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ES2572915T3 - Nanopartículas conjugadas a un agente antinucleolina - Google Patents

Nanopartículas conjugadas a un agente antinucleolina Download PDF

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Publication number
ES2572915T3
ES2572915T3 ES12730288.3T ES12730288T ES2572915T3 ES 2572915 T3 ES2572915 T3 ES 2572915T3 ES 12730288 T ES12730288 T ES 12730288T ES 2572915 T3 ES2572915 T3 ES 2572915T3
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gro
human
antinucleolin
nucleolin
agent
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Paula J. Bates
Mohammad Tariq MALIK
Kyung A. Kang
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University of Louisville Research Foundation ULRF
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K49/00—Preparations for testing in vivo
    • A61K49/04—X-ray contrast preparations
    • 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
    • A61K47/61—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 the organic macromolecular compound being a polysaccharide or a derivative thereof
    • 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
    • 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
    • A61K47/69—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 conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A61K47/6921—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 conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere
    • A61K47/6923—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 conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being an inorganic particle, e.g. ceramic particles, silica particles, ferrite or synsorb
    • 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
    • A61K47/69—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 conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit
    • A61K47/6921—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 conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere
    • A61K47/6927—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 conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores
    • A61K47/6929—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 conjugate being characterised by physical or galenical forms, e.g. emulsion, particle, inclusion complex, stent or kit the form being a particulate, a powder, an adsorbate, a bead or a sphere the form being a solid microparticle having no hollow or gas-filled cores the form being a nanoparticle, e.g. an immuno-nanoparticle
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
    • Y10T428/00—Stock material or miscellaneous articles
    • Y10T428/29—Coated or structually defined flake, particle, cell, strand, strand portion, rod, filament, macroscopic fiber or mass thereof
    • Y10T428/2982—Particulate matter [e.g., sphere, flake, etc.]

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
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  • Engineering & Computer Science (AREA)
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  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • Immunology (AREA)
  • Nanotechnology (AREA)
  • Ceramic Engineering (AREA)
  • Inorganic Chemistry (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicinal Preparation (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)

Abstract

Composición, que comprende un agente antinucleolina conjugado a nanopartículas, en la que las nanopartículas no son magnéticas, ni de óxido de hierro, ni de poliacrilamida, y en la que las nanopartículas comprenden al menos un miembro seleccionado del grupo que consiste en oro, platino, iridio y paladio, y en la que el agente antinucleolina comprende: a) un oligonucleótido antinucleolina; y/o b) un anticuerpo; y/o c) una proteína de reconocimiento de nucleolina; y/o d) un GRO (oligonucléotido rico en guanosina).

