.
[0015] Described herein, in certain embodiments, are methods for delivering a therapeutic agent or an imaging agent to a site of inflammation in a subject, comprising administering to the subject having inflammation a selective delivery molecule conjugate provided herein or a selective delivery molecule provided herein, thereby delivering a therapeutic agent or an imaging agent to the site of inflammation. In some embodiments, the methods further comprise imaging the site of
inflammation.
BRIEF DESCRIPTION OF THE DRAWINGS
[0016] Figure 1: Cleavage of SDM-145 by hMMP-9. A) HPLC chromatogram of SDM-145; B) HPLC chromatogram of the reaction mixture after incubation at 37 °C for 17 h. LC-MS confirmed that the molecular weight of the peak at ~9.4 min was consistent with the fragment generated by hMMP-9 cleavage at the expected cleavage site. The fragment's chemical structure was shown in the chromatogram.
[0017] Figure 2: Cleavage of SDM-145 by Cathepsin B. A) HPLC chromatogram of conjugate SDM-145 ; B) HPLC chromatogram of the reaction mixture after incubation at 37 °C for 17 h. LC- MS confirmed that the molecular weight of the peak at ~9.0 min was consistent with the freed doxorubicin.
[0018] Figure 3 illustrates schematics of exemplary protease activated antibody conjugates and protease cleavage steps.
[0019] Figure 4 illustrates a schematic of exemplary dual protease activated drug delivery conjugate and protease cleavage steps.
[0020] Figure 5 illustrates a higher resolution schematic of exemplary dual protease activated drug delivery conjugate. [0021] Figure 6 illustrates an exemplary scheme for protease cleavage and uptake of a dual protease activated drug delivery conjugate: (a) extracellular proteases (e.g. matrix
metalloproteinases) cleave conjugate near target cells, (b) cell penetrating peptide (CPP)-drug portion enters target cells, (c) lysosomal proteases (e.g. Cathepsin B) cleave linker, and (d) active therapeutic cargo (e.g. Doxorubicin) is released.
[0022] Figure 7 illustrates a schematic of exemplary thiol-reactive drug delivery conjugates. The SDMs react efficiently with albumin Cys(34)-SH in circulation albumin after injected into blood stream. Albumin-SDM conjugates have improved pharmacokinetic profiles and efficient targeted cargo delivery.
[0023] Figure 8 illustrates MALDI-TOF Spectrum of SDM-147.
[0024] Figure 9 illustrates therapeutic activity of SDM-147 in a 4T1 breast cancer mouse model.
DETAILED DESCRIPTION OF THE INVENTION
Certain Terminology
[0025] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of skill in the art to which the claimed subject matter belongs. All patents, patent applications, published applications and publications, GENBANK sequences, websites and other published materials referred to throughout the entire disclosure herein, unless noted otherwise, are incorporated by reference in their entirety. In the event that there is a plurality of definitions for terms herein, those in this section prevail. Where reference is made to a URL or other such identifier or address, it is understood that such identifiers can change and particular information on the internet can come and go, but equivalent information is known and can be readily accessed, such as by searching the internet and/or appropriate databases. Reference thereto evidences the availability and public dissemination of such information. Generally, the procedures for cell culture, cell infection, antibody production and molecular biology methods are methods commonly used in the art. Such standard techniques can be found, for example, in reference manual, such as, for example, Sambrook et al. (2000) and Ausubel et al. (1994).
[0026] As used herein, the singular forms "a," "an" and "the" include plural referents unless the context clearly dictates otherwise. In this application, the use of the singular includes the plural unless specifically stated otherwise. As used herein, the use of "or" means "and/or" unless stated otherwise. Furthermore, use of the term "including" as well as other forms (e.g., "include", "includes", and "included") is not limiting.
[0027] As used herein, ranges and amounts can be expressed as "about" a particular value or range. About also includes the exact amount. Hence "about 40 mg" means "about 40 mg" and also "40 mg." Generally, the term "about" includes an amount that would be expected to be within experimental error.
[0028] The terms "individual," "patient," or "subject" are used interchangeably. As used herein, they mean any mammal (i.e. species of any orders, families, and genus within the taxonomic classification animalia: chordata: vertebrata: mammalia). In some embodiments, the mammal is a human. None of the terms require or are limited to situation characterized by the supervision (e.g. constant or intermittent) of a health care worker (e.g. a doctor, a registered nurse, a nurse practitioner, a physician's assistant, an orderly, or a hospice worker).
[0029] An "alkyl" group refers to an aliphatic hydrocarbon group. The alkyl moiety may be a saturated alkyl or an unsaturated alkyl. Depending on the structure, an alkyl group can be a monoradical or a diradical (i.e., an alkylene group).
[0030] The "alkyl" moiety may have 1 to 10 carbon atoms (whenever it appears herein, a numerical range such as "1 to 10" refers to each integer in the given range; e.g., "1 to 10 carbon atoms" means that the alkyl group may consist of 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and including 10 carbon atoms, although the present definition also covers the occurrence of the term "alkyl" where no numerical range is designated). The alkyl group could also be a "lower alkyl" having 1 to 6 carbon atoms. The alkyl group of the compounds described herein may be designated as "C1-C4 alkyl" or similar designations. By way of example only, "C1-C4 alkyl" indicates that there are one to four carbon atoms in the alkyl chain, i.e., the alkyl chain is selected from: methyl, ethyl, propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, and t-butyl. Typical alkyl groups include, but are in no way limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tertiary butyl, pentyl, hexyl, ethenyl, propenyl, butenyl, and the like.
[0031] In some embodiments, the linker comprises a ring structure (e.g., an aryl). As used herein, the term "ring" refers to any covalently closed structure. Rings include, for example, carbocycles (e.g., aryls and cycloalkyls), heterocycles (e.g., heteroaryls and non-aromatic heterocycles), aromatics (e.g. aryls and heteroaryls), and non-aromatics (e.g., cycloalkyls and non-aromatic heterocycles). Rings can be optionally substituted. Rings can be monocyclic or polycyclic.
[0032] As used herein, the term "aryl" refers to an aromatic ring wherein each of the atoms forming the ring is a carbon atom. Aryl rings can be formed by five, six, seven, eight, nine, or more than nine carbon atoms. Aryl groups can be optionally substituted. Examples of aryl groups include, but are not limited to phenyl, naphthalenyl, phenanthrenyl, anthracenyl, fluorenyl, and indenyl. Depending on the structure, an aryl group can be a monoradical or a diradical (i.e., an arylene group). [0033] The term "cycloalkyl" refers to a monocyclic or polycyclic non-aromatic radical, wherein each of the atoms forming the ring (i.e. skeletal atoms) is a carbon atom. Cycloalkyls may be saturated, or partially unsaturated. Cycloalkyl groups include groups having from 3 to 10 ring atoms. Cycloalkyls include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.
[0034] In some embodiments, the ring is a cycloalkane. In some embodiments, the ring is a cycloalkene.
[0035] In some embodiments, the ring is an aromatic ring. The term "aromatic" refers to a planar ring having a delocalized π-electron system containing 4n+2 π electrons, where n is an integer.
Aromatic rings can be formed from five, six, seven, eight, nine, or more than nine atoms.
Aromatics can be optionally substituted. The term "aromatic" includes both carbocyclic aryl (e.g., phenyl) and heterocyclic aryl (or "heteroaryl" or "heteroaromatic") groups (e.g., pyridine). The term includes monocyclic or fused-ring polycyclic (i.e., rings which share adjacent pairs of carbon atoms) groups.
[0036] In some embodiments, the ring is a heterocycle. The term "heterocycle" refers to heteroaromatic and heteroalicyclic groups containing one to four heteroatoms each selected from O, S and N, wherein each heterocyclic group has from 4 to 10 atoms in its ring system, and with the proviso that the ring of said group does not contain two adjacent O or S atoms. Non-aromatic heterocyclic groups include groups having only 3 atoms in their ring system, but aromatic heterocyclic groups must have at least 5 atoms in their ring system. The heterocyclic groups include benzo-fused ring systems. An example of a 3-membered heterocyclic group is aziridinyl. An example of a 4-membered heterocyclic group is azetidinyl (derived from azetidine). An example of a 5-membered heterocyclic group is thiazolyl. An example of a 6-membered heterocyclic group is pyridyl, and an example of a 10-membered heterocyclic group is quinolinyl. Examples of non- aromatic heterocyclic groups are pyrrolidinyl, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, dihydropyranyl, tetrahydrothiopyranyl, piperidino, morpholino, thiomorpholino, thioxanyl, piperazinyl, azetidinyl, oxetanyl, thietanyl, homopiperidinyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl, thiazepinyl, 1,2,3,6-tetrahydropyridinyl, 2-pyrrolinyl, 3-pyrrolinyl, indolinyl, 2H-pyranyl, 4H-pyranyl, dioxanyl, 1,3-dioxolanyl, pyrazolinyl, dithianyl, dithiolanyl, dihydropyranyl, dihydrothienyl, dihydrofuranyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, 3- azabicyclo[3.1.0]hexanyl, 3-azabicyclo[4.1.0]heptanyl, 3H-indolyl and quinolizinyl. Examples of aromatic heterocyclic groups are pyridinyl, imidazolyl, pyrimidinyl, pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyridinyl, and furopyridinyl. The foregoing groups, may be C-attached or reattached where such is possible. For instance, a group derived from pyrrole may be pyrrol- 1-yl (reattached) or pyrrol-3-yl (C-attached). Further, a group derived from imidazole may be imidazol-1- yl or imidazol-3-yl (both N-attached) or imidazol-2-yl, imidazol-4-yl or imidazol-5-yl (all C- attached). The heterocyclic groups include benzo-fused ring systems and ring systems substituted with one or two oxo (=0) moieties such as pyrrolidin-2-one. Depending on the structure, a heterocycle group can be a monoradical or a diradical (i.e., a heterocyclene group).
[0037] In some embodiments, the ring is fused. The term "fused" refers to structures in which two or more rings share one or more bonds. In some embodiments, the ring is a dimer. In some embodiments, the ring is a trimer. In some embodiments, the ring is a substituted.
[0038] The term "carbocyclic" or "carbocycle" refers to a ring wherein each of the atoms forming the ring is a carbon atom. Carbocycle includes aryl and cycloalkyl. The term thus distinguishes carbocycle from heterocycle ("heterocyclic") in which the ring backbone contains at least one atom which is different from carbon (i.e., a heteroatom). Heterocycle includes heteroaryl and
heterocycloalkyl. Carbocycles and heterocycles can be optionally substituted.
[0039] In some embodiments, the linker is substituted. The term "optionally substituted" or "substituted" means that the referenced group may be substituted with one or more additional group(s) individually and independently selected from Ci-Cealkyl, C3-Cscycloalkyl, aryl, heteroaryl, C2-C6heteroalicyclic, hydroxy, Ci-Cealkoxy, aryloxy, Ci-C6alkylthio, arylthio, Ci- C6alkylsulfoxide, arylsulfoxide, Ci-C6alkylsulfone, arylsulfone, cyano, halo, C2-C8acyl, C2- Csacyloxy, nitro, Ci-Cehaloalkyl, Ci-Cefluoroalkyl, and amino, including Ci-Cealkylamino, and the protected derivatives thereof. By way of example, an optional substituents may be Ls s, wherein each Ls is independently selected from a bond, -0-, -C(=0)-, -S-, -S(=0)-, -S(=0)2-, -NH-, - NHC(=0)-, -C(=0)NH-, S(=0)2NH-, -NHS(=0)2-, -OC(=0)NH-, -NHC(=0)0-, -(Ci-C6alkyl)-, or -(C2-C6alkenyl)-; and each Rs is independently selected from H, (Ci-C4alkyl), (Cs-Cscycloalkyl), heteroaryl, aryl, and Ci-Ceheteroalkyl. Optionally substituted non-aromatic groups may be substituted with one or more oxo (=0). The protecting groups that may form the protective derivatives of the above substituents are known to those of skill in the art.
Overview:
[0040] Selective delivery molecules (SDMs) allow the targeted delivery of therapeutic agents and/or imaging agents to specific cells and/or tissues. In certain embodiments, selective delivery molecules comprise (a) an acidic sequence (portion of A) which is effective to inhibit or prevent uptake of the molecule into cells or tissue retention, (b) a molecular transport or retention sequence (portion of B) (e.g., a cell penetrating peptide (CPP), (c) a cleavable linker X located between portion of A and portion of B, (d) at least one cargo moiety (portion D) bound to portion of B, and optionally (e) a macromolecular carrier bound to portion of A. In some embodiments, cleavage of the X linker allows the separation of portion of A from portion of B, thereby promoting the uptake or retention of portion of B and the attached cargo into cells or tissue retention. In some embodiments, the therapeutic cargo is a chemotherapeutic agent. In some embodiments, the therapeutic cargo is a cytotoxin.
[0041] As described herein, in certain instances, conjugating a selective delivery molecule disclosed herein to a targeting ligand, such as an antibody, allows the SDM to be targeted to specific cells having a specific cell surface marker. Similarly, as described herein, in certain instances, the cell specific delivery of an existing targeting ligand-drug conjugate (e.g., an antibody-drug conjugate) is improved by attachment of a selective delivery molecule described herein to the targeting ligand. In some embodiments, the targeting antibody binds to a tumor antigen or a receptor that is upregulated in tumor cells. Accordingly, provided herein are improved antibody-drug conjugates for delivery of therapeutic agents to target cells and tissues, such as cancer cells and other diseased cells.
[0042] In some embodiments, the targeting ligand-conjugated SDMs described herein provide significant advantages over existing targeting ligand-drug conjugates (e.g., antibody-drug conjugates). In some embodiments, the targeting ligand-conjugated SDMs described herein provide improved tumor penetration and retention over existing targeting ligand -drug conjugates. In some embodiments, the targeting ligand -conjugated SDMs described herein provide dual targeting specificity. In some embodiments, the dual targeting mechanism comprises 1) ligand (e.g., antibody) targeting of cell specific markers on diseased cells and 2) pathological protease activity targeting of increased extracellular protease activity at physiological location of the diseased cell. In some embodiments, the targeting ligand-conjugated SDMs described herein provide different options for conjugating a variety of therapeutic cargo molecules for delivery to the diseased cell. For example, in certain embodiments, a variety of configurations is available for the attachment of the therapeutic cargo to the SDM or the ligand (e.g., antibody) itself. Figure 3 illustrates exemplary configurations of targeting ligand-conjugated SDMs, where the targeting ligand is an antibody. In some embodiments, the therapeutic cargo is internalized with the targeting ligand. In some embodiments, the therapeutic cargo is released from the targeting ligand-conjugated SDM prior to uptake by the cell. In addition, the modular design of the targeting ligand-conjugated SDMs described herein enables easy modification of the conjugates to change protease recognition sites and therapeutic cargo molecules.
[0043] In some embodiments, increased retention and penetration of the cells achieved by the SDM conjugates provided herein increase the efficacy of the therapeutic cargo. In some embodiments, a lower dosage of the therapeutic cargo is employed compared to existing targeting ligand-drug conjugates that lack an SDM. In some embodiments, a less toxic therapeutic cargo is needed to treat the target cell. In some embodiments, increased retention and penetration of the cells achieved by the conjugates provided herein allows for less toxic therapeutic cargo molecules to be employed compared to existing targeting ligand-drug conjugates that lack an SDM.
[0044] Described herein, in certain embodiments are SDMs that provide single protease targeting. For example, in some embodiments, cleavage of the X linker located between portion of A and portion of B of the SDM by a protease located near the target cell allows the separation of portion of A from portion of B, thereby promoting the uptake or retention of portion of B and the attached cargo into cells or tissue retention. In some embodiments, the protease exhibits higher expression in the extracellular area surrounding the target cell (e.g., a cancer cell) as compared to a non-target cell (e.g., non-diseased/non-cancerous cell). In some embodiments, the protease is a matrix metalloproteinase (MMP). In some embodiments, the therapeutic cargo is a
chemotherapeutic agent. In some embodiments, the therapeutic cargo is a cytotoxin. In some embodiments, the therapeutic cargo is doxorubicin, calicheamicin, maytansinoid, or auritstatin. In some embodiments, the therapeutic cargo is an anti-inflammatory agent. In some embodiments, the therapeutic cargo is a steroid. In some embodiments, the therapeutic cargo is cortisone or a derivative thereof. In some embodiments, the SDM is conjugated to a targeting ligand.
[0045] Described herein, in certain embodiments are SDMs that provide double protease targeting. For example, in some embodiments, the SDM comprises an X linker located between portion of A and portion of B and a second linker Y located between portion of B and the therapeutic cargo molecule (portion D). In some embodiments, the X linker comprises an extracellular protease cleavage site and the Y linker comprises an intracellular protease cleavage site. In some embodiments, cleavage of the X linker located between portion of A and portion of B of the selective delivery molecule by a protease located near the target cell allows the separation of portion of A from portion of B, thereby promoting the uptake or retention of portion of B and the attached cargo into cells or tissue retention, and then cleavage of the Y linker by an intracellular protease allows the therapeutic cargo to be released into the cell after uptake. In some
embodiments, such configurations allow therapeutic cargos to be maintained in an inactive state until the therapeutic cargos are taken up by the cell following cleavage of the X linker. In some embodiments, the extracellular protease that cleaves the X linker exhibits a higher expression in the extracellular area surrounding the target cell (e.g., a cancer cell) as compared to a non-target cell (e.g., non-diseased/non-cancerous cell). In some embodiments, the extracellular protease is a matrix metalloproteinase (MMP). In some embodiments, the intracellular protease that cleaves the Y linker is a lysosomal protease. In some embodiments, the protease is one that is activated at low pH. In some embodiments, the protease is one that is activated at low pH and is expressed in an endosome. In some embodiments, the protease is a cathepsin. In some embodiments, the cathepsin is cathepsin B or cathepsin D. In some embodiments, the cathepsin is cathepsin B. In some embodiments, the Y linker additionally comprises a self-immolative cleavage site located between the intracellular protease cleavage site and the attached therapeutic cargo. In some embodiments, the self-immolative cleavage site is a PABC (p-aminobenzylcarbonyl) spacer and analogs thereof. In some embodiments, the self-immolative cleavage site is a thiazole containing linker. In some embodiments, the selective delivery molecule comprising an X linker located between portion of A and portion of B and a second linker Y located between portion of B and the therapeutic cargo molecule (portion D) is conjugated to a targeting ligand, such as a targeting antibody. In some embodiments, the targeting ligand binds to a tumor antigen or a receptor that is upregulated in tumor cells. In some embodiments, the SDM is conjugated to a targeting ligand. Figures 4 and 5 illustrate schematics of exemplary dual protease drug delivery conjugates. Figure 6 illustrates delivery of an antibody-SDM conjugate with dual protease targeting.
[0046] In some embodiments, the SDMs provided herein are conjugated to albumin and/or contain a free thiol reactive group for interacting with albumin in vivo. Albumin is a carrier for tumor targeting because it accumulates in solid tumors due to the pathophysiology of tumor tissue, characterized by a high metabolic turnover, angiogenesis, hypervasculature, a defective vascular architecture and an impaired lymphatic drainage. The unique free sulfhydryl group (Cys-34) of albumin, which is not present in the majority of circulating serum proteins, is accessible for selective modifications. Albumin-drug conjugates show improved the pharmacokinetic profiles. However, albumin conjugates have limited tumor penetration and distribution due to their big molecular size and the tumor tissue's microenvironment, such as increased interstitial fluid pressure and dense extracellular matrix. In some embodiments, thiol-reactive SDMs provided herein form albumin conjugates in vivo. In some embodiments, the conjugates increase the drug's tumor penetration. In some embodiments, the conjugates increase improve drug's distribution and activity. In some embodiments, after injected into blood stream, thiol-reactive SDMs react with the free Cys34 thiol of the circulating albumin. The albumin-SDM conjugate is then transported and accumulated in the tumor tissues. The up-regulated MMPs in tumor tissues cleave the MMPs sensitive linker in ACPP part and release the poly-Arg-drug fragment. As poly-Arg has excellent cell penetrating capability, the poly-Arg-drug fragment is able to efficiently bind to the tumor cell and get internalized. After internalization, drug is regenerated after an enzymatic cleavage by intracellular proteases, such as Cathepsin B in the lysosome. In some embodiments, the thiol reactive group of the SDM is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. Figure 7 illustrates exemplary schematics of exemplary thiol- reactive SDMs.
Conjugated Selective Delivery Molecules (SDMs)
[0047] Disclosed herein, in certain embodiments, and carrier-conjugated SDMs. In some embodiments, a carrier modulates plasma half-life of a selective delivery molecule disclosed herein. In some embodiments, a carrier modulates solubility of a selective delivery molecule disclosed herein. In some embodiments, a carrier modulates bio-distribution of a selective delivery molecule disclosed herein.
[0048] In some embodiments, a carrier decreases uptake of a selective delivery molecule by non- target cells or tissues. In some embodiments, a carrier decreases uptake of a selective delivery molecule into cartilage. In some embodiments, a carrier decreases uptake of a selective delivery molecule into joints relative to target tissue.
[0049] In some embodiments, a carrier increases uptake of a selective delivery molecule by target cells or tissues. In some embodiments, a carrier decreases uptake of a selective delivery molecule into the liver relative to target tissue. In some embodiments, a carrier decreases uptake of a selective delivery molecule into kidneys. In some embodiments, a carrier enhances uptake into cancer tissue. In some embodiments, a carrier enhances uptake into lymphatic channels and/or lymph nodes.
[0050] In some embodiments, a carrier increases plasma half-life by reducing glomerular filtration. In some embodiments, a carrier modulates plasma half-life by increasing or decreases metabolism or protease degradation. In some embodiments, a carrier increases tumor uptake due to enhanced permeability and retention (EPR) of tumor vasculature. In some embodiments, a carrier increases the aqueous solubility of selective delivery molecule.
[0051] In some embodiments, any carrier is independently directly or indirectly (e.g., via CM) bound to A, B, or X. In some embodiments, any carrier is independently bound to A at the n- terminal poly glutamate. In some embodiments, any carrier is independently bound to A (or, the n- terminal poly glutamate) by a covalent linkage. In some embodiments, any carrier is independently bound to B at the c-terminal polyarginine. In some embodiments, any carrier is independently bound to B (or, the c-terminal polyarginine) by a covalent linkage. In some embodiments, any carrier is independently directly or indirectly bound to linkers between X and A, X and B, B and C N terminus, and A and C/N terminus. In some embodiments, the covalent linkage comprises an ether bond, thioether bond, amine bond, amide bond, oxime bond, carbon-carbon bond, carbon- nitrogen bond, carbon-oxygen bond, or carbon-sulfur bond.
[0052] In some embodiments, carrier is selected from a macromolecule such as a protein, a synthetic or natural polymer, or a dendrimer. In some embodiments, carrier is selected from dextran, a PEG polymer (e.g., a PEG polymer having an average molecular weight of
approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa),
approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)), albumin, or a combination thereof. In some embodiments, carrier is a PEG polymer.
[0053] In some embodiments, the size of carrier is between about 50kDa and about 70kDa.
[0054] In some embodiments, the selective delivery molecule is conjugated to albumin. In certain instances, albumin is excluded from the glomerular filtrate under normal physiological conditions. In some embodiments, the selective delivery molecule comprises a reactive group such as maleimide that can form a covalent conjugate with albumin. A selective delivery molecule comprising albumin results in enhanced accumulation of cleaved selective delivery molecules in tumors in a cleavage dependent manner. In some embodiments, albumin conjugates have good pharmacokinetic properties.
[0055] In some embodiments, the selective delivery molecule is conjugated to PEG polymers. In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately IkDa (PEG IkDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 2kDa (PEG 2kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 3kDa (PEG 3kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 4kDa (PEG 4kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 5kDa (PEG 5kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately lOkDa (PEG lOkDa). In some embodiments, the selective delivery molecule is conjugated PEG polymers having an average molecular weight of approximately 12 kDa ( PEG 12kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 15kDa (PEG 15kDa). In some embodiments, selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 20 kDa (PEG 20kDa). In some embodiments, selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 30 kDa (PEG 30kDa). In some embodiments, selective delivery molecules conjugated to PEG30kDa had a longer half-life as compared to free peptides. In some
embodiments, selective delivery molecules are conjugated to PEG polymers having an average molecular weight of between about 20 to about 40kDa which have hepatic and renal clearance.
[0056] In some embodiments, the selective delivery molecule is conjugated to a dextran. In some embodiments, the selective delivery molecule is conjugated to a dextran having an average molecular weight of approximately 70kDa. In some embodiments, dextran conjugates, being a mixture of molecular weights, are difficult to synthesize and purify reproducibly.
[0057] In some embodiments, the selective delivery molecule is conjugated to streptavidin.
[0058] In some embodiments, the selective delivery molecule is conjugated to a fifth generation PAMAM dendrimer.
[0059] In some embodiments, a carrier is capped. In some embodiments, capping a carrier improves the pharmacokinetics and reduces cytotoxicity of a carrier by adding hydrophilicity. In some embodiments, the cap is selected from: Acetyl, succinyl, 3-hydroxypropionyl, 2-sulfobenzoyl, glycidyl, PEG-2, PEG-4, PEG-8 and PEG-12.
[0060] Disclosed herein, in certain embodiments, are targeting ligand-conjugated SDMs. In some embodiments, a therapeutic cargo and an SDM are conjugated to a targeting ligand. In some embodiments, the SDM comprises a therapeutic cargo and the SDM comprising the therapeutic cargo is conjugated to a targeting ligand. Thus, in some embodiments, the therapeutic cargo and the SDM are conjugated to the same site on the targeting ligand. In some embodiments, the therapeutic cargo is first conjugated to an SDM, and then the SDM comprising the therapeutic cargo is attached to the targeting ligand.
[0061] In some embodiments, the therapeutic cargo and the SDM are conjugated to two different sites on the targeting ligand. In some embodiments, the SDM is conjugated to an existing ligand- drug conjugate. In some embodiments, an SDM is conjugated to a targeting ligand, and then a therapeutic cargo is conjugated to the targeting ligand -SDM conjugate. In some embodiments, a therapeutic cargo is conjugated to a targeting ligand, and then an SDM is conjugated to the targeting ligand-therapeutic cargo conjugate.
[0062] In some embodiments, the targeting ligand is conjugated to the acidic sequence (portion of A) of an SDM.
[0063] In some embodiments, the targeting ligand is conjugated to molecular transport or retention sequence (portion of B) of an SDM.
[0064] In some embodiments, any of a variety of known methods for conjugation of molecules to polypeptides such as targeting ligands are employed for the conjugation of the therapeutic cargo and/or SDMs provided herein.
[0065] Disclosed herein, in certain embodiments, are antibody-conjugated SDMs. In some embodiments, a therapeutic cargo and an SDM are conjugated to a targeting antibody. In some embodiments, the SDM comprises a therapeutic cargo and the SDM comprising the therapeutic cargo is conjugated to a targeting antibody. Thus, in some embodiments, the therapeutic cargo and the SDM are conjugated to the same site on the targeting antibody (e.g., Form 1 and Form 2 of Fig. 3). In some embodiments, the therapeutic cargo is first conjugated to an SDM, and then the SDM comprising the therapeutic cargo is attached to the targeting antibody.
[0066] In some embodiments, the therapeutic cargo and the SDM are conjugated to two different sites on the targeting antibody (e.g., Form 3 of Fig. 3). In some embodiments, the SDM is conjugated to an existing antibody-drug conjugate. In some embodiments, an SDM is conjugated to a targeting antibody, and then a therapeutic cargo is conjugated to the antibody-SDM conjugate. In some embodiments, a therapeutic cargo is conjugated to a targeting antibody, and then an SDM is conjugated to the antibody-therapeutic cargo conjugate.
[0067] In some embodiments, the targeting antibody is conjugated to the acidic sequence (portion of A) of an SDM (Form 1, Fig. 3).
[0068] In some embodiments, the targeting antibody is conjugated to molecular transport or retention sequence (portion of B) of an SDM (Form 2 and Form 3, Fig. 3).
[0069] In some embodiments, any of a variety of known methods for conjugation of molecules to antibodies are employed for the conjugation of the therapeutic cargo and/or SDMs provided herein. Targeting Ligands
[0070] In some embodiments, the targeting ligand is a molecule that binds to a cell surface molecule expressed on the surface of a target cell. In some embodiments, the targeting ligand binds to a receptor expressed on the surface of a target cell. In some embodiments, the targeting ligand binds to a cell surface antigen expressed on the surface of a target cell. In some embodiments, the targeting ligand binds to a carbohydrate, a polypeptide or glycoprotein expressed on the surface of a target cell. In some embodiments, the targeting ligand is a lectin or an integrin. In some embodiments, the targeting ligand is an antibody. In some embodiments, the targeting ligand is a targeting non-antibody. In some embodiments, the targeting ligand is a co-stimulatory molecule.
[0071] In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule of a diseased cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule that is specific to the diseased cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the diseased cell compared to non- diseased cells. In some embodiments, the targeting ligand is an antibody. In some embodiments, the diseased cell is a cancer cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule expressed by a hematopoietic cell. In some embodiments, the targeting ligand targets an inflammatory cell (e.g., neutrophil, macrophage, monocyte, eosinophil, basophil). In some embodiments, the targeting ligand targets a cell involved in an autoimmune disease (e.g. a lymphocyte, such as a B lymphocyte or a T lymphocyte).
[0072] In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule of a cancer cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule that is specific to the cancer cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the cancer cell compared to non-cancer cells. In some embodiments, targeting ligand is a ligand that binds to a cell surface molecule that that is expressed by the cancer cell but not a non-cancer cell. In some embodiments, the targeting ligand is a ligand that binds to a tumor antigen. In some embodiments, the targeting ligand is a ligand that binds to a cell surface receptor that is upregulated in the tumor cell (i.e., has a higher expression) compared to a non-tumor cell. In some embodiments, the targeting ligand is a ligand that binds to a cell surface receptor that that is expressed by the tumor cell but not a non-tumor cell. In some embodiments, the targeting ligand is an antibody.
[0073] In some embodiments, the targeting antibody used in the compositions and methods provided herein is an antibody that binds to a cell surface molecule on the targeted cell. In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule that is specific for the targeted cell. In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the target cell compared to non-targeted cells. [0074] In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule of a diseased cell. In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule that is specific to the diseased cell. In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the diseased cell compared to non- diseased cells.
[0075] In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule of a cancer cell. In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule that is specific to the cancer cell. In some embodiments, the targeting antibody is an antibody that binds to a cell surface molecule that is upregulated (i.e., has a higher expression) on the cancer cell compared to non-cancer cells. In some embodiments, the targeting antibody binds to a cell surface molecule that that is expressed by the cancer cell but not a non- cancer cell. In some embodiments, the antibody binds to a tumor antigen. In some embodiments, the targeting antibody binds to a cell surface receptor that is upregulated in the tumor cell (i.e., has a higher expression) compared to a non-tumor cell. In some embodiments, the targeting antibody binds to a cell surface receptor that that is expressed by the tumor cell but not a non-tumor cell.
[0076] In some embodiments, the targeting antibody is a tumor specific antibody. In some embodiments, the targeting antibody binds to CD3, CD19, CD20, CD22, CD25, CD30, CD33, CD52, interleukin-2 receptor (IL-2), HLA-DR10p, tenascin, CEA, MUC1, TAG72, EBBB2 receptor (HER2), CD56 (NCAM), CS-125, Cripto, glycoprotein NMB (osteoactivin), CD70, prostate specific membrane antigen (PSMA), SLC44A4 (AGS-5), folate receptor, an integrin, such as ανβ3 -integrin, transferrin receptor, granulocyte-macrophage colony-stimulating factor (GM- CSF) receptor, aminopeptidase N (CD 13), galactosamine receptor, leutenizing hormone releasing hormone (LHRH) receptor, vascular endothelial growth factor (VEGF) receptor (FLK1), ROR1, mesothelin, CD33/IL3Ra, c-Met; PSMA, Glycolipid F77, EGFRvllI, GD-2, NY-ESO- 1 TCR, MAGE A3 TCR.
[0077] In some embodiments, the targeting antibody is gemtuzumab, inotuumab, trastuzumab (Herceptin), HD37, M195, LMB2, lyml, 81C6, HMFG1, CC49, rituximab, epratuzumab, lorvotuzumab, 2C3, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN-75, PSMA ADC, ASG-5ME or mdx-1203. In some embodiments, the targeting antibody is a variant of gemtuzumab, inotuumab, trastuzumab (Herceptin), HD37, M195, LMB2, lyml, 81C6, HMFG1, CC49, rituximab, epratuzumab, lorvotuzumab, 2C3, imgn388, SAR3419, ΒΪ1Β062, brentixumab, glembatumumab, SGN-75, PSMA ADC, ASG-5ME or mdx-1203. In some embodiments, the comprises an antigen-binding fragment of gemtuzumab, inotuumab, trastuzumab (Herceptin), HD37, M195, LMB2, lyml, 81C6, HMFG1, CC49, rituximab, epratuzumab, lorvotuzumab, 2C3, imgn388, SAR3419, ΒΪ1Β062, brentixumab, glembatumumab, SGN-75, PSMA ADC, ASG-5ME or mdx-1203.
[0078] In some embodiments, the targeting ligand is a non-antibody ligand that binds to a receptor. In some embodiments, the targeting ligand binds to folate receptor, avp3-integrin, transferrin receptor, GM-CSF receptor, aminopeptidase N (CD 13), galactosamine receptor and LHRH receptor. In some embodiments, the targeting ligand comprises RGD, NGR, folate, transferrin, GM-CSF, or galactosamine.
[0079] In some embodiments, the targeting antibody is natural antibody. In some embodiments, the targeting antibody is a synthetic antibody. In some embodiments, the targeting antibody is a recombinant antibody. In some embodiments, the targeting antibody is an antibody fragment containing at least a portion of the variable region of the immunoglobulin molecule that retains the binding specificity ability of the full-length immunoglobulin. In some embodiments, the targeting antibody is any protein having a binding domain that is homologous or substantially homologous to an immunoglobulin antigen-binding domain (antibody combining site). In some embodiments, the targeting antibody is a multispecific antibodies (e.g., bispecific antibodies). In some embodiments, the targeting antibody is a human antibody or non-human antibody. In some embodiments, the targeting antibody is a humanized antibody. In some embodiments, the targeting antibody is a chimeric antibody. In some embodiments, the targeting antibody is an intrabody. In some embodiments, the targeting antibody is an antibody fragment, such as, but not limited to, Fab fragment, Fab' fragment, F(ab')2 fragment, Fv fragment, disulfide-linked Fv (dsFv), Fd fragment, Fd' fragment, single-chain Fv (scFv), single-chain Fab (scFab), diabody, anti-idiotypic (anti-Id) antibody, or antigen-binding fragments of any of the above In some embodiments, the targeting antibody is any member of any immunoglobulin type (e.g., IgG, IgM, IgD, IgE, IgA and IgY), any class (e.g. IgGl , IgG2, IgG3, IgG4, IgAl and IgA2) or subclass (e.g., IgG2a and IgG2b).
[0080] In some embodiments, the targeting antibody is a monoclonal antibody. In some embodiments, the targeting antibody is an IgG antibody. In some embodiments, the targeting antibody is a monovalent antibody. In some embodiments, the targeting antibody is multivalent antibody. In some embodiments, the targeting antibody is a bivalent antibody. In some
embodiments, the targeting antibody is an antibody fragment, such as a single-chain variable fragment (scFv). In some embodiments, the targeting antibody is a humanized antibody. In some embodiments, the targeting antibody is a variant of a known tumor specific antibody. In some embodiments, the targeting antibody is an antigen-binding fragment of a known tumor specific antibody. [0081] In some embodiments, the SDM and/or therapeutic cargo is conjugated to a portion of the antibody such that conjugation does not interfere with antibody-antigen binding. In some embodiments, the SDM and/or therapeutic cargo is conjugated to the Fc portion of the antibody. Selective delivery molecules
[0082] Disclosed herein, in certain embodiments, are selective delivery molecules (SDMs) for use in the compositions and methods described herein. In certain embodiments, the SDMs described herein are used alone or are conjugated to one or more molecules. In certain
embodiments, the SDMs described herein are conjugated to a targeting ligand. In some embodiments, the targeting ligand is an antibody. In certain embodiments, the SDMs described herein are conjugated to an existing antibody-drug conjugate. In certain embodiments, the SDMs described herein comprise a therapeutic cargo. In certain embodiments, the SDMs described herein comprising a therapeutic cargo are conjugated to a targeting ligand. In some embodiments, the targeting ligand is an antibody.
[0083] In certain embodiments, the SDM is an SDM of Formula I, having the structure:
Figure imgf000029_0001
Formula I
wherein,
X is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids;
CB comprises 0- 1 amino acid;
Db is a therapeutic agent; and
wherein [CB-Db] is bound to any amino acid of B. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some embodiments, CB is selected from a naturally-occurring amino acid or a non- naturally-occurring amino acid. In some embodiments, CB is selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a ΐ-amino acid. In some embodiments, cB is selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxyl amine or hydrazine group. In some embodiments, CB is selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some
embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQL (Ac). In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence RPLALWRS. In some embodiments, X comprises the amino acid sequence DPRSFL. In some embodiments, X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac). In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In certain embodiments, the SDM is an SDM comprising the structure of Formula I. In some embodiments, the selective delivery molecule of Formula I is: SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM- 110, SDM-1 1 1 , SDM- 1 12, SDM-1 13, SDM-114, SDM-115, SDM-116, SDM-1 17, SDM- 118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM- 129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141 , SDM-142, SDM-143, SDM-144, SDM-145, SDM- 146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151, SDM-152, and SDM-153.
[0084] In certain embodiments, the SDM is an SDM of Formula II, having the structure:
A-[CM-M]-X-B-[CB-DB]
Formula II
wherein,
X is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids; cB and cM each independently comprise 0-1 amino acid;
M is a macromolecule;
DB is a therapeutic agent; and
wherein [CM-M] is bound to at any position of A or X, and [CB-Db] is bound to any amino acid of B. In some embodiments, the therapeutic agent is cortisone. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some embodiments, CB and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CB and cM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a fi- amino acid, or a -amino acid. In some embodiments, CB and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxyl amine or hydrazine group. In some embodiments, CB and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by an extracellular protease. In some embodiments, X is cleavable by a soluble protease or cell surface associated protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALW S, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQL (Ac). In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence RPLALWRS. In some embodiments, X comprises the amino acid sequence
DPRSFL. In some embodiments, X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac). In some embodiments, M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a PEG polymer, albumin, or a combination thereof. In some embodiments, M is a PEG. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa),
approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)). In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In certain embodiments, the SDM is an SDM comprising the structure of Formula II. In some embodiments, the selective delivery molecule of Formula II is: SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM- 110, SDM-1 1 1 , SDM- 1 12, SDM-1 13, SDM-114, SDM-115, SDM-116, SDM-1 17, SDM- 118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM- 129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141 , SDM-142, SDM-143, SDM-144, SDM-145, SDM- 146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151, SDM-152, and SDM-153.
[0085] In certain embodiments, the SDM does not comprise a cargo molecule. In some embodiments, an SDM without a cargo molecule is conjugated to an existing antibody-drug conjugate (e.g. Form 3, Fig. 3). In certain embodiments, the SDM is an SDM of Formula III or IV, having the structure:
A-X-B
Formula III
A-[cM-M]-X-B
Formula IV
wherein,
X is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids; cM comprises 0-1 amino acid;
M is a macromolecule; and
wherein [CM-M] is bound to at any position of A or X. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some embodiments, CM is selected from a naturally-occurring amino acid or a non- naturally-occurring amino acid. In some embodiments, CM is selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a r-amino acid. In some embodiments, CM is selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxyl amine or hydrazine group. In some embodiments, CM is selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)- AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac). In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence RPLALWRS. In some embodiments, X comprises the amino acid sequence DPRSFL. In some embodiments, X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac). In some embodiments, M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a PEG polymer, albumin, or a combination thereof. In some embodiments, M is a PEG. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)). In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In certain embodiments, the SDM is an SDM comprising the structure of Formula III or IV. In some embodiments, the selective delivery molecule of Formula III or IV is: SDM- 101, SDM- 102, SDM- 103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM- 110, SDM-111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM-118, SDM-119, SDM- 120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM- 137, SDM-138, SDM-139, SDM-140, SDM-141, SDM-142, SDM-143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151, SDM-152, and SDM-153.
[0086] Dual protease Substrate SDMs
[0087] Described herein, in certain embodiments, is an SDM that comprises more than one protease cleavage site. In some embodiments, the SDM comprises a cleavage site for an
extracellular protease and a cleavage site for an intracellular protease. In some embodiments, the SDM comprises a cleavable linker X and a cleavable linker Y, where linker X comprises a cleavage site for an extracellular protease and linker Y comprises a cleavage site for an intracellular protease. In some embodiments, linker X is located between portion of A and portion of B, and linker Y is located between portion of B and the therapeutic cargo. In some embodiments, the intracellular protease that cleaves the Y linker is a lysosomal protease. In some embodiments, the protease is one that is activated at low pH. In some embodiments, the protease is one that is activated at low pH and is expressed in an endosome. In some embodiments, the protease is a cathepsin. In some embodiments, the cathepsin is cathepsin B. In some embodiments, the Y linker additionally comprises a self-immolative cleavage site located between the intracellular protease cleavage site and the attached therapeutic cargo. In some embodiments, the self-immolative cleavage site is a PABC (p-aminobenzylcarbonyl) spacer and its analogs thereof. In some embodiments, the self- immolative cleavage site is a thiazole containing linker.
[0088] In certain embodiments, the SDM is an SDM of Formula V, having the structure:
A-[CM-M]-X-B-Y-[CB-DB] Formula V
wherein,
X is a cleavable linker;
Y is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids;
CB and CM each independently comprise 0-1 amino acid;
M is a macromolecule;
DB is a therapeutic agent; and
wherein [CM-M] is bound to at any position of A or X, and [CB-Db] is bound to any amino acid of B. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some
embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some
embodiments, CB, and CM are each independently a 0-1 amino acid. In some embodiments, CB and CM are each independently selected from a naturally-occurring amino acid or a non-naturally- occurring amino acid. In some embodiments, CB and CM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a τ-amino acid. In some
embodiments, CB and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CB and CM are each independently selected from D-cysteine, D- glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CB is any amino acid having a free thiol group. In some embodiments, CB is D-cysteine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)- AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac). In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence RPLALWRS. In some embodiments, X comprises the amino acid sequence DPRSFL. In some embodiments, X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac). In some embodiments, M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a PEG polymer, albumin, or a combination thereof. In some embodiments, M is a PEG. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa),
approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), and approximately 40kDa (PEG 40kDa)). In some embodiments, Y is cleavable by a protease. In some embodiments, Y is cleavable by an intracellular protease. In some embodiments, Y comprises an amino acid sequence that is cleavable by Cathepsin B. In some embodiments, Y comprises the amino acid sequence Phe-Lys or Val-Cit (L-citrulline). In some embodiments, Y comprises a site for self-immolative cleavage. In some embodiments, Y comprises a PABC self-immolative spacer. In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In certain embodiments, the SDM is an SDM comprising the structure of Formula V. In some embodiments, the selective delivery molecule of Formula V is: SDM-101, SDM-102, SDM-103, SDM-104, SDM-105, SDM- 106, SDM-107, SDM-108, SDM-109, SDM-110, SDM-1 11, SDM-112, SDM-113, SDM-1 14, SDM-1 15, SDM-116, SDM-117, SDM-118, SDM-1 19, SDM-120, SDM-121 , SDM-122, SDM- 123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM- 140, SDM-141, SDM-142, SDM-143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151 , SDM-152, and SDM-153.
[0089] Other SDMs [0090] In certain embodiments, the SDM is an SDM comprising an imaging agent. In certain embodiments, the SDM is an SDM comprising an imaging agent and a therapeutic agent. In certain embodiments, the SDM is an SDM comprising two or more imaging agents or two or more therapeutic agents.
[0091] In certain embodiments, the SDM is an SDM comprising two or more imaging agents for Forster resonance energy transfer (FRET) imaging, where one imaging agent is conjugated to the A portion of the SDM and one imaging agent is conjugated to the B portion of the SDM.
[0092] In certain embodiments, the SDM is an SDM of Formula VI, having the structure:
[DA-CA]-A-[CM-M]-X-B-[CB-DB]
Formula VI
wherein,
X is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids;
CA, CB, and CM each independently comprise 0-1 amino acid;
M is a macromolecule; and
DA and DB are each independently selected from an imaging agent and a therapeutic; and wherein [CM-M] is bound to at any position of A or X, [DA-CA] is bound to any amino acid of A, and [CB-Db] is bound to any amino acid of B. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some embodiments, CA, CB, and CM are each independently a 0-1 amino acid. In some embodiments, CA, CB, and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CA, CB, and CM are each
independently selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a γ-amino acid. In some embodiments, CA, CB, and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CA, CB, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CB is any amino acid having a free thiol group. In some embodiments, CB is D- cysteine. In some embodiments, CA is any amino acid having a -terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CA is lysine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L- phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac). In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence RPLALWRS. In some embodiments, X comprises the amino acid sequence DPRSFL. In some embodiments, X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac). In some embodiments, M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer. In some embodiments, M is selected from dextran, a PEG polymer, albumin, or a combination thereof. In some embodiments, M is a PEG. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa),
approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)). In some embodiments, DA and DB are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other. In some embodiments, DA and DB are Cy5 and Cy7. In some embodiments, DA and DB are Cy5 and IRDye750. In some embodiments, DA and DB are Cy5 and IRDye800. In some embodiments, DA and DB are Cy5 and ICG. In some embodiments, DA and DB are a fluorescent moiety and a fluorescence-quenching moiety. In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In certain embodiments, the SDM is an SDM comprising the structure of Formula VI. In some embodiments, the molecule of Formula I is: SDM- 14, SDM- 15, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-32, or SDM-35.
[0093] In certain embodiments, the SDM is an SDM of Formula VI, having the structure:
[DA-CA]-A-[CM-M]-X-B-[CB-DB]
Formula VI
wherein,
X is a cleavable linker;
A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids;
CA, Cb, and cM each independently comprise 0-1 amino acid;
M is a polyethylene glycol (PEG) polymer; and
DA and DB are each independently an imaging agent; and
wherein [CM -M] is bound to at any position of A or X, [DA-CA] is bound to any amino acid of A, and [cB -DB] is bound to any amino acid of B. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, the number of basic amino acids in B is greater than the number of acidic amino acids in A. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates. In some embodiments, B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 or 9 consecutive glutamates and B is a peptide comprising 8 or 9 consecutive arginines. In some embodiments, A is a peptide comprising 5 consecutive glutamates and B is a peptide comprising 8 consecutive arginines. In some embodiments, CA, Cb, and CM are each independently a 0-1 amino acid. In some embodiments, CA, Cb, and cM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CA, Cb, and CM are each
independently selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a T-amino acid. In some embodiments, CA, CB, and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CA, CB, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CB is any amino acid having a free thiol group. In some embodiments, CB is D- cysteine. In some embodiments, CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CA is lysine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L- phenylalanine. In some embodiments, X is cleavable by a protease. In some embodiments, X is cleavable by a matrix metalloproteinase. In some embodiments, X comprises an amino acid sequence that is cleavable by MMP2, MMP7, MMP9, or MMP14. In some embodiments, X comprises a peptide linkage. In some embodiments, X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQL L, and RLQLK(Ac). In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence RPLALWRS. In some embodiments, X comprises the amino acid sequence DPRSFL. In some embodiments, X comprises the amino acid sequence PPRSFL. In some embodiments, X comprises the amino acid sequence RLQLKL. In some embodiments, X comprises the amino acid sequence RLQLK(Ac). In some embodiments, DA and DB are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other. In some embodiments, DA and DB are Cy5 and Cy7. In some embodiments, DA and DB are Cy5 and IRDye750. In some embodiments, DA and DB are Cy5 and IRDye800. In some embodiments, DA and DB are Cy5 and ICG. In some embodiments, DA and DB are a fluorescent moiety and a fluorescence-quenching moiety. In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In certain embodiments, the SDM is an SDM comprising the structure of Formula VI. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), and approximately 40kDa (PEG 40kDa)). In some embodiments, the molecule of Formula VI is: SDM-14, SDM-15, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-32; or SDM-35.
[0094] In certain embodiments, the SDM is an SDM of Formula VI, having the structure:
[DA-CA]-A-[CM-M]-X-B-[CB-DB]
Formula VI wherein,
X is a peptide linker cleavable by a matrix metalloproteinase;
A is a peptide with a sequence comprising 5 or 9 consecutive glutamates;
B is a peptide with a sequence comprising 8 or 9 consecutive arginines;
CA, CB, and cM each independently comprise 0-1 amino acid;
M is a polyethylene glycol (PEG) polymer; and
DA and DB are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other; and
wherein [CM -M] is bound to at any position of A or X, [DA-CA] is bound to any amino acid of A, and [CB -DB] is bound to any amino acid of B. In some embodiments, A and B do not have an equal number of acidic and basic amino acids. In some embodiments, CA, Cb, and cM are each
independently selected from any amino acid having a free thiol group, any amino acid having a N- terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CA, Cb, and CM are each independently a 0-1 amino acid. In some embodiments, CA, CB, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L- phenylalanine. In some embodiments, CB is any amino acid having a free thiol group. In some embodiments, CB is D-cysteine. In some embodiments, CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, DA and DB are Cy5 and Cy7. In some embodiments, DA and DB are Cy5 and Cy7. In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)). In certain embodiments, the SDM is an SDM comprising the structure of Formula VI.
[0095] In certain embodiments, the SDM is an SDM of Formula VI, having the structure:
[DA-CA]-A-[CM-M]-X-B-[CB-DB]
Formula VI
wherein,
X is a peptide linker cleavable by a matrix metalloproteinase;
A is a peptide with a sequence comprising 5 consecutive glutamates;
B is a peptide with a sequence comprising 8 consecutive arginines;
CA, Cb, and cM each independently comprise 0-1 amino acid;
M is a polyethylene glycol (PEG) polymer; and
DA and DB are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other; and
wherein [CM -M] is bound to at any position of A or X, [DA-CA] is bound to any amino acid of A, and [cB -DB] is bound to any amino acid of B. In some embodiments, CA, Cb, and cM are each independently a 0-1 amino acid. In some embodiments, CA, cB, and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CA, Cb, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, cB is any amino acid having a free thiol group. In some embodiments, cB is D- cysteine. In some embodiments, CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence
RPLALWRS. In some embodiments, DA and DB are Cy5 and Cy7. In some embodiments, DA and DB are Cy5 and IRDye750. In some embodiments, DA and DB are Cy5 and IRDye800. In some embodiments, DA and DB are Cy5 and ICG. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately IkDa (PEG IkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)). In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In certain embodiments, the SDM is an SDM comprising the structure of Formula VI.
[0096] In certain embodiments, the SDM is an SDM of Formula VI, having the structure:
[DA-CA]-A-[CM-M]-X-B-[CB-DB]
Formula VI
wherein,
X is a peptide linker cleavable by a matrix metalloproteinase;
A is a peptide with a sequence comprising 9 consecutive glutamates;
B is a peptide with a sequence comprising 9 consecutive arginines;
CA, CB, and CM each independently comprise 0-1 amino acid;
M is a polyethylene glycol (PEG) polymer; and
DA and DB are a pair of acceptor and donor fluorescent moieties that are capable of undergoing Forsters/fluorescence resonance energy transfer with the other; and
wherein [CM -M] is bound to at any position of A or X, [DA-CA] is bound to any amino acid of A, and [cB -DB] is bound to any amino acid of B. In some embodiments, CA, Cb, and cM are each independently a 0-1 amino acid. In some embodiments, CA, CB, and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CA, CB, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CB is any amino acid having a free thiol group. In some embodiments, CB is D- cysteine. In some embodiments, CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine. In some embodiments, X comprises the amino acid sequence PLGLAG. In some embodiments, X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, X comprises the amino acid sequence
RPLALWPvS. In some embodiments, DA and DB are Cy5 and Cy7. In some embodiments, DA and DB are Cy5 and IRDye750. In some embodiments, DA and DB are Cy5 and IRDye800. In some embodiments, DA and DB are Cy5 and ICG. In some embodiments, M is selected from a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa),
approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG 40kDa)). In some embodiments, A comprises a thiol reactive group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo. In certain embodiments, the SDM is an SDM comprising the structure of Formula VI.
[0097] In some embodiments, the SDM comprises a structure selected from: SDM-1 , SDM-2, SDM-3, SDM-4, SDM-5, SDM-6, SDM-7, SDM-8, SDM-9, SDM-10, SDM-11, SDM-12, SDM- 13, SDM-14, SDM-15, SDM-16, SDM-17, SDM-18, SDM-19, SDM-20, SDM-21 , SDM-22, SDM- 23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-28, SDM-29, SDM-30, SDM-31 , SDM-32, SDM- 33, SDM-34, SDM-35, SDM-36, SDM-37, SDM-38, SDM-39, and SDM-40 (see International PCT Pub. No. WO2013/019681). In some embodiments, the selective delivery molecule comprises a structure selected from: SDM-14, SDM-15, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-32, or SDM-35. In certain embodiments, the selective delivery molecule comprises Peptide P-3 (see International PCT Pub. No. WO2013/019681).
Portion of A
[0098] In some embodiments, A is a peptide with a sequence comprising 2 to 20 acidic amino acids. In some embodiments, peptide portion of A comprises between about 2 to about 20 acidic amino acids. In some embodiments, peptide portion of A comprises between about 5 to about 20 acidic amino acids. In some embodiments, A has a sequence comprising 5 to 9 acidic amino acids. In some embodiments, A has a sequence comprising 5 to 8 acidic amino acids. In some
embodiments, A has a sequence comprising 5 to 7 acidic amino acids. In some embodiments, A has a sequence comprising 5 acidic amino acids. In some embodiments, A has a sequence comprising 6 acidic amino acids. In some embodiments, A has a sequence comprising 7 acidic amino acids. In some embodiments, A has a sequence comprising 8 acidic amino acids. In some embodiments, A has a sequence comprising 9 acidic amino acids.
[0099] In some embodiments, peptide portion of A comprises between about 2 to about 20 consecutive acidic amino acids. In some embodiments, peptide portion of A comprises between about 5 to about 20 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 5 to 9 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 5 to 8 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 5 to 7 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 5 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 6 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 7 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 8 consecutive acidic amino acids. In some embodiments, A has a sequence comprising 9 consecutive acidic amino acids.
[00100] In some embodiments, peptide portion of A comprises between about 2 to about 20 acidic amino acids selected from, aspartates and glutamates. In some embodiments, peptide portion of A comprises between about 5 to about 20 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 9 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 8 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 7 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 6 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 7 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 8 acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 9 acidic amino acids selected from, aspartates and glutamates.
[00101] In some embodiments, peptide portion of A comprises between about 2 to about 20 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, peptide portion of A comprises between about 5 to about 20 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 9 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 8 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 to 7 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 5 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 6 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 7 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 8 consecutive acidic amino acids selected from, aspartates and glutamates. In some embodiments, A has a sequence comprising 9 consecutive acidic amino acids selected from, aspartates and glutamates.
[00102] In some embodiments, peptide portion of A comprises between about 2 to about 20 glutamates. In some embodiments, peptide portion of A comprises between about 5 to about 20 glutamates. In some embodiments, A has a sequence comprising 5 to 9 glutamates. In some embodiments, A has a sequence comprising 5 to 8 glutamates. In some embodiments, A has a sequence comprising 5 to 7 glutamates. In some embodiments, A has a sequence comprising 5 glutamates. In some embodiments, A has a sequence comprising 6 glutamates. In some
embodiments, A has a sequence comprising 7 glutamates. In some embodiments, A has a sequence comprising 8 glutamates. In some embodiments, A has a sequence comprising 9 glutamates.
[00103] In some embodiments, peptide portion of A comprises between about 2 to about 20 consecutive glutamates. In some embodiments, peptide portion of A comprises between about 5 to about 20 consecutive glutamates. In some embodiments, A has a sequence comprising 5 to 9 consecutive glutamates. In some embodiments, A has a sequence comprising 5 to 8 consecutive glutamates. In some embodiments, A has a sequence comprising 5 to 7 consecutive glutamates. In some embodiments, A has a sequence comprising 5 consecutive glutamates. In some embodiments, A has a sequence comprising 6 consecutive glutamates. In some embodiments, A has a sequence comprising 7 consecutive glutamates. In some embodiments, A has a sequence comprising 8 consecutive glutamates. In some embodiments, A has a sequence comprising 9 consecutive glutamates.
[00104] In some embodiments, portion of A comprises 5 consecutive glutamates (i.e., EEEEE or eeeee). In some embodiments, portion of A comprises 9 consecutive glutamates (i.e., EEEEEEEEE or eeeeeeeee).
[00105] An acidic portion of A may include amino acids that are not acidic. Acidic portion of A may comprise other moieties, such as negatively charged moieties. In embodiments of a selective delivery molecule disclosed herein, an acidic portion of A may be a negatively charged portion, preferably having about 2 to about 20 negative charges at physiological pH that does not include an amino acid. [00106] In some embodiments, the amount of negative charge in portion of A is approximately the same as the amount of positive charge in portion of B. In some embodiments, the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B. In some embodiments, improved tissue uptake is seen in a selective delivery molecule wherein the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B. In some embodiments, improved solubility is observed in a selective delivery molecule wherein the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B. In some embodiments, faster tissue uptake is seen in a selective delivery molecule wherein the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B. In some embodiments, greater tissue uptake is seen in a selective delivery molecule wherein the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B.
[00107] Portion of A is either L-amino acids or D-amino acids. In embodiments of the invention, D-amino acids are preferred in order to minimize immunogenicity and nonspecific cleavage by background peptidases or proteases. Cellular uptake of oligo-D-arginine sequences is known to be as good as or better than that of oligo-L-arginines.
[00108] In some embodiments, portion of A comprises a thiol reactive group. In some
embodiments, the thiol group is an N-terminal thiol group. In some embodiments, the thiol reactive group is selected from among haloacetyls, maleimides, aziridines, acryloyls, arylating agents, vinylsulfones, pyridyl disulfides, TNB-thiols and disulfide reducing agents. In some embodiments, the thiol reactive group covalently binds to a carrier protein. In some embodiments, the carrier protein is albumin. In some embodiments, the thiol reactive group covalently binds to Cysteine 34 of albumin. In some embodiments, the thiol reactive group covalently binds to albumin in vivo.
[00109] It will be understood that portion of A may include non-standard amino acids, such as, for example, hydroxylysine, desmosine, isodesmosine, or other non-standard amino acids. Portion of A may include modified amino acids, including post-translationally modified amino acids such as, for example, methylated amino acids (e.g., methyl histidine, methylated forms of lysine, etc.), acetylated Amino acids, amidated amino acids, formylated amino acids, hydroxylated amino acids, phosphorylated amino acids, or other modified amino acids. Portion of A may also include peptide mimetic moieties, including portions linked by non-peptide bonds and amino acids linked by or to non-amino acid portions.
[00110] The Selective Delivery Molecules disclosed herein are effective where A is at the amino terminus or where A is at the carboxy terminus, i.e., either orientation of the peptide bonds is permissible. Portion of B
[00111] In some embodiments, B is a peptide with a sequence comprising 5 to 15 basic amino acids. In some embodiments, peptide portion of B comprises between about 5 to about 20 basic amino acids. In some embodiments, peptide portion of B comprises between about 5 to about 12 basic amino acids. In some embodiments, peptide portion of B comprises between about 7 to about 9 basic amino acids. In some embodiments, peptide portion of B comprises between about 7 to about 8 basic amino acids. In some embodiments, peptide portion of B comprises 9 basic amino acids. In some embodiments, peptide portion of B comprises 8 basic amino acids. In some embodiments, peptide portion of B comprises 7 basic amino acids.
[00112] In some embodiments, peptide portion of B comprises between about 5 to about 20 consecutive basic amino acids. In some embodiments, peptide portion of B comprises between about 5 to about 12 consecutive basic amino acids. In some embodiments, peptide portion of B comprises between about 7 to about 9 consecutive basic amino acids. In some embodiments, peptide portion of B comprises between about 7 to about 8 consecutive basic amino acids. In some embodiments, peptide portion of B comprises 9 consecutive basic amino acids. In some embodiments, peptide portion of B comprises 8 consecutive basic amino acids. In some embodiments, peptide portion of B comprises 7 consecutive basic amino acids.
[00113] In some embodiments, peptide portion of B comprises between about 5 to about 20 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 5 to about 12 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 7 to about 9 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 7 to about 8 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises 9 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises 8 basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises 7 basic amino acids selected from arginines, histidines, and lysines.
[00114] In some embodiments, peptide portion of B comprises between about 5 to about 20 consecutive basic amino acids selected from arginines, histidines, and lysines. In some
embodiments, peptide portion of B comprises between about 5 to about 12 consecutive basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 7 to about 9 consecutive basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises between about 7 to about 8 consecutive basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises 9 consecutive basic amino acids selected from arginines, histidines, and lysines. In some embodiments, peptide portion of B comprises 8 consecutive basic amino acids selected from arginines, histidines, and lysines. In some
[00115] In some embodiments, peptide portion of B comprises between about 5 to about 20 arginines. In some embodiments, peptide portion of B comprises between about 5 to about 12 arginines. In some embodiments, peptide portion of B comprises between about 7 to about 9 arginines. In some embodiments, peptide portion of B comprises between about 7 to about 8 arginines. In some embodiments, peptide portion of B comprises 9 arginines. In some
embodiments, peptide portion of B comprises 8 arginines. In some embodiments, peptide portion of B comprises 7 arginines.
[00116] In some embodiments, peptide portion of B comprises between about 5 to about 20 consecutive arginines. In some embodiments, peptide portion of B comprises between about 5 to about 12 consecutive arginines. In some embodiments, peptide portion of B comprises between about 7 to about 9 consecutive arginines. In some embodiments, peptide portion of B comprises between about 7 to about 8 consecutive arginines. In some embodiments, peptide portion of B comprises 9 consecutive arginines. In some embodiments, peptide portion of B comprises 8 consecutive arginines. In some embodiments, peptide portion of B comprises 7 consecutive arginines.
[00117] A basic portion of B may include amino acids that are not basic. Basic portion of B may comprise other moieties, such as positively charged moieties. In embodiments, a basic portion of B may be a positively charged portion, preferably having between about 5 and about 20 positive charges at physiological pH, that does not include an amino acid. In some embodiments, the amount of negative charge in portion of A is approximately the same as the amount of positive charge in portion of B. In some embodiments, the amount of negative charge in portion of A is not the same as the amount of positive charge in portion of B.
[00118] Portion of B is either L-amino acids or D-amino acids. In embodiments of the invention, D-amino acids are preferred in order to minimize immunogenicity and nonspecific cleavage by background peptidases or proteases. Cellular uptake of oligo-D-arginine sequences is known to be as good as or better than that of oligo-L-arginines.
[00119] It will be understood that portion of B may include non-standard amino acids, such as, for example, hydroxylysine, desmosine, isodesmosine, or other non-standard amino acids. Portion of B may include modified amino acids, including post-translationally modified amino acids such as, for example, methylated amino acids (e.g., methyl histidine, methylated forms of lysine, etc.), acetylated amino acids, amidated amino acids, formylated amino acids, hydroxylated amino acids, phosphorylated amino acids, or other modified amino acids. Portion of B may also include peptide mimetic moieties, including portions linked by non-peptide bonds and amino acids linked by or to non-amino acid portions.
[00120] In embodiments where X is a peptide cleavable by a protease, it may be preferable to join the C-terminus of X to the N-terminus of B, so that the new amino terminus created by cleavage of X contributes an additional positive charge that adds to the positive charges already present in B. Conjugation Group (c)
[00121] In some embodiments, the cargo (e.g., DA and DB) and the macromolecule carriers (M) are attached indirectly to A-X-B.
[00122] In some embodiments, the cargo (e.g., DA and DB) and the macromolecule carriers (M) are attached indirectly to A-X-B by a conjugation group (CA, CB, and CM). In some embodiments, the cargo (e.g., DA and DB) and the macromolecule carriers (M) are attached indirectly to A-X-B by a reactive conjugation group (CA, CB, and CM). In some embodiments, the cargo (e.g., DA and DB) and the macromolecule carriers (M) are attached indirectly to A-X-B by an orthogonally reactive conjugation group (CA, Cb, and cM). In some embodiments, CA, Cb, and cM each independently comprise an amino acid. In some embodiments, CA, CB, and CM each independently comprise 0-10 amino acids. In some embodiments, CA, CB, and CM each independently comprise 1 amino acid. In some embodiments, CA, Cb, and cM each independently comprise 2 amino acids. In some embodiments, CA, Cb, and cM each independently comprise 3 amino acids. In some embodiments, CA, CB, and CM each independently comprise 4 amino acids. In some embodiments, CA, CB, and CM each independently comprise 5 amino acids. In some embodiments, CA, CB, and CM each
independently comprise 6 amino acids. In some embodiments, CA, Cb, and cM each independently comprise 7 amino acids. In some embodiments, CA, CB, and CM each independently comprise 8 amino acids. In some embodiments, CA, CB, and CM each independently comprise 9 amino acids. In some embodiments, CA, CB, and CM each independently comprise 10 amino acids.
[00123] In some embodiments, CA, CB, and CM each independently comprise a derivatized amino acid. In some embodiments, multiple cargos (D) are attached to a derivatized amino acid conjugation group.
[00124] In some embodiments, the conjugation group comprises a receptor ligand.
[00125] In some embodiments, CA, CB, and CM each independently comprise a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CA, CB, and CM each independently comprise from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a T-amino acid. In some embodiments, CA, CB, and CM each independently comprise any amino acid having a free thiol group, any amino acid containing a free amine group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CA, CB, and CM each independently comprise D-cysteine, D-glutamate, lysine, and para-4-acetyl L- phenylalanine. In some embodiments, CB comprises any amino acid having a free thiol group. In some embodiments, CB comprises D-cysteine. In some embodiments, CA comprises any amino acid having a N-terminal amine group. In some embodiments, CA comprises D-glutamate. In some embodiments, CA comprises lysine. In some embodiments, CM comprises any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM comprises para-4-acetyl L-phenylalanine.
[00126] In some embodiments, CA, Cb, and cM are each independently selected from a naturally- occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CA, CB, and cM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a β- amino acid, or a τ-amino acid. In some embodiments, CA, Cb, and cM are each independently any amino acid having a free thiol group, any amino acid containing a free amine group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CA, CB, and CM are each independently selected from: D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CB is any amino acid having a free thiol group. In some embodiments, cB is D-cysteine. In some embodiments, CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CA is lysine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine.
Cargo (D)
Therapeutic Agents
[00127] Disclosed herein, in certain embodiments, is the use of a selective delivery molecule disclosed herein for delivering a therapeutic agent to a tissue or a plurality of cells. In some embodiments, the therapeutic agent is an anti-inflammatory agent. In some embodiments, the therapeutic agent is an anti-cancer agent. In some embodiments, the selective delivery molecule is used to treat colorectal cancer.
[00128] In some embodiments, a D moiety is independently a therapeutic agent. In some embodiments, a D moiety comprises two or more therapeutic agents. In some embodiments, the two or more therapeutic agents are the same therapeutic agent. In some embodiments, the two or more therapeutic agents are different therapeutic agents. In some embodiments, a D moiety comprises 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 or more therapeutic agents. In some embodiments, the therapeutic agent is selected from: a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, an anti-inflammatory agent, or a combination thereof. In some embodiments, the therapeutic agent is a radiotherapeutic agent. In some embodiments, the therapeutic agent is a cytotoxin.
[00129] In some embodiments, the therapeutic agent is a B cell receptor pathway inhibitor. In some embodiments, the therapeutic agent is a CD79A inhibitor, a CD79B inhibitor, a CD 19 inhibitor, a Lyn inhibitor, a Syk inhibitor, a PI3K inhibitor, a Blnk inhibitor, a PLGy inhibitor, a PKCP inhibitor, or a combination thereof. In some embodiments, the therapeutic agent is an antibody, B cell receptor signaling inhibitor, a PI3K inhibitor, an IAP inhibitor, an mTOR inhibitor, a radioimmunotherapeutic, a DNA damaging agent, a proteosome inhibitor, a histone deacytlase inhibitor, a protein kinase inhibitor, a hedgehog inhibitor, an Hsp90 inhibitor, a telomerase inhibitor, a Jakl/2 inhibitor, a protease inhibitor, a PKC inhibitor, a PARP inhibitor, or a combination thereof. In some embodiments, the therapeutic agent is selected from: chlorambucil, ifosphamide, doxorubicin, mesalazine, thalidomide, lenalidomide, temsirolimus, everolimus, fludarabine, fostamatinib, paclitaxel, docetaxel, ofatumumab, rituximab, dexamethasone, prednisone, CAL-101 , ibritumomab, tositumomab, bortezomib, pentostatin, endostatin,
bendamustine, chlorambucil, chlormethine, cyclophosphamide, ifosfamide, melphalan,
prednimustine, trofosfamide, busulfan, mannosulfan, treosulfan, carboquone, thiotepa, triaziquone, carmustine, fotemustine, lomustine, nimustine, ranimustine, semustine, streptozocin, etoglucid, dacarbazine, mitobronitol, pipobroman, temozolomide, methotrexate, permetrexed, pralatrexate, raltitrexed, cladribine, clofarabine, fludarabine, mercaptopunne, nelarabine, tioguanine, azacitidine, capecitabine, carmofur, cytarabine, decitabine, fluorouracil, gemcitabine, tegafur, vinblastine, vincristine, vindesine, vinflunine, vinorelbine, etoposide, teniposide, demecolcine, docetaxel, paclitaxel, paclitaxel poliglumex, trabectedin, dactinomycin, aclarubicin, daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, pirarubicin, valrubicin, zorubincin, bleomycin, ixabepilone, mitomycin, plicamycin, carboplatin, cisplatin, oxaliplatin, satraplatin, procarbazine, aminolevulinic acid, efaproxiral, methyl aminolevulinate, porfimer sodium, temoporfm, dasatinib, erlotinib, everolimus, gefitinib, imatinib, lapatinib, nilotinib, pazonanib, sorafenib, sunitinib, temsirolimus, alitretinoin, altretamine, amzacrine, anagrelide, arsenic trioxide, asparaginase, bexarotene, bortezomib, celecoxib, denileukin diftitox, estramustine, hydroxycarbamide, irinotecan, lonidamine, masoprocol, miltefosein, mitoguazone, mitotane, oblimersen, pegaspargase, pentostatin, romidepsin, sitimagene ceradenovec, tiazofurine, topotecan, tretinoin, vorinostat, diethylstilbenol, ethinylestradiol, fosfestrol, polyestradiol phosphate, gestonorone, medroxyprogesterone, megestrol, buserelin, goserelin, leuprorelin, triptorelin, fulvestrant, tamoxifen, toremifene, bicalutamide, flutamide, nilutamide, aminoglutethimide, anastrozole, exemestane, formestane, letrozole, vorozole, abarelix, degarelix, histamine dihydrochloride, mifamurtide, pidotimod, plerixafor, roquinimex, thymopentin, everolimus, gusperimus, leflunomide, mycophenolic acid, sirolimus, ciclosporin, tacrolimus, azathioprine, lenalidomide, methotrexate, thalidomide, iobenguane, ancestim, filgrastim, lenograstim, molgramostim, pegfilgrastim, sargramostim, interferon alfa natural, interferon alfa-2a, interferon alfa-2b, interferon alfacon-1, interferon alfa-nl, interferon beta natural, interferon beta-la, interferon beta-lb, interferon gamma, peginterferon alfa-2a, peginterferon alfa-2b, aldesleukin, oprelvekin, BCG vaccine, glatiramer acetate, histamine dihydrochloride, immunocyanin, lentinan, melanoma vaccine, mifamurtide, pegademase, pidotimod, plerixafor, poly I:C, poly ICLC, roquinimex, tasonermin, thymopentin, abatacept, abetimus, alefacept, antilymphocyte immunoglobulin (horse), antithymocyte immunoglobulin (rabbit), eculizumab, efalizumab, everolimus, gusperimus, leflunomide, muromab-CD3, mycophenolic acid, natalizumab, sirolimus, adalimumab,
afelimomab, certolizumab pegol, etanercept, golimumab, infliximab, anakinra, basiliximab, canakinumab, daclizumab, mepolizumab, rilonacept, tocilizumab, ustekinumab, ciclosporin, tacrolimus, azathioprine, lenalidomide, methotrexate, thalidomide, adalimumab, alemtuzumab, bevacizumab, cetuximab, certolizumab pegol, , eculizumab, efalizumab, gemtuzumab, ibritumomab tiuxetan, muromonab-CD3, natalizumab, panitumumab, ranibizumab, rituximab, tositumomab, trastuzumab, catumaxomab, edrecolomab, ofatumumab, muromab-CD3, afelimomab, golimumab, ibritumomab tiuxetan, abagovomab, adecatumumab, alemtuzumab, anti-CD30 monoclonal antibody Xmab2513, anti-MET monoclonal antibody MetMab, apolizumab, apomab, arcitumomab, bispecific antibody 2B1, blinatumomab, brentuximab vedotin, capromab pendetide, cixutumumab, claudiximab, conatumumab, dacetuzumab, denosumab, eculizumab, epratuzumab, epratuzumab, ertumaxomab, etaracizumab, figitumumab, fresolimumab, galiximab, ganitumab, gemtuzumab ozogamicin, glembatumumab, ibritumomab, inotuzumab ozogamicin, ipilimumab, lexatumumab, lintuzumab, lintuzumab, lucatumumab, mapatumumab, matuzumab, milatuzumab, monoclonal antibody CC49, necitumumab, nimotuzumab, ofatumumab, oregovomab, pertuzumab,
ramacurimab, ranibizumab, siplizumab, sonepcizumab, tanezumab, tositumomab, trastuzumab, tremelimumab, tucotuzumab celmoleukin, veltuzumab, visilizumab, volociximab, zalutumumab, a syk inhibitor (e.g., R788), enzastaurin, dasatinib, erlotinib, everolimus, gefitinib, imatinib, lapatinib, nilotinib, pazonanib, sorafenib, sunitinib, temsirolimus, an angiogenesis inhibitor (e.g., GT-111, JI-101, R1530), a kinase inhibitors (e.g., AC220, AC480, ACE-041, AMG 900, AP24534, Arry-614, AT7519, AT9283, AV-951, axitinib, AZD1152, AZD7762, AZD8055, AZD8931, bafetinib, BAY 73-4506, BGJ398, BGT226, BI 811283, BI6727, BIBF 1120, BIBW 2992, BMS- 690154, BMS-777607, BMS-863233, BSK-461364, CAL-101, CEP-11981 , CYC116, DCC-2036, dinaciclib, dovitinib lactate, E7050, EMD 1214063, ENMD-2076, fostamatinib disodium,
GSK2256098, GSK690693, INCB18424, INNO-406, J J-26483327, JX-594, KX2-391, linifanib, LY2603618, MGCD265, MK-0457, MK1496, MLN8054, MLN8237, MP470, NMS-1116354, NMS-1286937, ON 01919.Na, OSI-027, OSI-930, Btk inhibitor, PF-00562271, PF-02341066, PF- 03814735, PF-04217903, PF-04554878, PF-04691502, PF-3758309, PHA-739358, PLC3397, progenipoietin, R547, R763, ramucirumab, regorafenib, R05185426, SAR103168, S3333333CH 727965, SGI-1176, SGX523, SNS-314, TAK-593, TAK-901, TKI258, TLN-232, TTP607, XL147, XL228, XL281R05126766, XL418, XL765), an inhibitor of mitogen-activated protein kinase signaling (e.g., U0126, PD98059, PD184352, PD0325901, ARRY-142886, SB239063, SP600125, BAY 43-9006, wortmannin, or LY294002), adriamycin, dactinomycin, bleomycin, vinblastine, cisplatin, acivicin, aclarubicin, acodazole hydrochloride, acronine, adozelesin, aldesleukin, altretamine, ambomycin, ametantrone acetate, aminoglutethimide, amsacrine, anastrozole, anthramycin, asparaginase, asperlin, azacitidine, azetepa, azotomycin, batimastat, benzodepa, bicalutamide, bisantrene hydrochloride, bisnafide dimesylate, bizelesin, bleomycin sulfate, brequinar sodium, bropirimine, busulfan, cactinomycin, calusterone, caracemide, carbetimer, carboplatin, carmustine, carubicin hydrochloride, carzelesin, cedeflngol, chlorambucil, cirolemycin, cladribine, crisnatol mesylate, cyclophosphamide, cytarabine, dacarbazine, daunorubicin hydrochloride, decitabine, dexormaplatin, dezaguanine, dezaguanine mesylate, diaziquone, doxorubicin, doxorubicin hydrochloride, droloxifene, droloxifene citrate, dromostanolone propionate, duazomycin, edatrexate, eflornithine hydrochloride, elsamitrucin, enloplatin, enpromate, epipropidine, epirubicin hydrochloride, erbulozole, esorubicin hydrochloride, estramustine, estramustine phosphate sodium, etanidazole, etoposide, etoposide phosphate, etoprine, fadrozole hydrochloride, fazarabine, fenretinide, floxuridine, fludarabine phosphate, fluorouracil, flurocitabine, fosquidone, fostriecin sodium, gemcitabine, gemcitabine hydrochloride, hydroxyurea, idarubicin hydrochloride, ifosfamide, iimofosine, interleukin II (including
recombinant interleukin II, or rlL2), interferon alfa-2a, interferon alfa -2b, interferon alfa -nl, interferon alfa -n3, interferon beta-1 a, interferon gamma-1 b, iproplatin, irinotecan hydrochloride, lanreotide acetate, letrozole, leuprolide acetate, liarozole hydrochloride, lometrexol sodium, lomustine, losoxantrone hydrochloride, masoprocol, maytansine, mechlorethamine hydrochloride, megestrol acetate, melengestrol acetate, melphalan, menogaril, mercaptopurine, methotrexate, methotrexate sodium, metoprine, meturedepa, mitindomide, mitocarcin, mitocromin, mitogillin, mitomalcin, mitomycin, mitosper, mitotane, mitoxantrone hydrochloride, mycophenolic acid, nocodazoie, nogalamycin, ormaplatin, oxisuran, pegaspargase, peliomycin, pentamustine, peplomycin sulfate, perfosfamide, pipobroman, piposulfan, piroxantrone hydrochloride, plicamycin, plomestane, porfimer sodium, porfiromycin, prednimustine, procarbazine
hydrochloride, puromycin, puromycin hydrochloride, pyrazofurin, riboprine, rogletimide, safingol, safingol hydrochloride, semustine, simtrazene, sparfosate sodium, sparsomycin, spirogermanium hydrochloride, spiromustine, spiroplatin, streptonigrin, streptozocin, sulofenur, talisomycin, tecogalan sodium, tegafur, teloxantrone hydrochloride, temoporfin, teniposide, teroxirone, testolactone, thiamiprine, thioguanine, thiotepa, tiazofurin, tirapazamine, toremifene citrate, trestolone acetate, triciribine phosphate, trimetrexate, trimetrexate glucuronate, triptorelin, tubulozole hydrochloride, uracil mustard, uredepa, vapreotide, verteporfin, vinblastine sulfate, vincristine sulfate, vindesine, vindesine sulfate, vinepidine sulfate, vinglycinate sulfate, vinleurosine sulfate, vinorelbine tartrate, vinrosidine sulfate, vinzolidine sulfate, vorozole, zeniplatin, zinostatin, zorubicin hydrochloride. In some embodiments, the therapeutic agent is selected from: 20-epi-l, 25 dihydroxyvitamin D3, 5-ethynyluracil, abiraterone, aclarubicin, acylfulvene, adecypenol, adozelesin, aldesleukin, ALL-TK antagonists, altretamine, ambamustine, amidox, amifostine, aminolevulinic acid, amrubicin, amsacrine, anagrelide, anastrozole, andrographolide, angiogenesis inhibitors, antagonist D, antagonist G, antarelix, anti-dorsalizing morphogenetic protein- 1, antiandrogen, prostatic carcinoma, antiestrogen, antineoplaston, antisense oligonucleotides, aphidicolin glycinate, apoptosis gene modulators, apoptosis regulators, apurinic acid, ara-CDP-DL-PTBA, arginine deaminase, asulacrine, atamestane, atrimustine, axinastatin 1 , axinastatin 2, axinastatin 3, azasetron, azatoxin, azatyrosine, baccatin III derivatives, balanol, batimastat, BCR/ABL antagonists, benzochlorins, benzoylstaurosporine, beta lactam derivatives, beta-alethine, betaclamycin B, betulinic acid, bFGF inhibitor, bicalutamide, bisantrene, bisaziridinylspermine, bisnafide, bistratene A, bizelesin, breflate, bropirimine, budotitane, buthionine sulfoximine, calcipotriol, calphostin C, camptothecin derivatives, canarypox IL-2, capecitabine, carboxamide-amino-triazole, carboxyamidotriazole, CaRest M3, CARN 700, cartilage derived inhibitor, carzelesin, casein kinase inhibitors (ICOS), castanospermine, cecropin B, cetrorelix, chlorlns, chloroquinoxaline sulfonamide, cicaprost, cis-porphyrin, cladribine, clomifene analogues, clotrimazole, collismycin A, collismycin B, combretastatin A4,
combretastatin analogue, conagenin, crambescidin 816, crisnatol, cryptophycin 8, cryptophycin A derivatives, curacin A, cyclopentanthraquinones, cycloplatam, cypemycin, cytarabine ocfosfate, cytolytic factor, cytostatin, dacliximab, decitabine, dehydrodidemnin B, deslorelin, dexamethasone, dexifosfamide, dexrazoxane, dexverapamil, diaziquone, didemnin B, didox, diethylnorspermine, dihydro-5-azacytidine, 9- dioxamycin, diphenyl spiromustine, docosanol, dolasetron, doxifiuridine, droloxifene, dronabinol, duocarmycin SA, ebselen, ecomustine, edelfosine, edrecolomab, eflornithine, elemene, emitefur, epirubicin, epristeride, estramustine analogue, estrogen agonists, estrogen antagonists, etanidazole, etoposide phosphate, exemestane, fadrozole, fazarabine, fenretinide, filgrastim, finasteride, flavopiridol, flezelastine, fluasterone, fludarabine,
fluorodaunorunicin hydrochloride, forfenimex, formestane, fostriecin, fotemustine, gadolinium texaphyrin, gallium nitrate, galocitabine, ganirelix, gelatinase inhibitors, gemcitabine, glutathione inhibitors, hepsulfam, heregulin, hexamethylene bisacetamide, hypericin, ibandronic acid, idarubicin, idoxifene, idramantone, ilmofosine, ilomastat, imidazoacridones, imiquimod, immunostimulant peptides, insulin-such as for example growth factor- 1 receptor inhibitor, interferon agonists, interferons, interleukins, iobenguane, iododoxorubicin, ipomeanol, 4-, iroplact, irsogladine, isobengazole, isohomohalicondnn B, itasetron, jasplakinolide, kahalalide F, lamellarin- N triacetate, lanreotide, leinamycin, lenograstim, lentinan sulfate, leptolstatin, letrozole, leukemia inhibiting factor, leukocyte alpha interferon, leuprolide+estrogen+progesterone, leuprorelin, levamisole, liarozole, linear polyamine analogue, lipophilic disaccharide peptide, lipophilic platinum compounds, lissoclinamide 7, lobaplatin, lombricine, lometrexol, lonidamine, losoxantrone, lovastatin, loxoribine, lurtotecan, lutetium texaphyrin, lysofylline, lytic peptides, maitansine, mannostatin A, marimastat, masoprocol, maspin, matrilysin inhibitors, matrix metalloproteinase inhibitors, menogaril, merbarone, meterelin, methioninase, metoclopramide, MIF inhibitor, mifepristone, miltefosine, mirimostim, mismatched double stranded RNA, mitoguazone, mitolactol, mitomycin analogues, mitonafide, mitotoxin fibroblast growth factor-saporin, mitoxantrone, mofarotene, molgramostim, monoclonal antibody, human chorionic gonadotrophin, monophosphoryl lipid A+myobacterium cell wall sk, mopidamol, multiple drug resistance gene inhibitor, multiple tumor suppressor 1 -based therapy, mustard anticancer agent, mycaperoxide B, mycobacterial cell wall extract, myriaporone, N-acetyldinaline, N-substituted benzamides, nafarelin, nagrestip, naloxone+pentazocine, napavin, naphterpin, nartograstim, nedaplatin, nemorubicin, neridronic acid, neutral endopeptidase, nilutamide, nisamycin, nitric oxide modulators, nitroxide antioxidant, nitrullyn, 06-benzylguanine, octreotide, okicenone,
oligonucleotides, onapristone, ondansetron, ondansetron, oracin, oral cytokine inducer, ormaplatin, osaterone, oxaliplatin, oxaunomycin, palauamine, palmitoylrhizoxin, pamidronic acid, panaxytriol, panomifene, parabactin, pazelliptine, pegaspargase, peldesine, pentosan polysulfate sodium, pentostatin, pentrozole, perflubron, perfosfamide, perillyl alcohol, phenazinomycin, phenylacetate, phosphatase inhibitors, picibanil, pilocarpine hydrochloride, pirarubicin, piritrexim, placetin A, placetin B, plasminogen activator inhibitor, platinum complex, platinum compounds, platinum- triamine complex, porfimer sodium, porfiromycin, prednisone, propyl bis-acridone, prostaglandin J2, proteasome inhibitors, protein A-based immune modulator, protein kinase C inhibitor, protein kinase C inhibitors, microalgal, protein tyrosine phosphatase inhibitors, purine nucleoside phosphorylase inhibitors, purpurins, pyrazoloacridine, pyridoxylated hemoglobin polyoxyethylerie conjugate, raf antagonists, raltitrexed, ramosetron, ras farnesyl protein transferase inhibitors, ras inhibitors, ras-GAP inhibitor, retelliptine demethylated, rhenium Re 186 etidronate, rhizoxin, ribozymes, RII retinamide, rogletimide, rohitukine, romurtide, roquinimex, rubiginone Bl, ruboxyl, safmgol, saintopin, SarCNU, sarcophytol A, sargramostim, Sdi 1 mimetics, semustine, senescence derived inhibitor 1 , sense oligonucleotides, signal transduction inhibitors, signal transduction modulators, single chain antigen-binding protein, sizofiran, sobuzoxane, sodium borocaptate, sodium phenylacetate, solverol, somatomedin binding protein, sonermin, sparfosic acid, spicamycin D, spiromustine, splenopentin, spongistatin 1, squalamine, stem cell inhibitor, stem-cell division inhibitors, stipiamide, stromelysin inhibitors, sulfmosine, superactive vasoactive intestinal peptide antagonist, suradista, suramin, swainsonine, synthetic glycosaminoglycans, tallimustine, tamoxifen methiodide, tauromustine, tazarotene, tecogalan sodium, tegafur, tellurapyrylium, telomerase inhibitors, temoporfm, temozolomide, teniposide, tetrachlorodecaoxide, tetrazomine, thaliblastine, thiocoraline, thrombopoietin, thrombopoietin mimetic, thymalfasin, thymopoietin receptor agonist, thymotrinan, thyroid stimulating hormone, tin ethyl etiopurpurin, tirapazamine, titanocene bichloride, topsentin, toremifene, totipotent stem cell factor, translation inhibitors, tretinoin, triacetyluridine, triciribine, trimetrexate, triptorelin, tropisetron, turosteride, tyrosine kinase inhibitors, tyrphostins, UBC inhibitors, ubenimex, urogenital sinus-derived growth inhibitory factor, urokinase receptor antagonists, vapreotide, variolin B, vector system, erythrocyte gene therapy, velaresol, veramine, verdins, verteporfin, vinorelbine, vinxaltine, vitaxin, vorozole, zanoterone, zeniplatin, zilascorb, zinostatin stimalamer, mechloroethamine, cyclophosphamide, chlorambucil, busulfan, carmustine, lomusitne, decarbazine, methotrexate, cytarabine,
mercaptopurine, thioguanine, pentostatin, mechloroethamine, cyclophosphamide, chlorambucil, meiphalan, ethylenimine, methylmelamine, hexamethlymelamine, thiotepa, busulfan, carmustine, lomusitne, semustine, streptozocin, decarbazine, fluorouracil, floxouridine, cytarabine,
mercaptopurine, thioguanine, pentostatin, erbulozole (also known as R-55104), Dolastatin 10 (also known as DLS-10 and NSC-376128), Mivobulin isethionate (also known as CI-980), Vincristine, NSC-639829, Discodermolide (also known as NVP-XX-A-296), ABT-751 (Abbott, also known as E-7010), Altorhyrtins (such as Altorhyrtin A and Altorhyrtin C), Spongistatins (such as
Spongistatin 1, Spongistatin 2, Spongistatin 3, Spongistatin 4, Spongistatin 5, Spongistatin 6, Spongistatin 7, Spongistatin 8, and Spongistatin 9), Cemadotin hydrochloride (also known as LU- 103793 and NSC-D-669356), Epothilones (such as Epothilone A, Epothilone B, Epothilone C (also known as desoxyepothilone A or dEpoA), Epothilone D (also referred to as KOS-862, dEpoB, and desoxyepothilone B ), Epothilone E, Epothilone F, Epothilone B N-oxide, Epothilone A N-oxide, 16-aza-epothilone B, 21-aminoepothilone B (also known as BMS-310705), 21 -hydroxyepothilone D (also known as Desoxyepothilone F and dEpoF), 26-fluoroepothilone), Auristatin PE (also known as NSC-654663), Soblidotin (also known as TZT-1027), LS-4559-P (Pharmacia, also known as LS-4577), LS-4578 (Pharmacia, also known as LS-477-P), LS-4477 (Pharmacia), LS- 4559 (Pharmacia), RPR-112378 (Aventis), Vincristine sulfate, DZ-3358 (Daiichi), FR-182877 (Fujisawa, also known as WS-9885B), GS-164 (Takeda), GS-198 (Takeda), KAR-2 (Hungarian Academy of Sciences), BSF-223651 (BASF, also known as ILX-651 and LU-223651 ), SAH- 49960 (Lilly/Novartis), SDZ-268970 (Lilly/Novartis), AM-97 (Armad/Kyowa Hakko), AM- 132 (Armad), AM-138 (Armad/Kyowa Hakko), IDN-5005 (Indena), Cryptophycin 52 (also known as LY-355703), AC-7739 (Ajinomoto, also known as AVE-8063A and CS-39.HCI), AC-7700 (Ajinomoto, also known as AVE-8062, AVE-8062A, CS-39-L-Ser.HCI, and RPR-258062A), Vitilevuamide, Tubulysin A, Canadensol, Centaureidin (also known as NSC-106969), T-138067 (Tularik, also known as T-67, TL-138067 and TI-138067), COBRA-1 (Parker Hughes Institute, also known as DDE-261 and WHI-261), H10 (Kansas State University), H16 (Kansas State University), Oncocidin Al (also known as BTO-956 and DIME), DDE-313 (Parker Hughes Institute), Fijianolide B, Laulimalide, SPA-2 (Parker Hughes Institute), SPA-1 (Parker Hughes Institute, also known as SPIKET-P), 3-IAABU (Cytoskeleton/Mt. Sinai School of Medicine, also known as MF-569), Narcosine (also known as NSC-5366), Nascapine, D-24851 (Asta Medica), A- 105972 (Abbott), Hemiasterlin, 3-BAABU (Cytoskeleton/Mt. Sinai School of Medicine, also known as MF-191), TMPN (Arizona State University), Vanadocene acetylacetonate, T-138026 (Tularik), Monsatrol, lnanocine (also known as NSC-698666), 3-lAABE (Cytoskeleton/Mt. Sinai School of Medicine), A-204197 (Abbott), T-607 (Tuiarik, also known as T-900607), RPR- 115781 (Aventis), Eleutherobins (such as Desmethyleleutherobin, Desaetyleleutherobin, lsoeleutherobin A, and Z-Eleutherobin), Caribaeoside, Caribaeolin, Halichondrin B, D-64131 (Asta Medica), D-68144 (Asta Medica), Diazonamide A, A-293620 (Abbott), NPI-2350 (Nereus), Taccalonolide A, TUB- 245 (Aventis), A-2 9754 (Abbott), Diozostatin, (-)-Phenylahistin (also known as NSCL-96F037), D-68838 (Asta Medica), D-68836 (Asta Medica), Myoseverin B, D-43411 (Zentaris, also known as D-81862), A-289099 (Abbott), A-318315 (Abbott), HTI-286 (also known as SPA-110, trifluoroacetate salt) (Wyeth), D-82317 (Zentaris), D-82318 (Zentaris), SC-12983 (NCI),
Resverastatin phosphate sodium, BPR-OY-007 (National Health Research Institutes), and SSR- 250411 (Sanofi). [00130] In some embodiments, the therapeutic agent is an anti-inflammatory agent. In some embodiments, the therapeutic agent is an anti-TNF agent, an IL-1 receptor antagonist, an IL-2 receptor antagonist, a cytotoxic agent, an immunomodulatory agent, an antibiotic, a T-cell co- stimulatory blocker, a B cell depleting agent, an immunosuppressive agent, an alkylating agent, an anti-metabolite, a plant alkaloid, a terpenoids, a topoisomerase inhibitor, an antitumour antibiotic, an antibody, a hormonal therapy, an anti-diabetes agent, a leukotriene inhibitor, or combinations thereof. In some embodiments, the therapeutic agent is selected from: alefacept, efalizumab, methotrexate, acitretin, isotretinoin, hydroxyurea, mycophenolate mofetil, sulfasalazine, 6- Thioguanine, Dovonex, Taclonex, betamethasone, tazarotene, hydroxychloroquine, etanercept, adalimumab, infliximab, abatacept, rituximab, tratuzumab, Anti-CD45 monoclonal antibody AHN- 12 (NCI), Iodine-131 Anti-Bl Antibody (Corixa Corp.), anti-CD66 monoclonal antibody BW 250/183 (NCI, Southampton General Hospital), anti-CD45 monoclonal antibody (NCI, Baylor College of Medicine), antibody anti-anb3 integrin (NCI), BIW-8962 (BioWa Inc.), Antibody BC8 (NCI), antibody muJ591 (NCI), indium In 11 1 monoclonal antibody MN-14 (NCI), yttrium Y 90 monoclonal antibody MN-14 (NCI), F105 Monoclonal Antibody (NIAID), Monoclonal Antibody RAVI 2 (Raven Biotechnologies), CAT- 192 (Human Anti-TGF-Betal Monoclonal Antibody, Genzyme), antibody 3F8 (NCI), 177Lu-J591 (Weill Medical College of Cornell University), TB- 403 (Biolnvent International AB), anakinra, azathioprine, cyclophosphamide, cyclosporine A, leflunomide, d-penicillamine, amitriptyline, or nortriptyline, chlorambucil, nitrogen mustard, prasterone, LJP 394 (abetimus sodium), LJP 1082 (La Jolla Pharmaceutical), eculizumab, belibumab, rhuCD40L (NIAID), epratuzumab, sirolimus, tacrolimus, pimecrolimus, thalidomide, antithymocyte globulin-equine (Atgam, Pharmacia Upjohn), antithymocyte globulin-rabbit (Thymoglobulin, Genzyme), Muromonab-CD3 (FDA Office of Orphan Products Development), basiliximab, daclizumab, riluzole, cladribine, natalizumab, interferon beta-lb, interferon beta- la, tizanidine, baclofen, mesalazine, asacol, pentasa, mesalamine, balsalazide, olsalazine, 6- mercaptopurine, AIN4 7 (Anti IL-17 Monoclonal Antibody, Novartis), theophylline, D2E7 (a human anti-TNF mAb from Knoll Pharmaceuticals), Mepolizumab (Anti-IL-5 antibody, SB 240563), Canakinumab (Anti-IL- 1 Beta Antibody, NIAMS), Anti-IL-2 Receptor Antibody (Daclizumab, NHLBI), CNTO 328 (Anti IL-6 Monoclonal Antibody, Centocor), ACZ885 (fully human anti-interleukin-lbeta monoclonal antibody, Novartis), CNTO 1275 (Fully Human Anti-IL- 12 Monoclonal Antibody, Centocor), (3S)-N-hydroxy-4-({4-[(4-hydroxy-2- butynyl)oxy]phenyl}sulfonyl)-2,2-dimet- hyl-3-thiomorpholine carboxamide (apratastat), golimumab (CNTO 148), Onercept, BG9924 (Biogen Idee), Certolizumab Pegol (CDP870, UCB Pharma), AZD9056 (AstraZeneca), AZD5069 (AstraZeneca), AZD9668 (AstraZeneca), AZD7928 (AstraZeneca), AZD2914 (AstraZeneca), AZD6067 (AstraZeneca), AZD3342 (AstraZeneca), AZD8309 (AstraZeneca), ), [(lR)-3-methyl-l-({(2S)-3-phenyl-2-[(pyrazin-2- ylcarbonyl)amino]propanoyl}amino)butyl]boronic acid (Bortezomib), AMG-714, (Anti-IL 15 Human Monoclonal Antibody, Amgen), ABT-874 (Anti IL-12 monoclonal antibody, Abbott Labs), MRA(Tocilizumab, an Anti IL-6 Receptor Monoclonal Antibody, Chugai Pharmaceutical), CAT- 354 (a human anti-interleukin-13 monoclonal antibody, Cambridge Antibody Technology, Medlmmune), aspirin, salicylic acid, gentisic acid, choline magnesium salicylate, choline salicylate, choline magnesium salicylate, choline salicylate, magnesium salicylate, sodium salicylate, diflunisal, carprofen, fenoprofen, fenoprofen calcium, flurobiprofen, ibuprofen, ketoprofen, nabutone, ketolorac, ketorolac tromethamine, naproxen, oxaprozin, diclofenac, etodolac, indomethacin, sulindac, tolmetin, meclofenamate, meclofenamate sodium, mefenamic acid, piroxicam, meloxicam, celecoxib, rofecoxib, valdecoxib, parecoxib, etoricoxib, lumiracoxib, CS-502 (Sankyo), JTE-522 (Japan Tobacco Inc.), L-745,337 (Almirall), NS398 (Sigma), betamethasone (Celestone), prednisone (Deltasone), alclometasone, aldosterone, amcinonide, beclometasone, betamethasone, budesonide, ciclesonide, clobetasol, clobetasone, clocortolone, cloprednol, cortisone, cortivazol, deflazacort, deoxycorticosterone, desonide, desoximetasone, desoxycortone, dexamethasone, difiorasone, diflucortolone, difiuprednate, fluclorolone, fludrocortisone, fludroxycortide, flumetasone, flunisolide, fluocinolone acetonide, fluocinonide, fluocortin, fiuocortolone, fiuorometholone, fiuperolone, fluprednidene, fluticasone, formocortal, formoterol, halcinonide, halometasone, hydrocortisone, hydrocortisone aceponate, hydrocortisone buteprate, hydrocortisone butyrate, loteprednol, medrysone, meprednisone, methylprednisolone, methylprednisolone aceponate, mometasone furcate, paramethasone, prednicarbate, prednisone, rimexolone, tixocortol, triamcinolone, ulobetasol, Pioglitazone, Rosiglitazone, Glimepiride, Glyburide, Chlorpropamide, Glipizide, Tolbutamide, Tolazamide, Glucophage, Metformin, (glyburide + metformin), Rosiglitazone + metformin, (Rosiglitazone+glimepiride), Exenatide, Insulin, Sitagliptin, (glipizide and metformin), Repaglinide, Acarbose, Nateglinide, Orlistat, cisplatin; carboplatin; oxaliplatin; mechlorethamine; cyclophosphamide; chlorambucil; vincristine; vinblastine; vinorelbine; vindesine; mercaptopurine; fludarabine; pentostatin; cladribine; 5- fluorouracil ( FU); floxuridine (FUDR); cytosine arabinoside; trimethoprim; pyrimethamine; pemetrexed; paclitaxel; docetaxel; etoposide; teniposide; irinotecan; topotecan; amsacrine;
etoposide; etoposide phosphate; teniposide; dactinomycin; doxorubicin; daunorubicin; valrubicine; idarubicine; epirubicin; bleomycin; plicamycin; mitomycin; finasteride; goserelin;
aminoglutethimide; anastrozole; letrozole; vorozole; exemestane; 4-androstene-3,6,17-trione ("6- OXO"; l,4,6-androstatrien-3,17-dione (ATD); formestane; testolactone; fadrozole; A-81834 (3-(3- (l,l-dimethylethylthio-5-(quinoline-2- ylmethoxy)- 1 -(4-chloromethylphenyl)indole-2-yl)-2,2- dimethylpropionaldehyde oxime-O-2-acetic acid; AME103 (Amira); AME803 (Amira); atreleuton; BAY-x-1005 ((R)-(+)-alpha-cyclopentyl-4-(2-quinolinylmethoxy)-Benzeneacetic acid); CJ-13610 (4-(3-(4-(2-Methyl-imidazol-l-yl)-phenylsulfanyl)- phenyl)-tetrahydro-pyran-4-carboxylic acid amide); DG-031 (DeCode); DG-051 (DeCode); MK886 (l-[(4-chlorophenyl)methyl]3-[(l,l- dimethylethyl)thio]-a,a-dimethyl-5-(l-methylethyl)-lH-indole-2-propanoic acid, sodium salt); MK591 (3-(l-4[(4-chlorophenyl)methyl]-3 (t-butylthio)-5-((2-quinoly)methoxy) H-indole-2]-, dimehtylpropanoic acid); RP64966 ([4-[5-(3-Phenyl-propyl)thiophen-2- yl]butoxy] acetic acid); SA6541 ((R)-S-[[4- (dimethylamino)phenyl]methyl] -N-(3-mercapto-2methyl- 1 -oxopropyl-L- cycteine); SC-56938 (ethyl- l-[2-[4-(phenylmethyl)phenoxy] ethyl] -4-piperidine- carboxylate); VIA-2291 (Via Pharmaceuticals); WY-47,288 (2-[(l-naphthalenyloxy)methyl]quinoline); zileuton; ZD-2138 (6-((3-f uoro-5- (tetrahydro-4-methoxy-2H-pyran-4yl)phenoxy)methyl)-l-methyl-2(lH)- quinlolinone); doxycycline; or combinations thereof.
[00131] In some embodiments, the therapeutic agent contains a radioactive moiety, for example a radioactive isotope such as 211At, 1 1I, 125I, 90Y, 186Re, 188Re, 153Sm, 212Bi, 32P, 64Cu radioactive isotopes of Lu, and others.
Macromolecular Carriers (M)
[00132] Polymers are characterized by a distribution of molecular weights, and, as such, the molecular weight, presented herein for polymers, is only an approximate average molecular weight of a distribution of molecular weights of individual polymers. Unless stated otherwise, the molecular weight of a polymeric component will have a typical (i.e., as known in the art) error and standard deviation.
[00133] In some embodiments, a carrier modulates plasma half-life of a selective delivery molecule disclosed herein. In some embodiments, a carrier modulates solubility of a selective delivery molecule disclosed herein. In some embodiments, a carrier modulates bio-distribution of a selective delivery molecule disclosed herein.
[00134] In some embodiments, a carrier decreases uptake of a selective delivery molecule by non- target cells or tissues. In some embodiments, a carrier decreases uptake of a selective delivery molecule into cartilage. In some embodiments, a carrier decreases uptake of a selective delivery molecule into joints relative to target tissue.
[00135] In some embodiments, a carrier increases uptake of a selective delivery molecule by target cells or tissues. In some embodiments, a carrier decreases uptake of a selective delivery molecule into the liver relative to target tissue. In some embodiments, a carrier decreases uptake of a selective delivery molecule into kidneys. In some embodiments, a carrier enhances uptake into cancer tissue. In some embodiments, a carrier enhances uptake into lymphatic channels and/or lymph nodes.
[00136] In some embodiments, a carrier increases plasma half-life by reducing glomerular filtration. In some embodiments, a carrier modulates plasma half-life by increasing or decreases metabolism or protease degradation. In some embodiments, a carrier increases tumor uptake due to enhanced permeability and retention (EPR) of tumor vasculature. In some embodiments, a carrier increases the aqueous solubility of selective delivery molecule.
[00137] In some embodiments, any M is independently directly or indirectly (e.g., via CM) bound to A, B, or X. In some embodiments, any M is independently bound to A at the n-terminal poly glutamate. In some embodiments, any M is independently bound to A (or, the n-terminal poly glutamate) by a covalent linkage. In some embodiments, any M is independently bound to B at the c-terminal polyarginine. In some embodiments, any M is independently bound to B (or, the c- terminal polyarginine) by a covalent linkage. In some embodiments, any M is independently directly or indirectly bound to linkers between X and A, X and B, B and C N terminus, and A and C/N terminus. In some embodiments, the covalent linkage comprises an ether bond, thioether bond, amine bond, amide bond, oxime bond, carbon-carbon bond, carbon-nitrogen bond, carbon-oxygen bond, or carbon-sulfur bond.
[00138] In some embodiments, M is selected from a protein, a synthetic or natural polymer, or a dendrimer. In some embodiments, M is selected from dextran, a PEG polymer (e.g., a PEG polymer having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa), approximately lkDa (PEG lkDa), approximately 2kDa (PEG 2kDa), approximately approximately (PEG 3kDa), approximately 4kDa (PEG 4kDa), approximately 5kDa (PEG 5kDa), approximately lOkDa (PEG lOkDa), approximately 12kDa (PEG 12kDa), approximately 15kDa (PEG 15kDa), approximately 20kDa (PEG 20kDa), approximately 30kDa (PEG 30kDa), or approximately 40kDa (PEG
40kDa))), albumin, or a combination thereof. In some embodiments, M is a PEG polymer.
[00139] In some embodiments, the size of M is between about 50kDa and about 70kDa.
[00140] In some embodiments, the selective delivery molecule is conjugated to albumin. In certain instances, albumin is excluded from the glomerular filtrate under normal physiological conditions. In some embodiments, the selective delivery molecule comprises a reactive group such as maleimide that can form a covalent conjugate with albumin. A selective delivery molecule comprising albumin results in enhanced accumulation of cleaved selective delivery molecules in tumors in a cleavage dependent manner. In some embodiments, albumin conjugates have good pharmacokinetic properties. [00141] In some embodiments, the selective delivery molecule is conjugated to PEG polymers. In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 0.5kDa (PEG 0.5kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately lkDa (PEG lkDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 2kDa (PEG 2kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 3kDa (PEG 3kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 4kDa (PEG 4kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 5kDa (PEG 5kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately lOkDa (PEG lOkDa). In some embodiments, the selective delivery molecule is conjugated PEG polymers having an average molecular weight of approximately 12 kDa ( PEG 12kDa). In some embodiments, the selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 15kDa (PEG 15kDa). In some embodiments, selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 20 kDa (PEG 20kDa). In some embodiments, selective delivery molecule is conjugated to PEG polymers having an average molecular weight of approximately 30 kDa (PEG 30kDa). In some embodiments, selective delivery molecules conjugated to PEG30kDa had a longer half-life as compared to free peptides. In some
embodiments, selective delivery molecules are conjugated to PEG polymers having an average molecular weight of between about 20 to about 40kDa which have hepatic and renal clearance.
[00142] The PEG groups are polydisperse and have a distribution of molecular weights. Thus, any characterization of a PEG group should be interpreted in light of the polydispersity of PEG, unless otherwise stated.
[00143] In some embodiments, the selective delivery molecule is conjugated to a dextran. In some embodiments, the selective delivery molecule is conjugated to a dextran having a molecular weight of approximately 70kDa. In some embodiments, dextran conjugates, being a mixture of molecular weights, are difficult to synthesize and purify reproducibly.
[00144] In some embodiments, the selective delivery molecule is conjugated to streptavidin.
[00145] In some embodiments, the selective delivery molecule is conjugated to a fifth generation PAMAM dendrimer. [00146] In some embodiments, a carrier is capped. In some embodiments, capping a carrier improves the pharmacokinetics and reduces cytotoxicity of a carrier by adding hydrophilicity. In some embodiments, the cap is selected from: Acetyl, succinyl, 3-hydroxypropionyl, 2-sulfobenzoyl, glycidyl, PEG-2, PEG-4, PEG-8 and PEG-12.
Portion X (Extracellular Cleavable Linkers)
[00147] In some embodiments, X is a linker consisting of one or more amino acids is used to join peptide sequence A (i.e., the sequence designed to inhibit the delivery action of peptide B) and peptide sequence B. Generally the peptide linker will have no specific biological activity other than to join the molecules or to preserve some minimum distance or other spatial relationship between them. However, the constituent amino acids of the linker may be selected to influence some property of the molecule such as the folding, net charge, or hydrophobicity.
[00148] In live cells, an intact selective delivery molecule disclosed herein may not be able to enter the cell because of the presence of portion of A. Thus, a strictly intracellular process for cleaving X would be ineffective to cleave X in healthy cells since portion of A, preventing uptake into cells, would not be effectively cleaved by intracellular enzymes in healthy cells since it would not be taken up and would not gain access to such intracellular enzymes. However, where a cell is injured or diseased (e.g., cancerous cells, hypoxic cells, ischemic cells, apoptotic cells, necrotic cells) such intracellular enzymes leak out of the cell and cleavage of A would occur, allowing entry of portion of B and/or cargo into the cell, effecting targeted delivery of portion of B and/or cargo D to neighboring cells. In some embodiments, X is cleaved in the extracellular space.
[00149] In some embodiments, the fact that capillaries are often leaky around tumors and other trauma sites enhances the ability of high molecular weight molecules (e.g., molecular weight of about 30 kDa or more) to reach the interstitial compartment. In some embodiments, cells that do not express the relevant protease but that are immediately adjacent to expressing cells pick up cargo from a selective delivery molecule because linkage of a X linker is typically extracellular. In some embodiments, such bystander targeting is beneficial in the treatment of tumors because of the heterogeneity of cell phenotypes and the wish to eliminate as high a percentage of suspicious cells as possible.
[00150] In some embodiments, X is a cleavable linker.
[00151] In some embodiments, the X linker is flexible. In some embodiments, the linker is rigid.
[00152] In some embodiments, the X linker comprises a linear structure. In some embodiments, the X linker comprises a non-linear structure. In some embodiments, the X linker comprises a branched structure. In some embodiments, the X linker comprises a cyclic structure. [00153] In some embodiments, X is about 5 to about 30 atoms in length. In some embodiments, X is about 6 atoms in length. In some embodiments, X is about 8 atoms in length. In some
embodiments, X is about 10 atoms in length. In some embodiments, X is about 12 atoms in length. In some embodiments, X is about 14 atoms in length. In some embodiments, X is about 16 atoms in length. In some embodiments, X is about 18 atoms in length. In some embodiments, X is about 20 atoms in length. In some embodiments, X is about 25 atoms in length. In some embodiments, X is about 30 atoms in length.
[00154] In some embodiments, the linker binds peptide portion of A (i.e., the peptide sequence which prevents cellular uptake) to peptide portion of B (i.e., the delivery sequence) by a covalent linkage. In some embodiments, the covalent linkage comprises an ether bond, thioether bond, amine bond, amide bond, oxime bond, hydrazone bond, carbon-carbon bond, carbon-nitrogen bond, carbon-oxygen bond, or carbon-sulfur bond.
[00155] In some embodiments, X comprises a peptide linkage. The peptide linkage comprises L- amino acids and/or D-amino acids. In embodiments of the invention, D-amino acids are preferred in order to minimize immunogenicity and nonspecific cleavage by background peptidases or proteases. Cellular uptake of oligo-D-arginine sequences is known to be as good as or better than that of oligo-L-arginines.
[00156] In some embodiments, a X linker is designed for cleavage in the presence of particular conditions or in a particular environment. In preferred embodiments, a X linker is cleavable under physiological conditions. Cleavage of such a X linker may, for example, be enhanced or may be affected by particular pathological signals or a particular environment related to cells in which cargo delivery is desired. The design of a X linker for cleavage by specific conditions, such as by a specific enzyme, allows the targeting of cellular uptake to a specific location where such conditions obtain. Thus, one important way that selective delivery molecules provide specific targeting of cellular uptake to desired cells, tissues, or regions is by the design of the linker portion X to be cleaved by conditions near such targeted cells, tissues, or regions.
[00157] In some embodiments, X is a pH-sensitive linker. In some embodiments, X is cleaved under basic pH conditions. In some embodiments, X is cleaved under acidic pH conditions. In some embodiments, X is cleaved by a protease, a matrix metalloproteinase, or a combination thereof. In some embodiments, X is cleaved by a reducing agent.
[00158] In some embodiments, X is cleaved by an MMP. The hydrolytic activity of matrix metalloproteinases (MMPs) has been implicated in the invasive migration of metastatic tumor cells. In certain instances, MMPs are found near sites of inflammation. In certain instances, MMPs are found near sites of stroke (i.e., a disorder characterized by brain damage following a decrease in blood flow). Thus, uptake of molecules having features of the invention are able to direct cellular uptake of cargo (at least one D moiety) to specific cells, tissues, or regions having active MMPs in the extracellular environment. In some embodiments, a X linker that includes the amino-acid sequences PLG-C(Me)-AG (SEQ ID NO: 1), PLGLAG (SEQ ID NO: 2) which are cleaved by the metalloproteinase enzymes MMP-2, MMP-9, or MMP-7 (MMPs involved in cancer and inflammation).
[00159] In some embodiments, X is cleaved by proteolytic enzymes or reducing environment, as may be found near cancerous cells. Such an environment, or such enzymes, are typically not found near normal cells.
[00160] In some embodiments, X is cleaved by serine proteases including but not limited to thrombin.
[00161] In some embodiments, X is cleaved in or near tissues suffering from hypoxia. In some embodiments, cleavage in or near hypoxic tissues enables targeting of cancer cells and cancerous tissues, infarct regions, and other hypoxic regions. In some embodiments, X comprises a disulfide bond. In some embodiments, a linker comprising a disulfide bond is preferentially cleaved in hypoxic regions and so targets cargo delivery to cells in such a region. Hypoxia is thought to cause cancer cells to become more resistant to radiation and chemotherapy, and also to initiate angiogenesis. In a hypoxic environment in the presence of, for example, leaky or necrotic cells, free thiols and other reducing agents become available extracellularly, while the 02 that normally keeps the extracellular environment oxidizing is by definition depleted. In some embodiments, this shift in the redox balance promotes reduction and cleavage of a disulfide bond within a X linker. In addition to disulfide linkages which take advantage of thiol-disulfide equilibria, linkages including quinones that fall apart when reduced to hydroquinones are used in a X linker designed to be cleaved in a hypoxic environment.
[00162] In some embodiments, X is cleaved in a necrotic environment. Necrosis often leads to the release of enzymes or other cell contents that may be used to trigger cleavage of a X linker. In some embodiments, cleavage of X by necrotic enzymes (e.g., by calpains) allows cargo to be taken up by diseased cells and by neighboring cells that had not yet become fully leaky.
[00163] In some embodiments, X is an acid-labile linker. In some embodiments, X comprises an acetal or vinyl ether linkage. Acidosis is observed in sites of damaged or hypoxic tissue, due to the Warburg shift from oxidative phosphorylation to anaerobic glycolysis and lactic acid production. In some embodiments, acidosis is used as a trigger of cargo uptake by replacing some of the arginines within B by histidines, which only become cationic below pH 7. [00164] It will be understood that a linker X disclosed herein may include non-standard amino acids, such as, for example, hydroxylysine, desmosine, isodesmosine, or other non-standard amino acids. A linker disclosed herein may include modified amino acids, including post-translationally modified amino acids such as, for example, methylated amino acids (e.g., methyl histidine, methylated forms of lysine, etc.), acetylated amino acids, amidated amino acids, formylated amino acids, hydroxylated amino acids, phosphorylated amino acids, or other modified amino acids. A linker disclosed herein may also include peptide mimetic moieties, including portions linked by non-peptide bonds and amino acids linked by or to non-amino acid portions.
[00165] In some embodiments, the linker X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac). In some embodiments, the linker X comprises the amino acid sequence PLGLAG. In some embodiments, the linker X comprises the amino acid sequence PLG-C(me)-AG. In some embodiments, the linker X comprises the amino acid sequence PLGxAG, wherein x is any amino acid (naturally-occuring or non-naturally occurring). In some embodiments, the linker X comprises the amino acid sequence RPLALWRS. In some embodiments, the linker X comprises the amino acid sequence ESPAYYTA. In some embodiments, the linker X comprises the amino acid sequence DPRSFL. In some embodiments, the linker X comprises the amino acid sequence PPRSFL. In some embodiments, the linker X comprises the amino acid sequence RLQLKL. In some embodiments, the linker X comprises the amino acid sequence RLQLK(Ac).
[00166] In some embodiments, the linker X comprises a peptide selected from: PR(S/T)(L/I)(S/T), where the letters in parentheses indicate that either one of the indicated amino acids may be at that position in the sequence); GGAANLVRGG; SGRIGFLRTA; SGRSA; GFLG; ALAL; FK;
PIC(Et)F-F, where C(Et) indicates S-ethylcysteine (a cysteine with an ethyl group attached to the thiol) and the "-" indicates the typical cleavage site in this and subsequent sequences);
GGPRGLPG; HSSKLQ; LVLA-SSSFGY; GVSQNY-PIVG; GVVQA-SCRLA; f(Pip)R-S, where "f ' indicates D-phenylalanine and "Pip" indicates piperidine-2-carboxylic acid (pipecolinic acid, a proline analog having a six-membered ring); DEVD; GWEHDG; RPLALWRS, or a combination thereof.
[00167] In some embodiments, X is cleaved under hypoxic conditions. In some embodiments, X comprises a disulfide linkage. In some embodiments, X comprises a quinine.
[00168] In some embodiments, X is cleaved under necrotic conditions. In some embodiments, X comprises a molecule cleavable by a calpain.
[00169] In some embodiments, X comprises 6-aminohexanoyl, 5-(amino)-3-oxapentanoyl, or a combination thereof. In some embodiments, X comprises a disulfide linkage. [00170] In some embodiments, the linker is an alkyl. In some embodiments, the linker is heteroalkyl.
[00171] In some embodiments, the linker is an alkylene. In some embodiments, the linker is an alkenylene. In some embodiments, the linker is an alkynylene. In some embodiments, the linker is a heteroalkylene.
[00172] In some embodiments, a selective delivery molecules disclosed herein comprises a single of linker. Use of a single mechanism to mediate uptake of both imaging and therapeutic cargoes is particularly valuable, because imaging with noninjurious tracer quantities can be used to test whether a subsequent therapeutic dose is likely to concentrate correctly in the target tissue.
[00173] In some embodiments, a selective delivery molecules disclosed herein comprises a plurality of linkers. Where a selective delivery molecule disclosed herein includes multiple X linkages, separation of portion of A from the other portions of the molecule requires cleavage of all X linkages. Cleavage of multiple X linkers may be simultaneous or sequential. Multiple X linkages may include X linkages having different specificities, so that separation of portion of A from the other portions of the molecule requires that more than one condition or environment ("extracellular signals") be encountered by the molecule. Cleavage of multiple X linkers thus serves as a detector of combinations of such extracellular signals. For example, a selective delivery molecule may include two linker portions Xa and Xb connecting basic portion of B with acidic portion of A. Both X linkers a and Xb must be cleaved before acidic portion of A is separated from basic portion of B allowing entry of portion of B and cargo moiety C (if any) to enter a cell. It will be understood that a linker region may link to either a basic portion of B or a cargo moiety C independently of another linker that may be present, and that, where desired, more than two linker regions X may be included.
[00174] Combinations of two or more X linkers may be used to further modulate the targeting and delivery of molecules to desired cells, tissue or regions. Combinations of extracellular signals are used to widen or narrow the specificity of the cleavage of X linkers if desired. Where multiple X linkers are linked in parallel, the specificity of cleavage is narrowed, since each X linker must be cleaved before portion of A may separate from the remainder of the molecule. Where multiple X linkers are linked in series, the specificity of cleavage is broadened, since cleavage of any one X linker allows separation of portion of A from the remainder of the molecule. For example, in order to detect either a protease OR hypoxia (i.e., to cleave X in the presence of either protease or hypoxia), a X linker is designed to place the protease- sensitive and reduction-sensitive sites in tandem, so that cleavage of either would suffice to allow separation of the acidic portion of A. Alternatively, in order to detect the presence of both a protease AND hypoxia (i.e., to cleave X in the presence of both protease and hypoxia but not in the presence of only one alone), a X linker is designed to place the protease sensitive site between at least one pair of cysteines that are disulfide- bonded to each other. In that case, both protease cleavage and disulfide reduction are required in order to allow separation of portion of A.
Portion Y linker (Intracellular Cleavable Linkers)
[00175] In some embodiments, Y is a linker consisting of one or more amino acids is used to join Cargo (D) to the remainder of the SDM. In some embodiments, Y is a linker consisting of one or more amino acids is used to join Cargo (D) to portion B. Generally the peptide linker will have no specific biological activity other than to join the molecules or to preserve some minimum distance or other spatial relationship between them. However, the constituent amino acids of the linker may be selected to influence some property of the molecule such as the folding, net charge, or hydrophobicity.
[00176] In some embodiments, the linker binds cargo portion of D to peptide portion of B (i.e., the delivery sequence) by a covalent linkage. In some embodiments, the covalent linkage comprises an ether bond, thioether bond, amine bond, amide bond, oxime bond, hydrazone bond, carbon-carbon bond, carbon-nitrogen bond, carbon-oxygen bond, or carbon-sulfur bond.
[00177] In some embodiments, the Y linker is flexible. In some embodiments, the Y linker is rigid. In some embodiments, the Y linker comprises a linear structure. In some embodiments, the Y linker comprises a non-linear structure. In some embodiments, the Y linker comprises a branched structure. In some embodiments, the linker comprises a cyclic structure.
[00178] In some embodiments, Y linker comprises a peptide linkage. The peptide linkage comprises L-amino acids and/or D-amino acids. In embodiments, D-amino acids are preferred in order to minimize immunogenicity and nonspecific cleavage by background peptidases or proteases. Cellular uptake of oligo-D-arginine sequences is known to be as good as or better than that of oligo-L-arginines.
[00179] In some embodiments, a Y linker is designed for cleavage in the presence of particular conditions or in a particular environment. In some embodiments, a Y linker is cleavable by an intracellular protease. In some embodiments, Y is cleavable by an intracellular protease. In some embodiments, a Y linker is cleavable by a lysosomal protease. In some embodiments, the intracellular protease is a cysteine protease. In some embodiments, the intracellular protease is an aspartyl protease. In some embodiments, the intracellular protease is a serine protease. In some embodiments, the cysteine protease is a caspase, a cathepsin, calpain, papain or a legumain. In some embodiments, the intracellular protease is an initiator caspase. In some embodiments, the intracellular protease is an effector caspase. In some embodiments, the Y linker is cleavable by a protease selected from among cathepsin B, cathepsin L, cathepsin H, cathepsin K, cathepsin W, cathepsin C, cathepsin F, cathepsin V, cathepsin X, cathepsin S, cathepsin D, cathepsin G, HCP-1, HCP-2, dipeptidyl-peptidase I, MEROPS C 13, CED-3 peptidase, caspase 2, caspase 3, caspase 6, caspase 7, caspase 8, caspase 9, caspase 10, caspase 11; caspase 12, caspase 13, and caspase 14. In some embodiments, the Y linker is cleavable by a protease selected from among cathepsin B, cathepsin L, caspase 3, caspase 7, caspase 8, and caspase 9. In some embodiments, a Y linker is cleavable by Cathepsin B a dipeptidyl carboxypeptidase. In some embodiments the linker has a lysine, citrulline, or arginine residue at the PI position and a large hydrophobic residue at the PI ' position.
[00180] In some embodiments, the Y linker comprises an acid sensitive chemical linker. In some embodiments, acid sensitive chemical linker is hydrazone or a derivative thereof. In some embodiments, a Y linker comprises a self-immolative spacer. In some embodiments, the self- immolative spacer is of sufficient length to prevent the occurrence of steric hindrance between the B portion of the SDM and the therapeutic cargo. In some embodiments, Y comprises a p- aminobenzyl alcohol (PABOH) spacer or a derivative thereof. In some embodiments, Y comprises a p-aminobenzyl carbonyl (PABC) spacer or a derivative thereof. In some embodiments, Y comprises a branched bis(hydroxymethyl)styrene (BHMS) spacer or a derivative thereof. In some embodiments, Y comprises a 2-aminoimidazol-5 -methanol derivative or an ortho or para- aminobenzylacetal spacer. In some embodiments Y comprises 2,6-bishydroxymethyl-p-cresol or hemithioaminal derivatives.
[00181] In some embodiments, the Y linker comprises the lysosomally cleavable peptide. In some embodiments, the Y linker comprises the lysosomally cleavable dipeptide Phe-Arg. In some embodiments, the Y linker comprises the lysosomally cleavable dipeptide Phe-Lys. In some embodiments, the Y linker comprises the lysosomally cleavable dipeptide Val-Cit (1-citrulline). In some embodiments, the Y linker comprises the lysosomally cleavable tetrapeptide Gly-Phe-Leu- Gly. In some embodiments, the Y linker comprises the lysosomally cleavable tetrapeptide Ala-Leu- Ala-Leu.
[00182] In some embodiments, the Y linker comprises the lysosomally cleavable peptide and a self-immolative spacer.
[00183] In some embodiments, Y is a pH-sensitive linker. In some embodiments, Y is cleaved under acidic pH conditions. In some embodiments, Y is cleaved under acidic pH conditions of the lysosome. [00184] It will be understood that a Y linker disclosed herein may include non-standard amino acids, such as, for example, hydroxylysine, desmosine, isodesmosine, or other non-standard amino acids. A linker disclosed herein may include modified amino acids, including post-translationally modified amino acids such as, for example, methylated amino acids (e.g., methyl histidine, methylated forms of lysine, etc.), acetylated amino acids, amidated amino acids, formylated amino acids, hydroxylated amino acids, phosphorylated amino acids, or other modified amino acids. A linker disclosed herein may also include peptide mimetic moieties, including portions linked by non-peptide bonds and amino acids linked by or to non-amino acid portions.
[00185]
Imaging Agents
[00186] In some embodiments, an SDM provided herein is conjugated to an imaging agent. In some embodiments, the imaging agent is conjugated to portion of A, portion of B or both portions A and B. In some embodiments, the imaging agent is conjugated to the target ligand.
[00187] In some embodiments, an imaging agent is a dye. In some embodiments, an imaging agent is a fluorescent moiety. In some embodiments, a fluorescent moiety is selected from: a fluorescent protein, a fluorescent peptide, a fluorescent dye, a fluorescent material or a combination thereof.
[00188] All fluorescent moieties are encompassed within the term "fluorescent moiety." Specific examples of fluorescent moieties given herein are illustrative and are not meant to limit the fluorescent moieties for use with the targeting molecules disclosed herein.
[00189] Examples of fluorescent dyes include, but are not limited to, xanthenes (e.g., rhodamines, rhodols and fluoresceins, and their derivatives); bimanes; coumarins and their derivatives (e.g., umbelliferone and aminomethyl coumarins); aromatic amines (e.g., dansyl; squarate dyes);
benzofurans; fluorescent cyanines; indocarbocyanines; carbazoles; dicyanomethylene pyranes; polymethine; oxabenzanthrane; xanthene; pyrylium; carbostyl; perylene; acridone; quinacridone; rubrene; anthracene; coronene; phenanthrecene; pyrene; butadiene; stilbene; porphyrin;
pthalocyanine; lanthanide metal chelate complexes; rare-earth metal chelate complexes; and derivatives of such dyes.
[00190] Examples of fluorescein dyes include, but are not limited to, 5-carboxyfluorescein, fluorescein-5-isothiocyanate, fiuorescein-6-isothiocyanate and 6-carboxyfluorescein.
[00191] Examples of rhodamine dyes include, but are not limited to, tetramethylrhodamine-6- isothiocyanate, 5-carboxytetramethylrhodamine, 5-carboxy rhodol derivatives, tetramethyl and tetraethyl rhodamine, diphenyldimethyl and diphenyldiethyl rhodamine, dinaphthyl rhodamine, rhodamine 101 sulfonyl chloride (sold under the trade name of TEXAS RED®). [00192] Examples of cyanine dyes include, but are not limited to, Cy3, Cy3B, Cy3.5, Cy5, Cy5.5, Cy7, IRDYE680, Alexa Fluor 750, IRDye800CW, ICG.
[00193] Examples of fluorescent peptides include GFP (Green Fluorescent Protein) or derivatives of GFP (e.g., EBFP, EBFP2, Azurite, mKalamal, ECFP, Cerulean, CyPet, YFP, Citrine, Venus, YPet).
[00194] Fluorescent labels are detected by any suitable method. For example, a fluorescent label may be detected by exciting the fluorochrome with the appropriate wavelength of light and detecting the resulting fluorescence, e.g., by microscopy, visual inspection, via photographic film, by the use of electronic detectors such as charge coupled devices (CCDs), photomultipliers, etc.
[00195] In some embodiments, the imaging agent is labeled with a positron-emitting isotope (e.g.,18F) for positron emission tomography (PET), gamma-ray isotope (e.g., 99mTc) for single photon emission computed tomography (SPECT), or a paramagnetic molecule or nanoparticle (e.g.,Gd3+ chelate or coated magnetite nanoparticle) for magnetic resonance imaging (MRI).
[00196] In some embodiments, the imaging agent is labeled with: a gadolinium chelate, an iron oxide particle, a super paramagnetic iron oxide particle, an ultra small paramagnetic particle, a manganese chelate or gallium containing agent.
[00197] Examples of gadolinium chelates include, but are not limited to diethylene triamine pentaacetic acid (DTPA), l ,4,7,10-tetraazacyclododecane-l ,4,7,10-tetraacetic acid (DOTA), and l,4,7-triazacyclononane-N,N',N"-triacetic acid (NOT A).
[00198] In some embodiments, the imaging agent is a near-infrared fluorophore for near-infra red (near-IR) imaging, a luciferase (firefly, bacterial, or coelenterate) or other luminescent molecule for bioluminescence imaging, or a perfluorocarbon- filled vesicle for ultrasound.
[00199] In some embodiments, the imaging agent is a nuclear probe. In some embodiments, the imaging agent is a SPECT or PET radionuclide probe. In some embodiments, the radionuclide probe is selected from: a technetium chelate, a copper chelate, a radioactive fluorine, a radioactive iodine, a indiuim chelate.
[00200] Examples of Tc chelates include, but are not limited to HYNIC, DTPA, and DOTA.
[00201] In some embodiments, the imaging agent contains a radioactive moiety, for example a radioactive isotope such as 211At, 1 1I, 1251, 90Y, 186Re, 18 Re, 153Sm, 212Bi, 32P, 64Cu radioactive isotopes of Lu, and others.
[00202] In some embodiments, a selective delivery molecule according to Formulas I-VI comprising an imaging agent is employed in guided surgery. In some embodiments, the selective delivery molecule preferentially localized to cancerous, or other undesirable tissues (i.e. necrotic tissues). In some embodiments, a selective delivery molecule according to Formula I comprising an imaging agent is employed in a guided surgery to remove colorectal cancer. In some embodiments, guided surgery employing the selective delivery molecule allows a surgeon to excise as little healthy (i.e., non-cancerous) tissue as possible. In some embodiments, guided surgery employing the selective delivery molecule allows a surgeon to visualize and excise more cancerous tissue than the surgeon would have been able to excise without the presence of the selective delivery molecule. In some embodiments, the surgery is fluorescence-guided surgery.
Exemplary Selective Delivery Molecules
[00203] In some embodiments, the selective delivery molecule comprises a structure selected from SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM-110, SDM-111 , SDM-112, SDM-1 13, SDM-114, SDM-115, SDM-1 16, SDM- 1 17, SDM-1 18, SDM-119, SDM-120, SDM-121 , SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM- 134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM- 141 , SDM-142, SDM-143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM- 151 , SDM-152, and SDM-153. In some embodiments, the selective delivery molecule is a derivative of SDM-101 , SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM- 108, SDM-109, SDM-110, SDM-111 , SDM-112, SDM-1 13, SDM-114, SDM-115, SDM-1 16, SDM-1 17, SDM-118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM- 125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131 , SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141 , SDM- 142, SDM-143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151, SDM-152, and SDM-153. In some embodiments, the selective delivery molecule the derivative comprises an imaging agent. In some embodiments, the selective delivery molecule the derivative comprises an additional therapeutic agent.
Figure imgf000074_0001
Figure imgf000075_0001
Figure imgf000076_0001
Figure imgf000077_0001
Figure imgf000078_0001
Figure imgf000079_0001
Figure imgf000080_0001
Figure imgf000081_0001
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Figure imgf000083_0001
Figure imgf000084_0001
Figure imgf000085_0001
Figure imgf000086_0001
Figure imgf000087_0001
Figure imgf000088_0001
[00204] In some embodiments, the selective delivery molecule comprises a structure selected from: SDM-1 , SDM-2, SDM-3, SDM-4, SDM-5, SDM-6, SDM-7, SDM-8, SDM-9, SDM-10, SDM-1 1 , SDM-12, SDM-13, SDM-14, SDM-15, SDM-16, SDM-17, SDM-18, SDM-19, SDM-20, SDM-21 , SDM-22, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-28, SDM-29, SDM-30, SDM-31 , SDM-32, SDM-33, SDM-34, SDM-35, SDM-36, SDM-37, SDM-38, SDM-39, SDM-40, SDM-41 , SDM-42, SDM-43, SDM-44, SDM-45, SDM-46, SDM-47, SDM-48, SDM-49, SDM-50, SDM- 1 , SDM-52, SDM-53, SDM-54, SDM-55, SDM-56, SDM-57, SDM-58, SDM-59, SDM-60, and SDM-61 (see International PCT Pub. No. WO2013/019681). In some embodiments, the selective delivery molecule comprises a structure selected from: SDM-14, SDM-15, SDM-23, SDM-24, SDM-25, SDM-26, SDM-27, SDM-32, or SDM-35. In certain embodiments, the selective delivery molecule is derived from Peptide P-l, P-2, P-3, P-4, P-5, P-6, P-7, P-8, P-9, P-10, P-l 1, P- 12, P-13, P-14, P-15, P-16, P-17, P-18, P-19, P-20, P-21 , P-21 , or P-3.
Figure imgf000090_0001
Figure imgf000091_0001
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Figure imgf000092_0001
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-102-
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-103-
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Figure imgf000108_0001
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Figure imgf000109_0001
-107- Further Modifications
[00205] In some embodiments, the antibody-conjugated SDMs described herein are optionally conjugated to high molecular weight molecules that increase the multivalency and avidity of labeling. In some embodiments, the high molecular weight molecules are water-soluble polymers. Examples of suitable water-soluble polymers include, but are not limited to, peptides, saccharides, poly(vinyls), poly(ethers), poly(amines), poly(carboxylic acids) and the like. In some embodiments, the water-soluble polymer is dextran, polyethylene glycol (PEG), polyoxyalkylene, polysialic acid, starch, or hydroxyethyl starch. Any suitable method is used to conjugate peptides to water-soluble polymers ( see Hermanson G., Bioconjugate Techniques 2nd Ed., Academic Press, Inc. 2008).
Pharmaceutical Compositions
[00206] Disclosed herein, in certain embodiments, are pharmaceutical compositions comprising any of SDMs as disclosed herein. In some embodiments, the pharmaceutical compositions comprising an SDM comprises an SDM of any of Formulas I-VI and a pharmaceutically acceptable carrier.
[00207] Disclosed herein, in certain embodiments, are pharmaceutical compositions comprising any of the antibody-conjugated SDMs as disclosed herein. In some embodiments, the
pharmaceutical compositions comprising an antibody-conjugated SDM comprises a targeting antibody conjugated to an SDM of any of Formulas I-VI and a pharmaceutically acceptable carrier.
[00208] Pharmaceutical compositions herein are formulated using one or more physiologically acceptable carriers including excipients and auxiliaries which facilitate processing of the active agents into preparations which are used pharmaceutically. Proper formulation is dependent upon the route of administration chosen. A summary of pharmaceutical compositions is found, for example, in Remington: The Science and Practice of Pharmacy, Nineteenth Ed (Easton, Pa.: Mack Publishing Company, 1995); Hoover, John E., Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, Pennsylvania 1975; Liberman, H.A. and Lachman, L., Eds.,
Pharmaceutical Dosage Forms, Marcel Decker, New York, N.Y., 1980; and Pharmaceutical Dosage Forms and Drug Delivery Systems, Seventh Ed. (Lippincott Williams & Wilkins, 1999).
[00209] In certain embodiments, a pharmaceutical composition disclosed herein further comprises a pharmaceutically acceptable diluent(s), excipient(s), or carrier(s). In some embodiments, the pharmaceutical compositions includes other medicinal or pharmaceutical agents, carriers, adjuvants, such as preserving, stabilizing, wetting or emulsifying agents, solution promoters, salts for regulating the osmotic pressure, and/or buffers. In addition, the pharmaceutical compositions also contain other therapeutically valuable substances. WSGR Docket No. 39088-711.601
[00210] In certain embodiments, a pharmaceutical composition disclosed herein is administered to a subject by any suitable administration route, including but not limited to, parenteral (intravenous, subcutaneous, intraperitoneal, intramuscular, intravascular, intrathecal, intravitreal, infusion, or local) administration.
[00211] Formulations suitable for intramuscular, subcutaneous, peritumoral, or intravenous injection include physiologically acceptable sterile aqueous or non-aqueous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Examples of suitable aqueous and non-aqueous carriers, diluents, solvents, or vehicles including water, ethanol, polyols (propyleneglycol, polyethylene-glycol, glycerol, cremophor and the like), suitable mixtures thereof, vegetable oils (such as olive oil) and injectable organic esters such as ethyl oleate. Proper fluidity is maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants. Formulations suitable for subcutaneous injection also contain optional additives such as preserving, wetting, emulsifying, and dispensing agents.
[00212] For intravenous injections, an active agent is optionally formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hank's solution, Ringer's solution, or physiological saline buffer.
[00213] Parenteral injections optionally involve bolus injection or continuous infusion.
Formulations for injection are optionally presented in unit dosage form, e.g., in ampoules or in multi dose containers, with an added preservative. In some embodiments, the pharmaceutical composition described herein are in a form suitable for parenteral injection as a sterile suspensions, solutions or emulsions in oily or aqueous vehicles, and contain formulatory agents such as suspending, stabilizing and/or dispersing agents. Pharmaceutical formulations for parenteral administration include aqueous solutions of an active agent in water soluble form. Additionally, suspensions are optionally prepared as appropriate oily injection suspensions.
[00214] In some embodiments, the pharmaceutical composition described herein is in unit dosage forms suitable for single administration of precise dosages. In unit dosage form, the formulation is divided into unit doses containing appropriate quantities of an active agent disclosed herein. In some embodiments, the unit dosage is in the form of a package containing discrete quantities of the formulation. Non-limiting examples are packaged tablets or capsules, and powders in vials or ampoules. In some embodiments, aqueous suspension compositions are packaged in single-dose non-reclosable containers. Alternatively, multiple-dose reclosable containers are used, in which case it is typical to include a preservative in the composition. By way of example only, WSGR Docket No. 39088-711.601
formulations for parenteral injection are presented in unit dosage form, which include, but are not limited to ampoules, or in multi dose containers, with an added preservative.
[00215] Methods of Use
[00216] The SDMs of Formulas I-VI and carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, allow the targeted delivery of therapeutic agents and/or imaging agents to specific cells and/or tissues. The molecules comprise a basic peptide sequence (B) which is designed to be transported across a cellular membrane, an acidic peptide sequence (A) which inhibits uptake of peptide B into cells, a linker X which is cleavable under specific conditions, cargo moieties (at least DA and DB) bound to peptides A and B, or X and a macromolecular carrier. In some embodiments, cleavage of the linker X linker frees peptide B from peptide A and allows the transport of peptide B (and any cargo attached thereto) across a cellular membrane. In some embodiments, the selective delivery molecules of Formulas I-IV enable targeted delivery of one or more cargos (e.g., therapeutic agents or imaging agents) to a cell tissue.
[00217] Disclosed herein, in certain embodiments, are methods of delivering cargo to a tissue of interest, comprising contacting the tissue of interest with an SDM of any of Formulas I-VI.
Disclosed herein, in certain embodiments, are methods of delivering cargo to a tissue of interest, comprising contacting the tissue of interest with an antibody-conjugated SDM comprising a targeting antibody conjugated to an SDM of any of Formulas I-VI.
Tissue of Interest
[00218] In some embodiments, the tissue of interest is cancerous tissue (or, cancer). In some embodiments, the cancerous tissue is: breast cancer tissue, colon cancer tissue, squamous cell carcinoma tissue, prostate cancer tissue, melanoma tissue, or thyroid cancer tissue. In some embodiments, the cancerous tissue is breast cancer tissue. In some embodiments, the cancerous tissue is colon cancer tissue.
[00219] In some embodiments, the tissue of interest is an inflamed tissue. In some embodiments, some embodiments, the inflamed tissue is the result if acute or chronic inflammation. In some embodiments, the inflamed tissue is caused by an inflammatory disease is or is associated with an inflammatory disease. In some embodiments, the inflamed tissue is caused by an inflammatory disease is or is associated with rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, sepsis, erythema nodosum leprosum, multiple sclerosis, psoriasis, systemic lupus erythematosis, type I diabetes, atherosclerosis, encephalomyelitis, Alzheimer's disease, stroke, traumatic brain injury, Parkinson's disease or septic shock.
Therapeutic Uses WSGR Docket No. 39088-711.601
[00220] The SDMs of Formulas I-VI and carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, allow the targeted delivery of therapeutic agents to specific cells and/or tissues (e.g., cancerous tissues). The molecules comprise a basic peptide sequence (B) which is designed to be transported across a cellular membrane, an acidic peptide sequence (A) which inhibits uptake of peptide B into cells, a linker X which is cleavable under specific conditions, therapeutic agents bound to peptides A and B, or X and a macromolecular carrier. In some embodiments, cleavage of the linker X linker frees peptide B from peptide A and allows the transport of peptide B (and any therapeutic agents attached thereto) across a cellular membrane. In some embodiments, the SDMs of Formulas I-VI and carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, enable targeted delivery of one or more therapeutic agents to a cell or tissue. In some embodiments, targeted delivery of a therapeutic agent to a cell or tissue enables a medical professional to treat a specific tissue.
[00221] In some embodiments, targeted delivery of a therapeutic agent to a cell or tissue enables a medical professional to treat a specific tissue (e.g., cancerous tissue). In some embodiments, targeted delivery of a therapeutic agent to a cell or tissue decreases the dosage of the therapeutic agent. In some embodiments, targeted delivery of a therapeutic agent to a cell or tissue decreases contact of the therapeutic agent with healthy tissue. In some embodiments, targeted delivery of a therapeutic agent to a cell or tissue decreases unwanted side-effects arising from use of high concentrations of a therapeutic agent or contact. In some embodiments, targeted delivery of a therapeutic agent to a cell or tissue decreases unwanted side-effects arising from contact between the therapeutic agent and healthy tissue.
[00222] In some embodiments, an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, is employed for the treatment of cancer.
[00223] In some embodiments, the cancer is AIDS-related cancers (e.g., AIDS-related lymphoma), anal cancer, basal cell carcinoma, bile duct cancer (e.g., extrahepatic), bladder cancer, bone cancer, (osteosarcoma and malignant fibrous histiocytoma), breast cancer, cervical cancer, colon cancer, colorectal cancer, endometrial cancer (e.g., uterine cancer), ependymoma, esophageal cancer, eye cancer (e.g., intraocular melanoma and retinoblastoma), gastric (stomach) cancer, germ cell tumor, (e.g., extracranial, extragonadal, ovarian), head and neck cancer, leukemia, lip and oral cavity cancer, liver cancer, lung cancer (e.g., small cell lung cancer, non-small cell lung cancer, adenocarcinoma of the lung, and squamous carcinoma of the lung), ovarian cancer, pancreatic cancer, pituitary tumor, prostate cancer, renal cancer, skin cancer, small intestine cancer, squamous cell cancer, testicular cancer, throat cancer, thyroid cancer, urethral cancer, and post-transplant lymphoproliferative disorder (PTLD).
-I l l- WSGR Docket No. 39088-711.601
[00224] In some embodiments, the cancer is a lymphoid cancer (e.g., lymphoma).
[00225] In some embodiments, the cancer is a B-cell cancer. In some embodiments, the cancer is precursor B-cell cancers (e.g., precursor B-lymphoblastic leukemia/lymphoma) and peripheral B- cell cancers (e.g., B-cell chronic lymphocytic leukemia/pro lymphocytic leukemia/small lymphocytic lymphoma (small lymphocytic (SL) NHL), lymphoplasmacytoid
lymphoma/immunocytoma, mantel cell lymphoma, follicle center lymphoma, follicular lymphoma (e.g., cytologic grades: I (small cell), II (mixed small and large cell), III (large cell) and/or subtype: diffuse and predominantly small cell type), low grade/follicular non-Hodgkin's lymphoma (NHL), intermediate grade/follicular NHL, marginal zone B-cell lymphoma (e.g., extranodal (e.g., MALT- type +/- monocytoid B cells) and/or Nodal (e.g., +/- monocytoid B cells)), splenic marginal zone lymphoma (e.g., +/- villous lymphocytes), Hairy cell leukemia, plasmacytoma/plasma cell myeloma (e.g., myeloma and multiple myeloma), diffuse large B-cell lymphoma (e.g., primary mediastinal (thymic) B-cell lymphoma), intermediate grade diffuse NHL, Burkitt's lymphoma, High-grade B-cell lymphoma, Burkitt-like, high grade immunoblastic NHL, high grade lymphoblastic NHL, high grade small non-cleaved cell NHL, bulky disease NHL, AIDS -related lymphoma, and Waldenstrom's macro globulinemia).
[00226] In some embodiments, the cancer is a T-cell and/or putative NK-cell cancer. In some embodiments, the cancer is precursor T-cell cancer (precursor T-lymphoblastic
lymphoma/leukemia) and peripheral T-cell and NK-cell cancers (e.g., T-cell chronic lymphocytic leukemia/prolymphocytic leukemia, and large granular lymphocyte leukemia (LGL) (e.g., T-cell type and/or NK-cell type), cutaneous T-cell lymphoma (e.g., mycosis fungoides/Sezary syndrome), primary T-cell lymphomas unspecified (e.g., cytological categories (e.g., medium-sized cell, mixed medium and large cell), large cell, lymphoepitheloid cell, subtype hepatosplenic γδ T-cell
lymphoma, and subcutaneous panniculitic T-cell lymphoma), angioimmunoblastic T-cell lymphoma (AILD), angiocentric lymphoma, intestinal T-cell lymphoma (e.g., +/- enteropathy associated), adult T-cell lymphoma/leukemia (ATL), anaplastic large cell lymphoma (ALCL) (e.g., CD30+, T- and null-cell types), anaplastic large-cell lymphoma, and Hodgkin's like).
[00227] In some embodiments, the cancer is Hodgkin's disease.
[00228] In some embodiments, the cancer is leukemia. In some embodiments, the cancer is chronic myelocytic I (granulocytic) leukemia, chronic myelogenous, and chronic lymphocytic leukemia (CLL), acute lymphoblastic leukemia (ALL), acute myeloid leukemia, acute lymphocytic leukemia, and acute myelocytic leukemia (e.g., myeloblastic, promyelocytic, myelomonocytic, monocytic, and erythroleukemia). WSGR Docket No. 39088-711.601
[00229] In some embodiments, the cancer is a liquid tumor or plasmacytoma. In some
embodiments, the cancer is extramedullary plasmacytoma, a solitary myeloma, and multiple myeloma. In some embodiments, the plasmacytoma is multiple myeloma.
[00230] In some embodiments, the cancer is lung cancer.
[00231] In some embodiments, the cancer is prostate cancer. In some embodiments, the prostate cancer is an adenocarcinoma. In some embodiments, the prostate cancer is a sarcoma,
neuroendocrine tumor, small cell cancer, ductal cancer, or a lymphoma. In some embodiments, the prostate cancer is stage A prostate cancer (the cancer cannot be felt during a rectal exam). In some embodiments, the prostate cancer is stage B prostate cancer (i.e., the tumor involves more tissue within the prostate, it can be felt during a rectal exam, or it is found with a biopsy that is done because of a high PSA level). In some embodiments, the prostate cancer is stage C prostate cancer (i.e., the cancer has spread outside the prostate to nearby tissues). In some embodiments, the prostate cancer is stage D prostate cancer. In some embodiments, the prostate cancer is androgen independent prostate cancer (AIPC). In some embodiments, the prostate cancer is androgen dependent prostate cancer. In some embodiments, the prostate cancer is refractory to hormone therapy. In some embodiments, the prostate cancer is substantially refractory to hormone therapy. In some embodiments, the prostate cancer is refractory to chemotherapy. In some embodiments, the prostate cancer is metastatic prostate cancer. In some embodiments, the individual is a human who has a gene, genetic mutation, or polymorphism associated with prostate cancer (e.g.,
RNASEL/HPC 1 , ELAC2/HPC2, SR-A/MSRl , CHEK2, BRCA2, PO 1 , OGG1 , MIC-1 , TLR4, and PTEN) or has one or more extra copies of a gene associated with prostate cancer. In some embodiments, the prostate cancer is HER2 positive. In some embodiments, the prostate cancer is HER2 negative.
[00232] In some embodiments, the cancer has metastasized and is characterized by circulating tumor cells.
[00233] In some embodiments, an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, is employed for the treatment of inflammation or an inflammatory disease. In some embodiments, the inflammation is chronic inflammation. In some embodiments, the inflammation is acute inflammation. In some embodiments, inflammation or inflammatory disease is or is associated with rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, sepsis, erythema nodosum leprosum, multiple sclerosis, psoriasis, systemic lupus erythematosis, type I diabetes, atherosclerosis,
encephalomyelitis, Alzheimer's disease, stroke, traumatic brain injury, Parkinson's disease or septic shock. WSGR Docket No. 39088-711.601
[00234] In some embodiments, an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, is employed for the treatment of an autoimmune disease. In some embodiments, the autoimmune disease is Celiac disease, diabetes mellitus type 1, Sarcoidosis, systemic lupus erythematosus (SLE), Sjogren's syndrome, Churg-Strauss Syndrome, Hashimoto's thyroiditis, Graves' disease, idiopathic thrombocytopenic purpura, Addison's Disease, rheumatoid arthritis (RA), Polymyositis (PM), or Dermatomyositis (DM).
Therapeutic Agents
[00235] In some embodiments, a therapeutic agent is selected from: a chemotherapeutic agent, a steroid, an immunotherapeutic agent, a targeted therapy, an anti-inflammatory agent, or a combination thereof.
[00236] In some embodiments, a therapeutic agent is a CD79A inhibitor, a CD79B inhibitor, a CD 19 inhibitor, a Lyn inhibitor, a Syk inhibitor, a PI3K inhibitor, a Blnk inhibitor, a PLCy inhibitor, a PKCp inhibitor, or a combination thereof. In some embodiments, a therapeutic agent is an antibody, B cell receptor signaling inhibitor, a PI3K inhibitor, an IAP inhibitor, an mTOR inhibitor, a radioimmunotherapeutic, a DNA damaging agent, a proteosome inhibitor, a histone deacytlase inhibitor, a protein kinase inhibitor, a hedgehog inhibitor, an Hsp90 inhibitor, a telomerase inhibitor, a Jakl/2 inhibitor, a protease inhibitor, a PKC inhibitor, a PARP inhibitor, or a combination thereof. In some embodiments, a therapeutic agent is a B cell receptor pathway inhibitor. In some embodiments, a therapeutic agent is selected from: chlorambucil, ifosphamide, doxorubicin, mesalazine, thalidomide, lenalidomide, temsirolimus, everolimus, fludarabine, fostamatinib, paclitaxel, docetaxel, ofatumumab, rituximab, dexamethasone, prednisone, CAL-101, ibritumomab, tositumomab, bortezomib, pentostatin, endostatin, bendamustine, chlorambucil, chlormethine, cyclophosphamide, ifosfamide, melphalan, prednimustine, trofosfamide, busulfan, mannosulfan, treosulfan, carboquone, thiotepa, triaziquone, carmustine, fotemustine, lomustine, nimustine, ranimustine, semustine, streptozocin, etoglucid, dacarbazine, mitobronitol, pipobroman, temozolomide, methotrexate, permetrexed, pralatrexate, raltitrexed, cladribine, clofarabine, fludarabine, mercaptopurine, nelarabine, tioguanine, azacitidine, capecitabine, carmofur, cytarabine, decitabine, fluorouracil, gemcitabine, tegafur, vinblastine, vincristine, vindesine, vinflunine, vinorelbine, etoposide, teniposide, demecolcine, docetaxel, paclitaxel, paclitaxel poliglumex, trabectedin, dactinomycin, aclarubicin, daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, pirarubicin, valrubicin, zorubincin, bleomycin, ixabepilone, mitomycin, plicamycin, carboplatin, cisplatin, oxaliplatin, satraplatin, procarbazine, aminolevulinic acid, efaproxiral, methyl aminolevulinate, porfimer sodium, temoporfin, dasatinib, erlotinib, everolimus, gefitinib, imatinib, lapatinib, nilotinib, pazonanib, sorafenib, sunitinib, temsirolimus, alitretinoin, WSGR Docket No. 39088-711.601
altretamine, amzacrine, anagrelide, arsenic trioxide, asparaginase, bexarotene, bortezomib, celecoxib, denileukin diftitox, estramustine, hydroxycarbamide, irinotecan, lonidamine, masoprocol, miltefosein, mitoguazone, mitotane, oblimersen, pegaspargase, pentostatin, romidepsin, sitimagene ceradenovec, tiazofurine, topotecan, tretinoin, vorinostat, diethylstilbenol, ethinylestradiol, fosfestrol, polyestradiol phosphate, gestonorone, medroxyprogesterone, megestrol, buserelin, goserelin, leuprorelin, triptorelin, fulvestrant, tamoxifen, toremifene, bicalutamide, flutamide, nilutamide, aminoglutethimide, anastrozole, exemestane, formestane, letrozole, vorozole, abarelix, degarelix, histamine dihydrochloride, mifamurtide, pidotimod, plerixafor, roquinimex, thymopentin, everolimus, gusperimus, leflunomide, mycophenolic acid, sirolimus, ciclosporin, tacrolimus, azathioprine, lenalidomide, methotrexate, thalidomide, iobenguane, ancestim, filgrastim, lenograstim, molgramostim, pegfilgrastim, sargramostim, interferon alfa natural, interferon alfa-2a, interferon alfa-2b, interferon alfacon-1, interferon alfa-nl, interferon beta natural, interferon beta-la, interferon beta-lb, interferon gamma, peginterferon alfa-2a, peginterferon alfa-2b, aldesleukin, oprelvekin, BCG vaccine, glatiramer acetate, histamine dihydrochloride, immunocyanin, lentinan, melanoma vaccine, mifamurtide, pegademase, pidotimod, plerixafor, poly I:C, poly ICLC, roquinimex, tasonermin, thymopentin, abatacept, abetimus, alefacept, antilymphocyte immunoglobulin (horse), antithymocyte immunoglobulin (rabbit), eculizumab, efalizumab, everolimus, gusperimus, leflunomide, muromab-CD3, mycophenolic acid, natalizumab, sirolimus, adalimumab, afelimomab, certolizumab pegol, etanercept, golimumab, infliximab, anakinra, basiliximab, canakinumab, daclizumab,
mepolizumab, rilonacept, tocilizumab, ustekinumab, ciclosporin, tacrolimus, azathioprine, lenalidomide, methotrexate, thalidomide, adalimumab, alemtuzumab, bevacizumab, cetuximab, certolizumab pegol, , eculizumab, efalizumab, gemtuzumab, ibritumomab tiuxetan, muromonab- CD3, natalizumab, panitumumab, ranibizumab, rituximab, tositumomab, trastuzumab,
catumaxomab, edrecolomab, ofatumumab, muromab-CD3, afelimomab, golimumab, ibritumomab tiuxetan, abagovomab, adecatumumab, alemtuzumab, anti-CD30 monoclonal antibody Xmab2513, anti-MET monoclonal antibody MetMab, apolizumab, apomab, arcitumomab, bispecific antibody 2B1, blinatumomab, brentuximab vedotin, capromab pendetide, cixutumumab, claudiximab, conatumumab, dacetuzumab, denosumab, eculizumab, epratuzumab, epratuzumab, ertumaxomab, etaracizumab, figitumumab, fresolimumab, galiximab, ganitumab, gemtuzumab ozogamicin, glembatumumab, ibritumomab, inotuzumab ozogamicin, ipilimumab, lexatumumab, lintuzumab, lintuzumab, lucatumumab, mapatumumab, matuzumab, milatuzumab, monoclonal antibody CC49, necitumumab, nimotuzumab, ofatumumab, oregovomab, pertuzumab, ramacurimab, ranibizumab, siplizumab, sonepcizumab, tanezumab, tositumomab, trastuzumab, tremelimumab, tucotuzumab WSGR Docket No. 39088-711.601
celmoleukin, veltuzumab, visilizumab, volociximab, zalutumumab, a syk inhibitor (e.g., R788), enzastaurin, dasatinib, erlotinib, everolimus, gefitinib, imatinib, lapatinib, nilotinib, pazonanib, sorafenib, sunitinib, temsirolimus, an angiogenesis inhibitor (e.g., GT-111, JI-101, R1530), a kinase inhibitors (e.g., AC220, AC480, ACE-041, AMG 900, AP24534, Arry-614, AT7519, AT9283, AV- 951, axitinib, AZD1152, AZD7762, AZD8055, AZD8931, bafetinib, BAY 73-4506, BGJ398, BGT226, BI 811283, BI6727, BIBF 1120, BIBW 2992, BMS-690154, BMS-777607, BMS- 863233, BS -461364, CAL-101, CEP-11981, CYC116, DCC-2036, dinaciclib, dovitinib lactate, E7050, EMD 1214063, ENMD-2076, fostamatinib disodium, GSK2256098, GSK690693,
INCB18424, ΓΝ Ο-406, JNJ-26483327, JX-594, KX2-391, linifanib, LY2603618, MGCD265, MK-0457, MK1496, MLN8054, MLN8237, MP470, NMS-1116354, NMS-1286937, ON
01919.Na, OSI-027, OSI-930, Btk inhibitor, PF-00562271, PF-02341066, PF-03814735, PF- 04217903, PF-04554878, PF-04691502, PF-3758309, PHA-739358, PLC3397, progenipoietin, R547, R763, ramucirumab, regorafenib, R05185426, SAR103168, S3333333CH 727965, SGI- 1176, SGX523, SNS-314, TAK-593, TAK-901, TKI258, TLN-232, TTP607, XL147, XL228, XL281R05126766, XL418, XL765), an inhibitor of mitogen-activated protein kinase signaling (e.g., U0126, PD98059, PD184352, PD0325901, ARRY-142886, SB239063, SP600125, BAY 43- 9006, wortmannin, or LY294002), adriamycin, dactinomycin, bleomycin, vinblastine, cisplatin, acivicin, aclarubicin, acodazole hydrochloride, acronine, adozelesin, aldesleukin, altretamine, ambomycin, ametantrone acetate, aminoglutethimide, amsacrine, anastrozole, anthramycin, asparaginase, asperlin, azacitidine, azetepa, azotomycin, batimastat, benzodepa, bicalutamide, bisantrene hydrochloride, bisnafide dimesylate, bizelesin, bleomycin sulfate, brequinar sodium, bropirimine, busulfan, cactinomycin, calusterone, caracemide, carbetimer, carboplatin, carmustine, carubicin hydrochloride, carzelesin, cedefmgol, chlorambucil, cirolemycin, cladribine, crisnatol mesylate, cyclophosphamide, cytarabine, dacarbazine, daunorubicin hydrochloride, decitabine, dexormaplatin, dezaguanine, dezaguanine mesylate, diaziquone, doxorubicin, doxorubicin hydrochloride, droloxifene, droloxifene citrate, dromostanolone propionate, duazomycin, edatrexate, eflornithine hydrochloride, elsamitrucin, enloplatin, enpromate, epipropidine, epirubicin hydrochloride, erbulozole, esorubicin hydrochloride, estramustme, estramustme phosphate sodium, etanidazole, etoposide, etoposide phosphate, etoprine, fadrozole hydrochloride, fazarabine, fenretinide, floxuridine, fludarabine phosphate, fluorouracil, flurocitabine, fosquidone, fostriecin sodium, gemcitabine, gemcitabine hydrochloride, hydroxyurea, idarubicin hydrochloride, ifosfamide, iimofosine, interleukin II (including recombinant interleukin II, or rlL2), interferon alfa- 2a, int interferon alfa -2b, interferon alfa -nl, interferon alfa -n3, interferon beta-1 a, interferon gamma-1 b, iproplatin, irinotecan hydrochloride, lanreotide acetate, letrozole, leuprolide acetate, WSGR Docket No. 39088-711.601
liarozole hydrochloride, lometrexol sodium, lomustine, losoxantrone hydrochloride, masoprocol, maytansine, mechlorethamine hydrochloride, megestrol acetate, melengestrol acetate, melphalan, menogaril, mercaptopurine, methotrexate, methotrexate sodium, metoprine, meturedepa, mitindomide, mitocarcin, mitocromin, mitogillin, mitomalcin, mitomycin, mitosper, mitotane, mitoxantrone hydrochloride, mycophenolic acid, nocodazoie, nogalamycin, ormaplatin, oxisuran, pegaspargase, peliomycin, pentamustine, peplomycin sulfate, perfosfamide, pipobroman, piposulfan, piroxantrone hydrochloride, plicamycin, plomestane, porfrmer sodium, porfiromycin, prednimustine, procarbazine hydrochloride, puromycin, puromycin hydrochloride, pyrazofurin, riboprine, rogletimide, safingol, safingol hydrochloride, semustine, simtrazene, sparfosate sodium, sparsomycin, spirogermanium hydrochloride, spiromustine, spiroplatin, streptonigrin, streptozocin, sulofenur, talisomycin, tecogalan sodium, tegafur, teloxantrone hydrochloride, temoporfin, teniposide, teroxirone, testolactone, thiamiprine, thioguanine, thiotepa, tiazofurin, tirapazamine, toremifene citrate, trestolone acetate, triciribine phosphate, trimetrexate, trimetrexate glucuronate, triptorelin, tubulozole hydrochloride, uracil mustard, uredepa, vapreotide, verteporfm, vinblastine sulfate, vincristine sulfate, vindesine, vindesine sulfate, vinepidine sulfate, vinglycinate sulfate, vinleurosine sulfate, vinorelbine tartrate, vinrosidine sulfate, vinzolidine sulfate, vorozole, zeniplatin, zinostatin, zorubicin hydrochloride. In some embodiments, a therapeutic agent is selected from: 20-epi-l, 25 dihydroxyvitamin D3, 5-ethynyluracil, abiraterone, aclarubicin, acylfulvene, adecypenol, adozelesin, aldesleukin, ALL-TK antagonists, altretamine, ambamustine, amidox, amifostine, aminolevulinic acid, amrubicin, amsacrine, anagrelide, anastrozole, andrographolide, angiogenesis inhibitors, antagonist D, antagonist G, antarelix, anti-dorsalizing morphogenetic protein- 1, antiandrogen, prostatic carcinoma, antiestrogen, antineoplaston, antisense oligonucleotides, aphidicolin glycinate, apoptosis gene modulators, apoptosis regulators, apurinic acid, ara-CDP-DL-PTBA, arginine deaminase, asulacrine, atamestane, atrimustine, axinastatin 1 , axinastatin 2, axinastatin 3, azasetron, azatoxin, azatyrosine, baccatin III derivatives, balanol, batimastat, BCR/ABL antagonists, benzochlorins, benzoylstaurosporine, beta lactam derivatives, beta-alethine, betaclamycin B, betulinic acid, bFGF inhibitor, bicalutamide, bisantrene, bisaziridinylspermine, bisnafide, bistratene A, bizelesin, breflate, bropirimine, budotitane, buthionine sulfoximine, calcipotriol, calphostin C, camptothecin derivatives, canarypox IL-2, capecitabine, carboxamide-amino-triazole, carboxyamidotriazole, Ca est M3, CARN 700, cartilage derived inhibitor, carzelesin, casein kinase inhibitors (ICOS), castanospermine, cecropin B, cetrorelix, chlorlns, chloroquinoxaline sulfonamide, cicaprost, ΰίβ-ροφΐιντίη, cladribine, clomifene analogues, clotrimazole, collismycin A, collismycin B, combretastatin A4,
combretastatin analogue, conagenin, crambescidin 816, crisnatol, cryptophycin 8, cryptophycin A WSGR Docket No. 39088-711.601
derivatives, curacin A, cyclopentanthraquinones, cycloplatam, cypemycin, cytarabine ocfosfate, cytolytic factor, cytostatin, dacliximab, decitabme, dehydrodidemnm B, deslorelin, dexamethasone, dexifosfamide, dexrazoxane, dexverapamil, diaziquone, didemnin B, didox, diethylnorspermine, dihydro-5-azacytidine, 9- dioxamycin, diphenyl spiromustine, docosanol, dolasetron, doxifluridine, droloxifene, dronabinol, duocarmycin SA, ebselen, ecomustine, edelfosine, edrecolomab, eflomithine, elemene, emitefur, epirubicin, epristeride, estramustine analogue, estrogen agonists, estrogen antagonists, etanidazole, etoposide phosphate, exemestane, fadrozole, fazarabine, fenretinide, filgrastim, finasteride, flavopiridol, flezelastine, fluasterone, fludarabine,
fluorodaunorunicin hydrochloride, forfenimex, formestane, fostriecin, fotemustine, gadolinium texaphyrin, gallium nitrate, galocitabine, ganirelix, gelatinase inhibitors, gemcitabine, glutathione inhibitors, hepsulfam, heregulin, hexamethylene bisacetamide, hypericin, ibandronic acid, idarubicin, idoxifene, idramantone, ilmofosine, ilomastat, imidazoacridones, imiquimod, immunostimulant peptides, insulin-such as for example growth factor- 1 receptor inhibitor, interferon agonists, interferons, interleukins, iobenguane, iododoxorubicin, ipomeanol, 4-, iroplact, irsogladine, isobengazole, isohomohalicondnn B, itasetron, jasplakinolide, kahalalide F, lamellarin- N triacetate, lanreotide, leinamycin, lenograstim, lentinan sulfate, leptolstatin, letrozole, leukemia inhibiting factor, leukocyte alpha interferon, leuprolide+estrogen+progesterone, leuprorelin, levamisole, liarozole, linear polyamine analogue, lipophilic disaccharide peptide, lipophilic platinum compounds, lissoclinamide 7, lobaplatin, lombricine, lometrexol, lonidamine, losoxantrone, lovastatin, loxoribine, lurtotecan, lutetium texaphyrin, lysofylline, lytic peptides, maitansine, mannostatin A, marimastat, masoprocol, maspin, matrilysin inhibitors, matrix metalloproteinase inhibitors, menogaril, merbarone, meterelin, methioninase, metoclopramide, MIF inhibitor, mifepristone, miltefosine, mirimostim, mismatched double stranded RNA, mitoguazone, mitolactol, mitomycin analogues, mitonafide, mitotoxin fibroblast growth factor-saporin, mitoxantrone, mofarotene, molgramostim, monoclonal antibody, human chorionic gonadotrophin, monophosphoryl lipid A+myobacterium cell wall sk, mopidamol, multiple drug resistance gene inhibitor, multiple tumor suppressor 1 -based therapy, mustard anticancer agent, mycaperoxide B, mycobacterial cell wall extract, myriaporone, N-acetyldinaline, N-substituted benzamides, nafarelin, nagrestip, naloxone+pentazocine, napavin, naphterpin, nartograstim, nedaplatin, nemorubicin, neridronic acid, neutral endopeptidase, nilutamide, nisamycin, nitric oxide modulators, nitroxide antioxidant, nitrullyn, 06-benzylguanine, octreotide, okicenone,
oligonucleotides, onapristone, ondansetron, ondansetron, oracin, oral cytokine inducer, ormaplatin, osaterone, oxaliplatin, oxaunomycin, palauamine, palmitoylrhizoxin, pamidronic acid, panaxytriol, panomifene, parabactin, pazelliptine, pegaspargase, peldesine, pentosan polysulfate sodium, WSGR Docket No. 39088-711.601
pentostatin, pentrozole, perfiubron, perfosfamide, perillyl alcohol, phenazinomycin, phenylacetate, phosphatase inhibitors, picibanil, pilocarpine hydrochloride, pirarubicin, piritrexim, placetin A, placetin B, plasminogen activator inhibitor, platinum complex, platinum compounds, platinum- triamine complex, porfimer sodium, porfiromycin, prednisone, propyl bis-acridone, prostaglandin J2, proteasome inhibitors, protein A-based immune modulator, protein kinase C inhibitor, protein kinase C inhibitors, microalgal, protein tyrosine phosphatase inhibitors, purine nucleoside phosphorylase inhibitors, purpurins, pyrazoloacridine, pyridoxylated hemoglobin polyoxyethylerie conjugate, raf antagonists, raltitrexed, ramosetron, ras farnesyl protein transferase inhibitors, ras inhibitors, ras-GAP inhibitor, retelliptine demethylated, rhenium Re 186 etidronate, rhizoxin, ribozymes, RII retinamide, rogletimide, rohitukine, romurtide, roquinimex, rubiginone Bl, ruboxyl, safmgol, saintopin, SarCNU, sarcophytol A, sargramostim, Sdi 1 mimetics, semustine, senescence derived inhibitor 1 , sense oligonucleotides, signal transduction inhibitors, signal transduction modulators, single chain antigen-binding protein, sizofiran, sobuzoxane, sodium borocaptate, sodium phenylacetate, solverol, somatomedin binding protein, sonermin, sparfosic acid, spicamycin D, spiromustine, splenopentin, spongistatin 1, squalamine, stem cell inhibitor, stem-cell division inhibitors, stipiamide, stromelysin inhibitors, sulfinosine, superactive vasoactive intestinal peptide antagonist, suradista, suramin, swainsonine, synthetic glycosaminoglycans, tallimustine, tamoxifen methiodide, tauromustine, tazarotene, tecogalan sodium, tegafur, tellurapyrylium, telomerase inhibitors, temoporfm, temozolomide, teniposide, tetrachlorodecaoxide, tetrazomine, thaliblastine, thiocoraline, thrombopoietin, thrombopoietin mimetic, thymalfasin, thymopoietin receptor agonist, thymotrinan, thyroid stimulating hormone, tin ethyl etiopurpurin, tirapazamine, titanocene bichloride, topsentin, toremifene, totipotent stem cell factor, translation inhibitors, tretinoin, triacetyluridine, triciribine, trimetrexate, triptorelin, tropisetron, turosteride, tyrosine kinase inhibitors, tyrphostins, UBC inhibitors, ubenimex, urogenital sinus-derived growth inhibitory factor, urokinase receptor antagonists, vapreotide, variolin B, vector system, erythrocyte gene therapy, velaresol, veramine, verdins, verteporfm, vinorelbine, vinxaltine, vitaxin, vorozole, zanoterone, zeniplatin, zilascorb, zinostatin stimalamer, mechloroethamine, cyclophosphamide, chlorambucil, busulfan, carmustine, lomusitne, decarbazine, methotrexate, cytarabine,
mercaptopurine, thioguanine, pentostatin, mechloroethamine, cyclophosphamide, chlorambucil, meiphalan, ethylenimine, methylmelamine, hexamethlymelamine, thiotepa, busulfan, carmustine, lomusitne, semustine, streptozocin, decarbazine, fluorouracil, floxouridine, cytarabine,
mercaptopurine, thioguanine, pentostatin, erbulozole (also known as R-55104), Dolastatin 10 (also known as DLS-10 and NSC-376128), Mivobulin isethionate (also known as CI-980), Vincristine, NSC-639829, Discodermolide (also known as NVP-XX-A-296), ABT-751 (Abbott, also known as WSGR Docket No. 39088-711.601
E-7010), Altorhyrtins (such as Altorhyrtin A and Altorhyrtin C), Spongistatins (such as
Spongistatin 1, Spongistatin 2, Spongistatin 3, Spongistatin 4, Spongistatin 5, Spongistatin 6, Spongistatin 7, Spongistatin 8, and Spongistatin 9), Cemadotin hydrochloride (also known as LU- 103793 and NSC-D-669356), Epothilones (such as Epothilone A, Epothilone B, Epothilone C (also known as desoxyepothilone A or dEpoA), Epothilone D (also referred to as KOS-862, dEpoB, and desoxyepothilone B ), Epothilone E, Epothilone F, Epothilone B N-oxide, Epothilone A N-oxide, 16-aza-epothilone B, 21-aminoepothilone B (also known as BMS-310705), 21 -hydroxyepothilone D (also known as Desoxyepothilone F and dEpoF), 26-fluoroepothilone), Auristatin PE (also known as NSC-654663), Soblidotin (also known as TZT-1027), LS-4559-P (Pharmacia, also known as LS-4577), LS-4578 (Pharmacia, also known as LS-477-P), LS-4477 (Pharmacia), LS- 4559 (Pharmacia), RPR-112378 (Aventis), Vincristine sulfate, DZ-3358 (Daiichi), FR-182877 (Fujisawa, also known as WS-9885B), GS-164 (Takeda), GS-198 (Takeda), KAR-2 (Hungarian Academy of Sciences), BSF-2236 1 (BASF, also known as ILX-651 and LU-2236 1 ), SAH- 49960 (Lilly/Novartis), SDZ-268970 (Lilly/Novartis), AM-97 (Armad/Kyowa Hakko), AM- 132 (Armad), AM-138 (Armad/Kyowa Hakko), IDN-5005 (Indena), Cryptophycin 52 (also known as LY-355703), AC-7739 (Ajinomoto, also known as AVE-8063A and CS-39.HCI), AC-7700 (Ajinomoto, also known as AVE-8062, AVE-8062A, CS-39-L-Ser.HCI, and RPR-258062A), Vitilevuamide, Tubulysin A, Canadensol, Centaureidin (also known as NSC-106969), T-138067 (Tularik, also known as T-67, TL-138067 and TI-138067), COBRA-1 (Parker Hughes Institute, also known as DDE-261 and WHI-261), H10 (Kansas State University), H16 (Kansas State University), Oncocidin Al (also known as BTO-956 and DIME), DDE-313 (Parker Hughes Institute), Fijianolide B, Laulimalide, SPA-2 (Parker Hughes Institute), SPA-1 (Parker Hughes Institute, also known as SPIKET-P), 3-IAABU (Cytoskeleton/Mt. Sinai School of Medicine, also known as MF-569), Narcosine (also known as NSC-5366), Nascapine, D-24851 (Asta Medica), A- 105972 (Abbott), Hemiasterlin, 3-BAABU (Cytoskeleton/Mt. Sinai School of Medicine, also known as MF-191), TMPN (Arizona State University), Vanadocene acetylacetonate, T-138026 (Tularik), Monsatrol, lnanocine (also known as NSC-698666), 3-lAABE (Cytoskeleton/Mt. Sinai School of Medicine), A-204197 (Abbott), T-607 (Tuiarik, also known as T-900607), RPR- 115781 (Aventis), Eleutherobins (such as Desmethyleleutherobin, Desaetyleleutherobin, lsoeleutherobin A, and Z-Eleutherobin), Caribaeoside, Caribaeolin, Halichondrin B, D-64131 (Asta Medica), D-68144 (Asta Medica), Diazonamide A, A-293620 (Abbott), NPI-2350 (Nereus), Taccalonolide A, TUB- 245 (Aventis), A-259754 (Abbott), Diozostatin, (-)-Phenylahistin (also known as NSCL-96F037), D-68838 (Asta Medica), D-68836 (Asta Medica), Myoseverin B, D-43411 (Zentaris, also known as D-81862), A-289099 (Abbott), A-318315 (Abbott), HTI-286 (also known as SPA-110, WSGR Docket No. 39088-711.601
trifluoroacetate salt) (Wyeth), D-82317 (Zentaris), D-82318 (Zentaris), SC-12983 (NCI),
esverastatin phosphate sodium, BPR-OY-007 (National Health Research Institutes), and SSR- 25041 1 (Sanofi).
[00237] In some embodiments, a therapeutic agent is an anti-inflammatory agent. In some embodiments, a therapeutic agent is an anti-TNF agent, an IL-1 receptor antagonist, an IL-2 receptor antagonist, a cytotoxic agent, an immunomodulatory agent, an antibiotic, a T-cell co- stimulatory blocker, a B cell depleting agent, an immunosuppressive agent, an alkylating agent, an anti-metabolite, a plant alkaloid, a terpenoids, a topoisomerase inhibitor, an antitumour antibiotic, an antibody, a hormonal therapy, an anti-diabetes agent, a leukotriene inhibitor, or combinations thereof. In some embodiments, a therapeutic agent is selected from: alefacept, efalizumab, methotrexate, acitretin, isotretinoin, hydroxyurea, mycophenolate mofetil, sulfasalazine, 6- Thioguanine, Dovonex, Taclonex, betamethasone, tazarotene, hydroxychloroquine, etanercept, adalimumab, infliximab, abatacept, rituximab, tratuzumab, Anti-CD45 monoclonal antibody AHN- 12 (NCI), Iodine-131 Anti-Bl Antibody (Corixa Corp.), anti-CD66 monoclonal antibody BW 250/183 (NCI, Southampton General Hospital), anti-CD45 monoclonal antibody (NCI, Baylor College of Medicine), antibody anti-anb3 integrin (NCI), BIW-8962 (BioWa Inc.), Antibody BC8 (NCI), antibody muJ591 (NCI), indium In 11 1 monoclonal antibody MN-14 (NCI), yttrium Y 90 monoclonal antibody MN-14 (NCI), F105 Monoclonal Antibody (NIAID), Monoclonal Antibody RAVI 2 (Raven Biotechnologies), CAT- 192 (Human Anti-TGF-Betal Monoclonal Antibody, Genzyme), antibody 3F8 (NCI), 177Lu-J591 (Weill Medical College of Cornell University), TB- 403 (Biolnvent International AB), anakinra, azathioprine, cyclophosphamide, cyclosporine A, leflunomide, d-penicillamine, amitriptyline, or nortriptyline, chlorambucil, nitrogen mustard, prasterone, LJP 394 (abetimus sodium), LJP 1082 (La Jolla Pharmaceutical), eculizumab, belibumab, rhuCD40L (NIAID), epratuzumab, sirolimus, tacrolimus, pimecrolimus, thalidomide, antithymocyte globulin-equine (Atgam, Pharmacia Upjohn), antithymocyte globulin-rabbit (Thymoglobulin, Genzyme), Muromonab-CD3 (FDA Office of Orphan Products Development), basiliximab, daclizumab, riluzole, cladribine, natalizumab, interferon beta-lb, interferon beta- la, tizanidine, baclofen, mesalazine, asacol, pentasa, mesalamine, balsalazide, olsalazine, 6- mercaptopurine, AIN4 7 (Anti IL-17 Monoclonal Antibody, Novartis), theophylline, D2E7 (a human anti-TNF mAb from Knoll Pharmaceuticals), Mepolizumab (Anti-IL-5 antibody, SB 240563), Canakinumab (Anti-IL- 1 Beta Antibody, NIAMS), Anti-IL-2 Receptor Antibody (Daclizumab, NHLBI), CNTO 328 (Anti IL-6 Monoclonal Antibody, Centocor), ACZ885 (fully human anti-interleukin-lbeta monoclonal antibody, Novartis), CNTO 1275 (Fully Human Anti-IL- 12 Monoclonal Antibody, Centocor), (3S)-N-hydroxy-4-({4-[(4-hydroxy-2- WSGR Docket No. 39088-711.601
butynyl)oxy]phenyl}sulfonyl)-2,2-dimet- hyl-3-thiomorpholine carboxamide (apratastat), golimumab (CNTO 148), Onercept, BG9924 (Biogen Idee), Certolizumab Pegol (CDP870, UCB Pharma), AZD9056 (AstraZeneca), AZD5069 (AstraZeneca), AZD9668 (AstraZeneca), AZD7928 (AstraZeneca), AZD2914 (AstraZeneca), AZD6067 (AstraZeneca), AZD3342 (AstraZeneca), AZD8309 (AstraZeneca), ), [(lR)-3-methyl-l-({(2S)-3-phenyl-2-[(pyrazin-2- ylcarbonyl)amino]propanoyl}amino)butyl]boronic acid (Bortezomib), AMG-714, (Anti-IL 15 Human Monoclonal Antibody, Amgen), ABT-874 (Anti IL-12 monoclonal antibody, Abbott Labs), MRA(Tocilizumab, an Anti IL-6 Receptor Monoclonal Antibody, Chugai Pharmaceutical), CAT- 354 (a human anti-interleukin-13 monoclonal antibody, Cambridge Antibody Technology, Medlmmune), aspirin, salicylic acid, gentisic acid, choline magnesium salicylate, choline salicylate, choline magnesium salicylate, choline salicylate, magnesium salicylate, sodium salicylate, diflunisal, carprofen, fenoprofen, fenoprofen calcium, flurobiprofen, ibuprofen, ketoprofen, nabutone, ketolorac, ketorolac tromethamine, naproxen, oxaprozin, diclofenac, etodolac, indomethacin, sulindac, tolmetin, meclofenamate, meclofenamate sodium, mefenamic acid, piroxicam, meloxicam, celecoxib, rofecoxib, valdecoxib, parecoxib, etoricoxib, lumiracoxib, CS-502 (Sankyo), JTE-522 (Japan Tobacco Inc.), L-745,337 (Almirall), NS398 (Sigma), betamethasone (Celestone), prednisone (Deltasone), alclometasone, aldosterone, amcinonide, beclometasone, betamethasone, budesonide, ciclesonide, clobetasol, clobetasone, clocortolone, cloprednol, cortisone, cortivazol, deflazacort, deoxycorticosterone, desonide, desoximetasone, desoxycortone, dexamethasone, difiorasone, diflucortolone, difiuprednate, fluclorolone, fludrocortisone, fludroxycortide, flumetasone, flunisolide, fluocinolone acetonide, fluocinonide, fluocortin, fiuocortolone, fiuorometholone, fiuperolone, fluprednidene, fluticasone, formocortal, formoterol, halcinonide, halometasone, hydrocortisone, hydrocortisone aceponate, hydrocortisone buteprate, hydrocortisone butyrate, loteprednol, medrysone, meprednisone, methylprednisolone, methylprednisolone aceponate, mometasone furcate, paramethasone, prednicarbate, prednisone, rimexolone, tixocortol, triamcinolone, ulobetasol, Pioglitazone, Rosiglitazone, Glimepiride, Glyburide, Chlorpropamide, Glipizide, Tolbutamide, Tolazamide, Glucophage, Metformin, (glyburide + metformin), Rosiglitazone + metformin, (Rosiglitazone+glimepiride), Exenatide, Insulin, Sitagliptin, (glipizide and metformin), Repaglinide, Acarbose, Nateglinide, Orlistat, cisplatin; carboplatin; oxaliplatin; mechlorethamine; cyclophosphamide; chlorambucil; vincristine; vinblastine; vinorelbine; vindesine; mercaptopurine; fludarabine; pentostatin; cladribine; 5- fluorouracil ( FU); floxuridine (FUDR); cytosine arabinoside; trimethoprim; pyrimethamine; pemetrexed; paclitaxel; docetaxel; etoposide; teniposide; irinotecan; topotecan; amsacrine;
etoposide; etoposide phosphate; teniposide; dactinomycin; doxorubicin; daunorubicin; valrubicine; WSGR Docket No. 39088-711.601
idarubicine; epirubicin; bleomycin; plicamycin; mitomycin; finasteride; goserelin;
aminoglutethimide; anastrozole; letrozole; vorozole; exemestane; 4-androstene-3,6,17-trione ("6- OXO"; l,4,6-androstatrien-3,17-dione (ATD); formestane; testolactone; fadrozole; A-81834 (3-(3- (l,l-dimethylethylthio-5-(quinoline-2- ylmethoxy)- 1 -(4-chloromethylphenyl)indole-2-yl)-2,2- dimethylpropionaldehyde oxime-O-2-acetic acid; AME103 (Amira); AME803 (Amira); atreleuton; BAY-x-1005 ((R)-(+)-alpha-cyclopentyl-4-(2-quinolinylmethoxy)-Benzeneacetic acid); CJ-13610 (4-(3-(4-(2-Methyl-imidazol-l-yl)-phenylsulfanyl)- phenyl)-tetrahydro-pyran-4-carboxylic acid amide); DG-031 (DeCode); DG-051 (DeCode); MK886 (l-[(4-chlorophenyl)methyl]3-[(l,l- dimethylethyl)thio]-a,a-dimethyl-5-(l-methylethyl)-lH-indole-2-propanoic acid, sodium salt); MK591 (3-(l-4[(4-chlorophenyl)methyl]-3 (t-butylthio)-5-((2-quinoly)methoxy) H-indole-2 dimehtylpropanoic acid); RP64966 ([4-[5-(3-Phenyl-propyl)thiophen-2- yl]butoxy] acetic acid); SA6541 ((R)-S-[[4- (dimethylamino)phenyl]methyl] -N-(3-mercapto-2methyl- 1 -oxopropyl-L- cycteine); SC-56938 (ethyl- l-[2-[4-(phenylmethyl)phenoxy] ethyl] -4-piperidine- carboxylate); VIA-2291 (Via Pharmaceuticals); WY-47,288 (2-[(l-naphthalenyloxy)methyl]quinoline); zileuton; ZD-2138 (6-((3-fluoro-5- (tetrahydro-4-methoxy-2H-pyran-4yl)phenoxy)methyl)-l-methyl-2(lH)- quinlolinone); doxycycline; or combinations thereof.
[00238] In some embodiments, an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, is administered with one or more additional therapeutic agents. In some embodiments, the additional therapeutic agent is selected from among the therapeutic agents listed herein. In some embodiments, the additional therapeutic agent is administered prior to, following, or simultaneously (i.e., concurrently) with an SDM of any of Formulas -VI or a carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, provided herein.
[00239] Imaging Uses
[00240] The SDMs of Formulas I-VI and carrier-conjugated SDMs comprising a targeting ligand, e.g. an antibody, allow the targeted delivery of imaging agents to specific cells and/or tissues (e.g., cancerous tissues). The SDMs comprise a basic peptide sequence (B) which is designed to be transported across a cellular membrane or retained by tissue, an acidic peptide sequence (A) which inhibits uptake and retention of peptide B into cells, a linker X which is cleavable under specific conditions, imaging moieties bound to peptides A and B, or X and a macromolecular carrier. In some embodiments, cleavage of the linker X linker frees peptide B from peptide A and allows the transport of peptide B (and any imaging moieties attached thereto) across a cellular membrane or retention of B to tissue. In some embodiments, the SDMs enable targeted delivery of one or more WSGR Docket No. 39088-711.601
imaging agents to a cell or tissue. In some embodiments, targeted delivery of an imaging agent to a cell or tissue enables a medical professional to visualize/image a specific tissue.
[00241] In some embodiments, targeted delivery of an imaging agent to a cell or tissue enables a medical professional to visualize/image a specific tissue (e.g., cancerous tissue). In some embodiments, targeted delivery of an imaging agent to a cell or tissue enables a medical professional to remove (or, surgically excise) the tissue of interest (e.g., cancerous tissue). In some embodiments, targeted delivery of an imaging agent to a cell or tissue enables a medical professional to remove (or, surgically excise) the tissue of interest (e.g., cancerous tissue) with a decrease in surgical margins. In some embodiments, targeted delivery of an imaging agent to a cell or tissue enables a medical professional to remove (or, surgically excise) a tumor/cancerous tissue and decreases the chance that some of the tumor/cancerous tissue will not be removed. In some embodiments, targeted delivery of an imaging agent to a cell or tissue enables a medical professional to maximally debulk a tumor/cancerous tissue. In some embodiments, targeted delivery of an imaging agent to cancerous breast tissue decreases the chances of an unnecessary operations and re-operations.
[00242] In some embodiments, targeted delivery of an imaging agent to a cell or tissue enables a medical professional to more accurately sample (e.g., biopsy (e.g., excision biopsy, incision, biopsy, aspiration biopsy, or needle biopsy)) tissue of interest (e.g., cancerous tissue). In some embodiments, targeted delivery of an imaging agent to a cell or tissue enables a medical professional to visualize/image a specific tissue (e.g., cancerous tissue) within an excised tissue containing healthy tissue. Enabling identification of target tissue (e.g., cancerous tissue) can guide the pathologist on where to section of pathological evaluation and decreases the chances of a pathologist missing unhealthy tissue (e.g., cancerous tissue) and sampling healthy tissue which may produce a false negative. In some embodiments, tissue (e.g., cancerous tissue) removed following use of a compound of Formula I is used to prepare a pathology section or slide. In some embodiments, cancerous tissue removed following use of a compound of Formula I is used to prepare a pathology section or slide which is used to diagnose a tissue as malignant or benign.
[00243] In some embodiments, targeted delivery of an imaging agent to cancerous breast tissue enables a medical professional to accurately stage cancer enabling medical treatment decisions. In some embodiments, targeted delivery of an imaging agent to cancerous tissue enables a medical professional to observe the size of a tumor (cancerous tissue) or the spread (e.g., metastatic lesions) of cancerous tissue. In some embodiments, targeted delivery of an imaging agent to a cell or tissue enables a medical professional to design an efficacious treatment regimen. WSGR Docket No. 39088-711.601
[00244] In some embodiments, a selective delivery molecule according to Formula I comprising an imaging agent is employed in guided surgery. In some embodiments, the selective delivery molecule preferentially localized to cancerous, or other pathological tissues with up-regulated protease activity (e.g. tissues undergoing inflammatory response). In some embodiments, a selective delivery molecule according to Formula I comprising an imaging agent is employed in a guided surgery to remove colorectal cancer. In some embodiments, guided surgery employing the selective delivery molecule allows a surgeon to excise as little healthy (i.e., non-cancerous) tissue as possible. In some embodiments, guided surgery employing the selective delivery molecule allows a surgeon to visualize and excise more cancerous tissue than the surgeon would have been able to excise without the presence of the selective delivery molecule. In some embodiments, the surgery is fluorescence-guided surgery.
Imaging Agents
[00245] In some embodiments, an imaging agent is a dye. In some embodiments, an imaging agent is a fluorescent moiety. In some embodiments, a fluorescent moiety is selected from: a fluorescent protein, a fluorescent peptide, a fluorescent dye, a fluorescent material or a combination thereof.
[00246] All fluorescent moieties are encompassed within the term "fluorescent moiety." Specific examples of fluorescent moieties given herein are illustrative and are not meant to limit the fluorescent moieties for use with the targeting molecules disclosed herein.
[00247] Examples of fluorescent dyes include, but are not limited to, xanthenes (e.g., rhodamines, rhodols and fluoresceins, and their derivatives); bimanes; coumarins and their derivatives (e.g., umbelliferone and aminomethyl coumarins); aromatic amines (e.g., dansyl; squarate dyes);
benzofurans; fluorescent cyanines; indocarbocyanines; carbazoles; dicyanomethylene pyranes; polymethine; oxabenzanthrane; xanthene; pyrylium; carbostyl; perylene; acridone; quinacridone; rubrene; anthracene; coronene; phenanthrecene; pyrene; butadiene; stilbene; porphyrin;
pthalocyanine; lanthanide metal chelate complexes; rare-earth metal chelate complexes; and derivatives of such dyes.
[00248] Examples of fluorescein dyes include, but are not limited to, 5-carboxyfluorescein, fluorescein-5-isothiocyanate, fluorescein-6-isothiocyanate and 6-carboxyfluorescein.
[00249] Examples of rhodamine dyes include, but are not limited to, tetramethylrhodamine-6- isothiocyanate, 5-carboxytetramethylrhodamine, 5-carboxy rhodol derivatives, tetramethyl and tetraethyl rhodamine, diphenyldimethyl and diphenyldiethyl rhodamine, dinaphthyl rhodamine, rhodamine 101 sulfonyl chloride (sold under the tradename of TEXAS RED®).
[00250] Examples of cyanine dyes include, but are not limited to, Cy3, Cy3B, Cy3.5, Cy5, Cy5.5,
Cy7, IRDYE680, Alexa Fluor 750, IRDye800CW, ICG. WSGR Docket No. 39088-711.601
[00251] Examples of fluorescent peptides include GFP (Green Fluorescent Protein) or derivatives of GFP (e.g., EBFP, EBFP2, Azurite, mKalamal, ECFP, Cerulean, CyPet, YFP, Citrine, Venus, YPet).
[00252] Fluorescent labels are detected by any suitable method. For example, a fluorescent label may be detected by exciting the fluorochrome with the appropriate wavelength of light and detecting the resulting fluorescence, e.g., by microscopy, visual inspection, via photographic film, by the use of electronic detectors such as charge coupled devices (CCDs), photomultipliers, etc.
[00253] In some embodiments, the imaging agent is labeled with a positron-emitting isotope (e.g.,18F) for positron emission tomography (PET), gamma-ray isotope (e.g., 99mTc) for single photon emission computed tomography (SPECT), or a paramagnetic molecule or nanoparticle
3+
(e.g.,Gd chelate or coated magnetite nanoparticle) for magnetic resonance imaging (MRI).
[00254] In some embodiments, the imaging agent is labeled with: a gadolinium chelate, an iron oxide particle, a super paramagnetic iron oxide particle, an ultra small paramagnetic particle, a manganese chelate or gallium containing agent.
[00255] Examples of gadolinium chelates include, but are not limited to diethylene triamine pentaacetic acid (DTPA), l ,4,7,10-tetraazacyclododecane-l ,4,7,10-tetraacetic acid (DOTA), and l,4,7-triazacyclononane-N,N',N"-triacetic acid (NOT A).
[00256] In some embodiments, the imaging agent is a near-infrared fluorophore for near-infra red (near-IR) imaging, a luciferase (firefly, bacterial, or coelenterate) or other luminescent molecule for bioluminescence imaging, or a perfluorocarbon- filled vesicle for ultrasound.
[00257] In some embodiments, the imaging agent is a nuclear probe. In some embodiments, the imaging agent is a SPECT or PET radionuclide probe. In some embodiments, the radionuclide probe is selected from: a technetium chelate, a copper chelate, a radioactive fluorine, a radioactive iodine, a indiuim chelate.
[00258] Examples of Tc chelates include, but are not limited to HYNIC, DTPA, and DOTA.
[00259] In some embodiments, the imaging agent contains a radioactive moiety, for example a radioactive isotope such as 211At, 1311, 1251, 90Y, 186Re, 188Re, 153Sm, 212Bi, 32P, 64Cu radioactive isotopes of Lu, and others.
Starting Materials
[00260] Disclosed herein, in certain embodiments, are molecules of Formula VII, having the structure:
Ai-Xi-Bi;
Formula VII
wherein, WSGR Docket No. 39088-711.601
Xi is a cleavable linker;
Ai is a peptide with a sequence comprising 5 to 9 acidic amino acids and having a first reactive amino acid moiety CA;
Bi is a peptide with a sequence comprising 7 to 9 basic amino acids and having a second reactive amino acid moiety CB; and
Ai-Xi-Bi has a third reactive amino acid moiety CM of Ai or Xi; and
wherein CA is capable of reacting with a first cargo moiety comprising DA, CB is capable of reacting with a second cargo moiety comprising DB, and CM is capable of reacting with a macromolecular carrier comprising M to form a molecule of Formula I. In some embodiments, the CA, CB, and CM have functional groups that are orthogonally reactive. In some embodiments, CA, CB, and CM are each independently selected from a naturally-occurring amino acid or a non-naturally-occurring amino acid. In some embodiments, CA, CB, and CM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a B-amino acid, or a y-amino acid. In some embodiments, CA, CB, and CM are each independently selected from any amino acid having a free thiol group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CA, CB, and CM are each independently selected from D-cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine. In some embodiments, CB is any amino acid having a free thiol group. In some embodiments, cB is D-cysteine. In some embodiments, CA is any amino acid having a N-terminal amine group. In some embodiments, CA is D-glutamate. In some embodiments, CA is lysine. In some embodiments, CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group. In some embodiments, CM is para-4-acetyl L-phenylalanine.
[00261] As used herein, "orthogonally reactive" means a plurality of groups can be attached to a molecule via a sequence of reactions that do not cross react enabling specific attachment of each group in the presence of the others. In some embodiments, the three groups (DA, Db, and DM) are able to be attached to Ai-Xi-Bi via CA, CB, and CM using a sequence of 3 independent reactions that do not cross react so that each group is attached to only one site of Ai-Xi-Bi.
[00262] Disclosed herein, in certain embodiments, is a molecule having the amino acid sequence:
(D-Glu)5_F(4-Ac)-o-Pro-Leu-Gly-Cys(Me)-Ala-Gly-(D-Arg)8-(D-Cys) wherein o represent 5-(amino-3-oxapentanoyl); F(4_Ac) represent ?ara-acetyl-(L)-phenylalanine; and C(Me) represents S-methyl-(L)-cysteine.
[00263] In some embodiments, the molecule further comprises a polyethylene glycol (PEG) polymer. In some embodiments, the PEG polymer is covalently linked to the molecule at the F(4- WSGR Docket No. 39088-711.601
Ac) subunit. In some embodiments, the molecule comprises groups that can be orthogonally reacted. In some embodiments, the groups that can be orthogonally reacted are chosen from: an amine, thiol and an acetyl phenylalanine. In some embodiments, the molecule comprises an amine, a thiol, and an acetyl phenylalanine.
[00264] In some embodiments, the PEG polymer has an average molecular weight of 500 daltons. In some embodiments, the PEG polymer has an average molecular weight of 2,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 3,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 4,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 5,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 10,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 12,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 15,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 20,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 30,000 daltons. In some embodiments, the PEG polymer has an average molecular weight of 40,000 daltons.
[00265] Disclosed herein, in certain embodiments, is the use of the molecule in the synthesis of a molecule according to Formulas I- VI.
[00266] Disclosed herein, in certain embodiments, is a molecule having the amino acid sequence:
(D-Glu)5-o-Pro-Leu-Glys-Cys(me)-Ala-Gly-(D-Arg)8-(D-Cys)-[PEG(3K)] wherein all glutamates and arginines are D-amino acids; o represents 5-(amino-3-oxapentanoyl); C(me) represents S-methyl-(L)-cysteine; and PEG(3K) represents a-amino-co-amide poly(ethylene glycol) with an average three thousand Dalton molecular weight. In some embodiments, the molecule further comprises a fluorescent moiety. Disclosed herein, in certain embodiments, is the use of the molecule in the synthesis of a molecule according to Formulas I- VI.
Figure imgf000131_0001
Figure imgf000132_0001
Figure imgf000133_0001
-131-
Figure imgf000134_0001
Figure imgf000135_0001
-133-
Figure imgf000136_0001
Figure imgf000137_0001
EXAMPLES
[00267] These examples are provided for illustrative purposes only and not to limit the scope of the claims provided herein.
Materials and Methods
[00268] All reaction solvents were freshly opened Aldrich "Sure-Seal" quality. All the reagents were reagent-grade and used without further purification unless otherwise indicated. HPLC-grade acetonitrile was purchased from Fisher Scientific (Phillipsburg, PA). Water used in HPLC was collected through Milli-Q water purification system (Millipore, Bedford, MA). PBS-EDTA buffer was purchased from Teknova (Hollister, CA). a-Mercaptoethyl-co-methoxy, poly-oxyethylene (average molecular weight around 2,000, 5,000, 20,000 and 40,000 daltons) [mPEG(2 )-SH, mPEG(5 )-SH, mPEG(20 )-SH, mPEG(40K)-SH] and a-aminoxyl-co-methoxy, polyoxyethylene (average molecular weight around 2,000, 5,000, 20,000 and 40,000) [mPEG(2K)-ONH2, mPEG(5 )-ONH2, mPEG(20K)-ONH2, mPEG(40K)-ONH2] were purchased from NOF America Corporation (Irvine, CA). Compound 1 was supplied by GL Biochem Ltd. (Shanghai, China). Doxorubicin was purchased from NuB locks LLC (Oceanside, CA). Lyophilized peptide P1-P18 was supplied by Polypeptide Group (San Diego, CA). 3-Maleimidopropionic acid pentafluorophenyl ester 7 was purchased from Molecular Biosciences (Boulder, CO). Compound 17 was purchased from MedChem Express (Princeton, CO).
[00269] LC-MS analysis was carried out on an Agilent 1200 SL series in combination with AB SCIEX API 3200, equipped with CTC PAL autosampler operating at 4°C, a vacuum degasser, binary pump, UV-VIS detector, associated Analyst 1.5 analytical software and a Phenomenex column (Kinetex 2.6μ C18 100A, 100 x 2.1 mm) or a Waters 2695 separation module equipped with a Waters 2487 dual λ absorbance detector in combination with Finnigan LCQ Deca XP mass spectrometer. The equipment is associated with Xcalibur analytical software and Peeke Scientific columns (Titan 200 5μιη, C18-MC, 50/100 x 2.1 mm).
[00270] Preparation HPLC were carried out on an Agilent system (Agilent 1200 series) and a Thermo Scientific column (Hypersil Gold C I 8, 5μ, 250 x 10 mm), or a Waters Delta Prep preparative HPLC System and a Varian column (F75L, C I 8, 15μ, 1200g), or a Waters PrepLC System equipped with a Waters 2487 dual λ absorbance detector, Fraction Collector III, Masslynx software and a Thermo Scientific column (Hypersil Gold C18, 5μ, 250 x 10 mm) or a Phenomenex column (luna, C 18(2), 5μ, 100A AX 150 x 30 mm). The mobile phase consisted of a water (0.05% TFA)(solvent A)/acetonitrile (0.05% TFA)(solvent B) gradient unless otherwise specified. W SijK DocKeS NO. ifi) ~ ( i i .'At i
Centrifagation was earned out at 4 °C on an Eppendorf centrifuge 5417 or a Beckman Mserofuget.- 18. Lyophi fixation was carried out on a Labconco FreeZone 4.5,
Exam le ..i ;. Svn thes s of Intermediate 5
Figure imgf000139_0001
Synthesis vf Intermediate 3
1602711 o a solution of peptide 1 (LOI g, 1 .2 mmol) and 2 (2.0 g, 6. mmol) in CH2<¾ \ ml.) at room temperature was added DIEA (0.65 nvLs 3.7 finmoi.). The mixture was stirred at room temperature for ! day. After the solvent was removed, the residue was dissolved i« EtOAc (' 00 mL). The organic layer was washed with sodium acetate buffer (pH 5, 100 mL, 1 M), water (100 mL) and brine (50 mL), dried, and evaporated. The residue was purified by flash chromatography on silica gel, eluting with 1 : 1 ethyl acetate/hexane, to afford 3 (920 mg, 76%) as a light yellow oil. 1H NMR (500 MHz, CDC13): δ 8.46 (br. s, 1H), 8.25 (d, J= 9.0 Hz, 2H), 8.10 (d, J= 9.0 Hz, 1H), 7.60 (d, J= 8.0 Hz, 2H), 7.43-7.34 (m, 7H), 7.30 (d, J= 8.0 Hz, 2H), 7.23 (t, J= 8.0 Hz, 4H), 7.17- 7.11 (m, 7H), 7.04 (d, J= 8.0 Hz, 2H), 6.82 (d, J = 9.0 Hz, 2H), 6.60 (s, 2H), 5.25 (s, 2H), 4.65 (q, J= 7.0 Hz, 1H), 4.43 (q, J= 7.0 Hz, 1H), 3.43 (t, J= 7.0 Hz, 2H), 3.09 (dd, J= 14.0; 7.0 Hz, 1H), 3.03 (dd, J= 14.0; 7.0 Hz, 1H), 2.28 (s, 3H), 2.13 (t, J = 7.5 Hz, 2H), 2.08 (t, J = 7.0 Hz, 2H), 1.89- 1.92 (m, 1H), 1.46-1.65 (m, 6H), 1.27-1.32 (m, 4H), 1.18-1.27 (m, 2H); 13C NMR (125 MHz, CDC13): δ 173.9, 171.7, 171.0, 169.6, 163.8, 155.8, 152.7, 146.6, 145.6, 143.4, 138.7, 136.0, 135.9, 134.3, 130.4, 129.9, 129.3, 128.8, 128.7, 128.7, 127.9, 127.6, 126.5, 126.4, 125.5, 122.0, 120.4, 120.3, 116.2, 70.9, 70.8, 55.1 , 54.5, 43.5, 38.1 , 37.7, 36.3, 31.5, 30.7, 28.3, 26.3, 25.0, 23.7, 21.1, 19.3; MS (ESI): m/e 1013 [M + H]+, 1036 [M + Na]+.
Synthesis of Intermediate 5
[00272] To a solution of intermediate 3 (137 mg, 0.14 mmol) and doxorubicin 4 (80 mg, 0.14 mmol) in N-methyl-2-pyrrolidone (ΝΜΡ) (5 mL) at room temperature was added DIEA (25 μί, 0.14 mmol). The mixture was stirred at room temperature in the dark for 72 h. To the reaction mixture was added EtOAc (100 mL). The organic layer was washed with water (100 mL x 2) and brine (50 mL), dried, and evaporated. The residue was purified by flash chromatography on silica gel, eluting with 3 : 1 ethyl acetate/hexane, to afford 5 (1 18 mg, 76%) as an orange powder. 1H NMR (500 MHz, DMSO-dg): δ 14.0 (s, 1H), 13.3 (s, 1H), 8.13 (d, J= 8.0 Hz, 1H), 7.97 (d, J = 8.0 Hz, 1H), 7.93-7.89 (m, 2H), 7.66-7.64 (m, 1H), 7.53 (d, J= 8.0 Hz, 2H), 7.47-7.43 (m, 6H), 7.30-7.15 (m, 16H), 6.99 (s, 2H), 6.81 (d, J = 8.0 Hz, 1H), 5.45 (br. s, 1H), 5.22 (d, J = 3.0 Hz, lH), 4.95 (t, J = 4.0 Hz, 1H), 4.90 (d, J= 12.5 Hz, 1H), 4.88 (d, J = 12.5 Hz, 1H), 4.57 (s, 2H), 4.52-4.46 (m, 1H), 4.32-3.30 (m, 1H), 4.16-4.14 (m, 1H), 3.98 (s, 4H), 3.44 (s, 1H), 2.39 (t, J = 7.0 Hz, 3H), 2.99-2.94 (m, 3H), 2.73-2.70 (m, 3H), 2.31 (s, 3H), 2.22-2.13 (m, 2H), 2.00-1.93 (m, 2H), 1.87-1.82 (m, 1H), 1.75-1.45 (m, 5H), 1.42-1.28 (m, 4H), 1.25-1.15 (m, 3H), 1.12 (d, J= 6.5 Hz, 3H), 1.10-0.98 (m, 2H); 13C NMR (125 MHz, DMSO-d6): δ 213.5, 186.6, 186.5, 172.1 , 171.4, 170.9, 170.1, 160.8, 156.0, 155.2, 154.5, 138.2, 137.8, 136.2, 135.5, 134.7, 134.4, 134.1 , 132.0, 129.1 , 129.0, 128.6, 128.5, 128.4, 127.9, 126.1 , 120.1, 1 19.7, 1 19.1, 1 19.0, 110.8, 110.7, 100.1 , 75.0, 69.8, 68.0, 66.6, 64.8, 63.6, 56.6, 53.8, 53.2, 47.1 , 37.2, 36.9, 36.7, 34.9, 32.1 , 31.4, 29.8, 27.6, 25.7, 25.5, 24.6, 22.6, 20.4, 16.9; MS (ESI): m/e 1417 [M + H]+, 1440 [M + Na . te 2; Synthesis of SPM-lOl and SDM-14S
Figure imgf000141_0001
30 mitt !
»< * ■<« v« ..... 5Li . ¾ ,Ο
a; i>» <M». . ¾N ^ ^ W N)?I TIH ¾M NH ¾ <?M ?
Synthesis of Intermediate 6
IM273J To a solution of intermediate S (3,8 rag, 2.7 μ¾ο!) and peptide P-2 (8.0 $¾g, 2,2 umo!) in DMF (0,7 mL) at room temperature in the dark was added Λ'-methyImorph iine ( MM) (5 μΙ.., 46 moi) with stitring. The reaction was followed by LC-MS md completed in I h. The mixture was directly used m the nexs step without {urtber pui-ification. MS (ESI): /»·'<? 49 M + 3H]i ;. Synthesis of Intermediate $ [00274] To the reaction mixture above was added 3-maleimidopropionic acid-Pfp ester 7 (1 mg, 3.0 μηιοΐ). The resulting mixture was stirred at room temperature in the dark for 3 h. Purification by RP-HPLC afforded intermediate 8 (7.7 mg, 83% for two steps). MS (ESI): m/e 1400 [M + 3H]3+. Synthesis ofSDM-101
[00275] A stirred solution of intermediate 8 (2.4 mg, 0.57 μπιοΐ) and thioanisole (10 μί, 85 μηιοΐ) in CH2CI2 (1 mL) was treated with trifluoroacetic acid (5 μΐ^, 65 μηιοΐ). The mixture was stirred at room temperature in the dark for 30 min. After the solvent was removed, the residue was purified by RP-HPLC to afford SDM-101 (0.6 mg, 27%) as a free flowing red powder after lyophilization. MS (ESI): m/e 1315 [M + 3H]3+.
Synthesis ofSDM-145
[00276] The mixture of SDM-101 (0.6 mg, 0.15 μπιοΐ) and mPEG(40 )-SH 9 (5 mg, 0.12 μηιοΐ) in PBS-EDTA buffer (0.5 mL, 137 mM NaCl, 7 mM Na2HP04, 3 mM C1, 1.4 mM 3PO4, 4 mM EDTA, pH 7.4) was stirred at room temperature in the dark for 5 h. Purification by RP- HPLC afforded SDM-145 as a red powder after lyophilization (3.0 mg, 60%>).
Example 3; Cleavage of SDM-145 by hMMP-9
[00277] Conjugate SDM-145 (3.0 mg) was dissolved in water (135 μί) to make a stock solution (0.5 mM). To a TCNB buffer (50 mM tris, 10 mM CaCl2, 150 mM NaCl, 0.05% Brij35, pH 7.5, 480 iV> in a HPLC sample vial was added SDM-145 stock solution (10 μί) and hMMP-9 (10 μΕ, 100 nM) purchased from EMD Millipore (Billerica, MA). The resulting solution was gently mixed well and incubated 37 °C.
[00278] Aliquots (15 uL) were removed at various time points and injected into an LC-MS equipped with a fluorescence spectrometer (ex: 480 ran; em: 560 nm). The cleavage reaction was complete after 17 hour. The peak at retention time -9.4 min was confirmed to be the cleaved poly- arginine fragment by MS (ESI): m/e 1388.9 [M + 2H]2+.
Example 4; Cleavage of SDM-145 by Cathepsin B
[00279] Conjugate SDM-145 (3.0 mg) was dissolved in water (135 μί) to make a stock solution (0.5 mM). To a sodium acetate buffer (25 mM NaAc, 1 mM EDTA, pH 5.0, 480 μί) in a HPLC sample vial was added conjugate SDM-145 stock solution (10 μΕ) and Cathepsin B, human liver (10 \L, 100 nM) purchased from EMD Millipore (Billerica, MA). The resulting solution was gently mixed well and incubated 37 °C.
[00280] Aliquots (15 uL) were removed at various time points and injected into an LC-MS equipped with a fluorescence spectrometer (ex: 480 nm; em: 560 nm). The cleavage reaction was su vacant , «yw»*s i complete after 17 hour. The peak at. retention time -9.0 mm was confirmed to be the freed doxorubicin by MS (ESI); m/e 566.4 [ ÷ Naf.
41j Exam le 5: Synthesis of SDM-143 and SP 46
= c-«. a*« . Ί ' ti i ti 5 I ¾ i s. c t i 6 I 8♦ ! m ·
P-5
:■ ;;: .i' .;' ¾;
* «<i
S* ; mm. O P, 1 h
Figure imgf000143_0001
19
7 ff : pen¾f!uot»piieno!
Figure imgf000143_0002
TFA, CHaC¾ :
ΐ
;;■ w,* >*..«< w . .,Λ.Χ .>«Γ
>*. <*. w. ». «i «
>TiPEGM0 )-SH
9
Figure imgf000143_0003
Synthesis of Intermediate 10
100282-1 o a solution of Intermediate S (230 mg, 0.16 mmol) aid e tide F-3 (700 mg, 0.17 mmoi.) in DM (5.0 J L) at room temperature in the dark was added N-mctJhylmorpholine (N M) (200 j*L, L8 mmo!) with stimng. The reaction was followed by LC-MS and completed m I h. Purification by RP-HPLC afforded intermediate 10 (426 mg, 62%) MS (ESI): m e' 1412.6 [M +
Synthesis of Intermediate 11.
|0O283| To the solution of 16 (148 mg, 2S.7 umol) in anhydrous DMF (4 raL) was added 3- rodeimicfopropionie acid-Pip ester 7 (20 rng, 59,7 ωοΐ) and 'N M (100 uL, 0.9 mrnof). The resulting mixture was stirred at room temperature m the dark for 20 h. Purification by RP-HPLC afforded intermediate 1 1 { 148 nig, 97%). MS (ESI): mfe 1463 | ÷ 3ΒΓ.
Synthesis efSD f-143
$0284] A stirred solution of intermediate 1 (120 mg, 23,3 μτήοί) in CHjCb (40 ml..} was treated with trifluoroacetic acid (100 uL), The mixture was stirred at room temperature in the dark for 5 h. After the solven was removed, the residue was purified by RP-HPLC to afford SD -143 (102 mg, 87%) as a free flowing red powder after iyophii Nation. MS (ESI / 377.9 [M + † 1 529.6 M - 4 FA. ÷ 3Hf*.
Synthesis of 8DM-146
I.0028SJ The mixture of SDM-I43 (26 mg, 5.0 μιηοί) and mPEG(4 K)-SH 9 (225 mg, 5,6 μτηοΐ) in PBS buffer (5.0 mL, p 7.4) was stirred at room temperature in the dark for 5 k Purification by RP-HPLC afforded SDM-J46 as a red powder after lyopbiltzation (172 mg, 77%).
Example 6: Synthesis of S M- 147
Figure imgf000144_0001
rf F.Q;S )-SH PH 74, P8S. S H
Figure imgf000144_0002
Synthesis of$DM~147
I0O28SJ The mixture of SDM-143 (1 17 mg5 22,7 μιηοι) and mPEG(2K>SH 12 (63 mg, 29,4 μτηοΐ) in PBS buffer (5,0 ml.., pH 7.4) was stirred at room temperature w the dark for 5 h.
Purification by RP-HPLC afforded SDM44? as a red powder after lyophiikatieu (172 mg, 7?%s. ALDl-TOF: ion clusters observed between approximately m z 5300-6700, with 44 Da differences and centered at a roximatel m/z 6082 (Figure S}»
Figure imgf000145_0001
13
NMM, DMF. 16 h
y Pip: peRtafSti TOiitetxil » o« *
Figure imgf000145_0002
SBM-14 raP£i3{¾¾-SH pH 7. , pggi s *
52
Figure imgf000145_0003
Sy thesi of 'Intermediate 14
100290) To a .solution of intermediate (I \ 5 mg5 0.08 mmoi) aid peptide F 6 (200 rag, 0.04 mjno!) hi F (5.0 ml.. } at ro m temperature hi the dark was added XniethylmorphoUne (NMM) (?ø μΐ,, 0,63 rnmol) with stirring. The reaction was followed by LC-MS and completed in 1 h. After the addition of 3~m&!eimidopropk>nk a id-Pfp ester 7 (30 ag, 89,6 μιηοΐ), the resulting mixture was stirred at mom temperature in the dark for 18 k Purification by RP-HPLC afforded intermediate 14 (Ϊ 1Θ mg, 43% tor 2 steps) MS (ESI): e 1618.8 \ M + 3H
SyMk&m ofSDM-144
002911 A stirred solution of intermediate 14 (10$ mg, 17.7 μ∞οΙ) in C¾C¾ (40 ml) was treated with trifluoroaeetic acid (100 μΕ). The mixture was stirred at room temperature in the dark tor 3 h. After the sol ent was removed, the residue was purified by RP-HPLC to aiford SOM-144 (84 mg, 81%) as a tree flowing red powder after lyephllization. MS (ESI); m e i 150,3 [M +
Figure imgf000146_0001
1533.2 [M + 3Hj3+, 1684.5 [M TFA + 3H]3*.
S ttthesii of SDM-148
[002 21 The mixture of SBM-1 4 (28 mg, 4.8 u ol) and raPEG(2K)~SH 12 (9.0 mg, 4.2 .mol) in PBS buffer (5.0 mL, pH 7.4) was stirred at room temperature in the dark for 5 h. Purification by RP-HPLC afforded SDM-148 as & red powder alter lyopfcilization (33 mg, 86%).
2931 Example 8: Synthesis of SDM-149
Figure imgf000146_0002
Synthesis of . Intermediate 16
00294] T a solution of Peptide P~3 (60 mg, 0.15 msnol) in glycine buffer (0.1 M, 20 U aniline. pH . , 2.0 m.L) at room temperature was added mPEO(2 )-ONH-» 15 GO nig, 0.14 mmol) with stirring. The reaction was followed by LC-MS and completed m 15 . Purification by RP- HPLC afforded intermediate 16 (70 mg, 83%),
Synthesis ofS.DM~J 9
|00295j To s mixture of 17 (8.7 mg, 10 μο>οΙ) and 16 (42 mg, 6.9 μηιοΐ) m anhydrous DMF ( 1 mL) was added N-methy!mofphoiine (10 μΐ, The mixture was stirred at room temperature for 20 h, Purification by RP-HPLC (mobile phase A: water; mobik phase B: acetonitrile) afforded SUM- 149 (22 mgs 46%),
00296] Example 9: Synthesis of SDM- 150
Figure imgf000147_0001
Synthesis ofSl)M~l$0
The mixture of 1.8 (8.7 mg, 6.8 μιηο!) and 16 (42 mg, 6.9 pmoi) n PBS (2.0 mL, pH
Figure imgf000147_0002
(1 mL.) mixed solvent was stirred at room temperature for 2 h. Purification by R! HPLC afforded S'OM-iSO (40.2 mg, 81%).
xample 10; Synthesis of Cathespta B 'Labile ACPP-Cortisoisc€oni¾g¾¾&
A Cathespin B Labile AC PP -Cortisone conjugate was synthesized as follows:
Figure imgf000148_0001
Pfp: pentafluorophenol
Figure imgf000148_0002
TFA, CH2<¾
Figure imgf000148_0003
[00299] Example 11 : Breast cancer Mouse Therapeutic model and Assay
[00300] Female BALB/c mice (8-10 weeks old) purchased from Harlan (Indianapolis, IN, 46259) or Charles River (Wilmington, MA, 01887) were used after 4-7 day of acclimatization period. All studies were conducted at research facility under the Institutional Animal Care and Use Committee (IACUC) approved protocol # EBl 1-002-009. On the first day of study, animals were weighed and assessed for health status. Only animals with no sign of disease were selected for the study. Each involved animal was lightly anesthetized with a mixture of ketamine/xylazine administered intraperitoneally to subdue voluntary movement. Highly metastatic 4T1 tumor cells (ATCC® Number CRL-2539™) suspended in DPBS/Matrigel™ (1 : 1 vol) were then injected subcutaneously (4xl05 tumor cells/50
Figure imgf000149_0001
into the right upper mammary fat pad of the lightly anesthetized animal. Each involved animal was then allowed to recover from anesthesia, housed back in the vivarium and kept under controlled environmental conditions.
[00301] Ten days after the subcutaneous implantation of 4T1 tumor cells into the upper mammary fat pad, the tumor size (width and length) of each involved animal was measured using a Mitutoyo 500-196-20 Absolute Digimatic Digital Caliper (Mitutoyo Corporation, Kanagawa, 213-0012,
3 2
Japan) and the individual tumor volume (mm ) calculated as follows: tumor volume= width x length/2. Tumor-bearing mice were then divided into 3-5 experimental groups (n= 3-4 tumor- bearing mice / group) based on identical averaged (Mean + SEM) tumor volume before the intravenous administration of vehicle or test compound. For each experimental group, each involved tumor-bearing mouse was restrained using the tail rotating tail injector (Cat.# RTI, Braintree Scientific, Inc., Braintree, MA 02185) and dosed with vehicle or test compound
1/2
administered intravenously using a 28G1'" insulin syringe (Cat.# 14-826-79, Becton Dickinson and Company, Franklin Lakes, NJ 07417). Vehicle or test compound was administered intravenously every other day or every three days based on the pharmacokinetic profile of the screened chemical entity. Individual body weight and tumor volume were systematically recorded on the day of dosing.
[00302] Twelve days after the first dosing, the individual body weight was recorded and the tumor size measured and the volume calculated as above. Each involved animal was then terminally anesthetized with the mixture of ketamine/xylazine. A blood sample was collected from each anesthetized tumor-bearing by cardiac puncture using a 1 cc Terumo Syringe Tuberculin with needle (25Gx5/8", Ref: SS-01T2516; Somerset, NJ 08873) and processed for plasma separation. The tumor was then dissected and collected. Plasma and tumor samples were kept frozen (-80°C) before being used for in vitro analyses.
[00303] Averaged tumor volume from individual experimental groups were compared to determine the therapeutic effect of the test compound. As shown in Figure 9, SDM-147 reduces the tumor volume in the murine 4T1 breast cancer model compared to vehicle. [00304] The examples and embodiments described herein are for illustrative purposes only and various modifications or changes suggested to persons skilled in the art are to be included within the spirit and purview of this application and scope of the appended claims.

Claims

WHAT IS CLAIMED IS:
1. A selective delivery molecule conjugate comprising:
(a) a selective delivery molecule of Formula I or Formula II, having the structure:
A-X-B-[CB-DB]
Formula I
Figure imgf000151_0001
Formula II
wherein,
X of Formula I or Formula II is a cleavable linker;
A of Formula I or Formula II is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B of Formula I or Formula II is a peptide with a sequence comprising 7 to 9 basic amino acids;
CB of Formula I or Formula II is 0-1 amino acid;
CM of Formula I or Formula II is 0- 1 amino acid;
M of Formula II is a macromolecule;
DB of Formula I or Formula II is a therapeutic agent or an imaging agent;
wherein [CM -M] of Formula II is bound to at any position on A or X; [CB-DB] of Formula I or Formula II is bound to any amino acid on B; and
(b) a carrier or targeting ligand, wherein the carrier or targeting ligand is covalently bound to the selective delivery molecule.
2. The molecule of claim 1, wherein the carrier or targeting ligand is covalently bound to any amino acid of A or any amino acid of B.
3. The molecule of claim 1 , wherein the targeting ligand is an antibody or a ligand that binds to a cell surface receptor.
4. The molecule of claim 1, wherein the targeting ligand binds to a tumor antigen or tumor- specific receptor.
5. The molecule of claim 1, wherein the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203.
6. The molecule of claim 1, wherein the carrier is a polyethylene glycol (PEG) polymer.
7. The molecule of claim 1, wherein the therapeutic agent is a chemotherapeutic agent, a cytotoxin, a steroid, an immunotherapeutic agent, a targeted therapy, or an anti-inflammatory agent.
8. The molecule of claim 1, wherein the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, auritstatin, taxol, or cortisone.
9. The molecule of claim 1, wherein CB and CM are each selected from any amino acid having a free thiol group, any amino acid with free amine, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxyl amine or hydrazine group.
10. The molecule of claim 1, wherein CB and CM are each selected from D-cysteine, D- glutamate, lysine, and para-4-acetyl L-phenylalanine.
11. The molecule of claim 1 , wherein CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxyl amine or hydrazine group.
12. The molecule of claim 1, wherein X is cleavable by an extracellular protease.
13. The molecule of claim 1, wherein X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac).
14. The molecule of claim 1, wherein M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer.
15. The molecule of claim 1 , wherein M is selected from dextran, a polyethylene glycol (PEG) polymer, albumin, or a combination thereof.
16. The molecule of claim 1, wherein M is selected from PEG lkDa, PEG 2kDa, PEG 3kDa, PEG 4kDa, PEG 5kDa, PEG lOkDa, PEG 12kDa, PEG 15kDa, PEG 20kDa, PEG 30kDa, and PEG40kDa.
17. The molecule of claim 1, wherein the selective delivery molecule is: SDM-101, SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM-110, SDM- 111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM-118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM- 128, SDM-129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM-135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141, SDM-142, SDM-143, SDM-144, SDM- 145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151, SDM-152, and SDM-153.
18. A selective delivery molecule conjugate comprising:
(a) a selective delivery molecule of Formula V, having the structure:
A-[CM-M]-X-B-Y-[CB-DB]
Formula V
wherein,
X is a cleavable linker;
Y is a cleavable linker; A is a peptide with a sequence comprising 5 to 9 acidic amino acids;
B is a peptide with a sequence comprising 7 to 9 basic amino acids;
CB and CM each independently comprise 0-1 amino acid;
M is a macromolecule;
DB is a therapeutic agent or an imaging agent,
wherein [CM-M] is bound to at any position on A or X, and [CB-Db] is bound to any amino acid on B.
19. The molecule of claim 18, further comprising a carrier or targeting ligand, wherein the carrier or targeting ligand covalently bound to the selective delivery molecule
20. The molecule of claim 19, wherein the carrier or targeting ligand is covalently bound to any amino acid of A or any amino acid of B.
21. The molecule of claim 19, wherein the targeting ligand is an antibody or a ligand that binds to a cell surface receptor.
22. The molecule of claim 19, wherein the targeting antibody binds to a tumor antigen or a tumor antigen or tumor-specific receptor.
23. The molecule of claim 19, wherein the targeting antibody is gemtuzumab, inotuumab, trastuzumab, lorvotuzumab, imgn388, SAR3419, BilB062, brentixumab, glembatumumab, SGN- 75, PSMA ADC, ASG-5ME or mdx-1203.
24. The molecule of claim 18, wherein the therapeutic agent is a chemotherapeutic agent, a cytotoxin, a steroid, an immunotherapeutic agent, a targeted therapy, or an anti-inflammatory agent.
25. The molecule of claim 18, wherein the therapeutic agent is doxorubicin, calicheamicin, maytansinoid, auritstatin or cortisone.
26. The molecule of claim 18, wherein CB and CM are each independently selected from a D amino acid, a L amino acid, an a-amino acid, a β-amino acid, or a τ-amino acid.
27. The molecule of claim 18, wherein CB and CM are each independently selected from any amino acid having a free thiol group, any amino acid with amine group, any amino acid having a N-terminal amine group, and any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
28. The molecule of claim 18, wherein CB and CM are each independently selected from D- cysteine, D-glutamate, lysine, and para-4-acetyl L-phenylalanine.
29. The molecule of claim 18, wherein CM is any amino acid with a side chain capable of forming an oxime or hydrazone bond upon reaction with a hydroxylamine or hydrazine group.
30. The molecule of claim 18, wherein X is cleavable by an extracellular protease.
31. The molecule of claim 18, wherein X comprises an amino acid sequence selected from: PLGLAG, PLG-C(me)-AG, RPLALWRS, ESPAYYTA, DPRSFL, PPRSFL, RLQLKL, and RLQLK(Ac).
32. The molecule of claim 18, wherein Y is cleavable by an intracellular protease.
33. The molecule of claim 18, wherein Y is cleavable by a lysosomal protease.
34. The molecule of claim 18, wherein Y is cleavable by a cathepsin or a caspase.
35. The molecule of claim 18, wherein Y is cleavable by Cathepsin B.
36. The molecule of claim 18, wherein Y comprises a self-immolative spacer.
37. The molecule of claim 18, wherein Y comprises a PABC spacer, a PABOH spacer, a BHMS spacer or any derivative thereof.
38. The molecule of claim 18, wherein M is selected from a protein, a natural polymer, a synthetic polymer, or a dendrimer.
39. The molecule of claim 18, wherein M is selected from dextran, a polyethylene glycol (PEG) polymer, albumin, or a combination thereof.
40. The molecule of claim 18, wherein M is selected from PEG lkDa, PEG 2kDa, PEG 3kDa, PEG 4kDa, PEG 5kDa, PEG lOkDa, PEG 12kDa, PEG 15kDa, PEG 20kDa, PEG 30kDa, and PEG40kDa.
41. The molecule of claim 18, wherein the selective delivery molecule of Formula V is: SDM- 101, SDM-102, SDM-103, SDM-104, SDM-105, SDM-106, SDM-107, SDM-108, SDM-109, SDM-110, SDM-111, SDM-112, SDM-113, SDM-114, SDM-115, SDM-116, SDM-117, SDM- 118, SDM-119, SDM-120, SDM-121, SDM-122, SDM-123, SDM-124, SDM-125, SDM-126, SDM-127, SDM-128, SDM-129, SDM-130, SDM-131, SDM-132, SDM-133, SDM-134, SDM- 135, SDM-136, SDM-137, SDM-138, SDM-139, SDM-140, SDM-141, SDM-142, SDM-143, SDM-144, SDM-145, SDM-146, SDM-147, SDM-148, SDM-149, SDM-150, SDM-151, SDM- 152, and SDM- 153.
42. A pharmaceutical composition comprising a selective delivery molecule conjugate of claim 1 or claim 18 and one or more pharmaceutically acceptable carriers, glidants, diluents, or excipients.
43. A method for treating cancer in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a selective delivery molecule conjugate of claim 1 or claim 18, thereby treating the cancer.
44. The method of claim 43, wherein the cancer is a breast cancer, colorectal cancer, ovarian cancer, lung cancer, esophageal cancer, pancreatic cancer, gastro-intestinal cancer, squamous cell carcinoma, prostate cancer, melanoma, or thyroid cancer.
45. A method for treating inflammation or an inflammatory disease in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a selective delivery molecule of claim 1 or claim 18, thereby treating the inflammation or inflammatory disease.
46. The method of claim 45, wherein the inflammation is associated with rheumatoid arthritis, osteoarthritis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, sepsis, erythema nodosum leprosum, multiple sclerosis, psoriasis, systemic lupus erythematosis, type I diabetes, atherosclerosis, encephalomyelitis, Alzheimer's disease, stroke, traumatic brain injury, Parkinson's disease or septic shock.
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