Description

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Las Figuras 13A y B muestran la biodistribución de AS1411-GNP-Cy5: los ratones fueron tratados, sacrificados y se fotografiaron los órganos (A) y se examinaron para analizar la fluorescencia (B).
5 DESCRIPCIÓN DETALLADA
[0026] La presente invención hace uso del descubrimiento de que los agentes antinucleolina, conjugados a partículas, tales como aptámero conjugado a nanopartículas de oro, tienen un efecto antiproliferativo sobre el cáncer y tumores. Además, el aptámero conjugado a nanopartículas de oro en particular, tiene un efecto antiproliferativo 10 similar o mayor que el aptámero (oligonucleótido antinucleolina) por sí solo, demostrando efectos similares a solo de 1/10 a 1/100 de la dosis. Además, estos mismos agentes, que tienen preferiblemente un colorante fluorescente conjugado a la partícula o unida al agente antinucleolina, también se pueden usar tambbién como agentes de obtención de imágenes, tanto in vivo como ex vivo.
15 [0027] Los agentes antinucleolina incluyen (i) aptámeros, tales como GRO; (ii) anticuerpos antinucleolina; y (iii) proteínas que reconocen nucleolina. Ejemplos de aptámeros incluyen oligonucleótidos ricos en guanosina (GRO). Ejemplos de oligonucleótidos y ensayos adecuados se proporciona también en Miller et al. [7]. Las características de GRO incluyen:
(1) que tienen al menos 1 motivo GGT, 20 (2) que tienen preferiblemente 4-100 nucleótidos, aunque GRO que tienen muchos más nucleótidos son posibles,
(3) que tienen opcionalmente modificaciones químicas para mejorar la estabilidad.
[0028] Los GRO especialmente útiles forman estructuras de cuarteto de G, tal como se indica por un perfil de desnaturalización/renaturalización térmica reversible a 295 nm [6]. Los GRO preferidos también compiten con un 25 oligonucleótido de telómero para la unión a una proteína celular diana en un ensayo de desplazamiento por movilidad electroforética [6]. En algunos casos, la incorporación de los nucleótidos GRO en secuencias de ácidos nucleicos más grandes puede ser ventajosa; por ejemplo, para facilitar la unión de un ácido nucleico de GRO a un sustrato sin desnaturalizar el sitio de unión a nucleolina. Los ejemplos de oligonucleótidos se muestran en la Tabla 1; los oligonucleótidos preferidos incluyen las SEQ ID NOs: 1-7; 9-16; 19-30 y 31 de la Tabla 1. 30 Tabla 1. Gros no antisentido que se unen nucleolina y controles que no se unen1,2,3 .
GRO
Secuencia SEQ ID NO:
GRO29A1
tttggtggtg gtggttgtgg tggtggtgg 1
GRO29-2
tttggtggtg gtggttttgg tggtggtgg 2
GRO29-3
tttggtggtg gtggtggtgg tggtggtgg 3
GRO29-5
tttggtggtg gtggtttggg tggtggtgg 4
GRO29-13
tggtggtggt ggt 5
GRO14C
ggtggttgtg gtgg 6
GRO15A
gttgtttggg gtggt 7
GRO15B2
ttgggggggg tgggt 8
GRO25A
ggttggggtg ggtggggtgg gtggg 9
GRO26B1
ggtggtggtg gttgtggtgg tggtgg 10
GRO28A
tttggtggtg gtggttgtgg tggtggtg 11
GRO28B
tttggtggtg gtggtgtggt ggtggtgg 12
GRO29-6
ggtggtggtg gttgtggtgg tggtggttt 13
GRO32A
ggtggttgtg gtggttgtgg tggttgtggt gg 14
GRO32B
tttggtggtg gtggttgtgg tggtggtggt tt 15
GRO56A
ggtggtggtg gttgtggtgg tggtggttgt ggtggtggtg gttgtggtgg tggtgg 16
CRO
tttcctcctc ctccttctcc tcctcctcc 18
GRO A
ttagggttag ggttagggtt aggg 19
GRO B
ggtggtggtg g 20
GRO C
ggtggttgtg gtgg 21
GRO D
ggttggtgtg gttgg 22
GRO E
gggttttggg 23
GRO F
ttggttttgg ggttttggtt 24
GRO G1
ggttggtgtg gttgg 25
GRO H1
ggggttttgg 26
GRO I1
gggttttggg 27
GRO J1
ggggttttgg ggttttgggg ttttgggg 28
GRO K1
ttggggttgg ggttggggtt gggg 29
GRO L1
gggtgggtgg gtgggt 30
GRO M1
ttggttttgg ggttttggtt ttttgg 31
5
GRO N2
tttcctcctc ctccttctcc tcctcctcc 32
GRO O2
cctcctcctc cttctcctcc tcctcc 33
GRO P2
tggggt 34
GRO Q2
gcatgct 35
GRO R2
gcggtttgcg g 36
GRO S2
tagg 37
GRO T2
ggggttgggg tgtggggttg 38
1Indica un buen GRO que se une a nucleolina de membrana plasmática. 2Indica un control de nucleolina (unión a nucleolina no de membrana plasmática). 3Secuencias de GRO sin las denominaciones 1 o 2 tienen cierta actividad antiproliferativa.
[0029] También se puede usar cualquier anticuerpo que se une a nucleolina. En ciertos casos se prefieren anticuerpos monoclonales ya que se unen a epítopos individuales, específicos y definidos. En otros casos, sin embargo, se pueden usar anticuerpos policlonales capaces de interactuar con más de un epítopo en la nucleolina. Muchos anticuerpos antinucleolina están disponibles comercialmente, y de cualquier forma, se fabrican fácilmente. La Tabla 2 enumera unos cuantos anticuerpos antinucleolina disponibles comercialmente.
Tabla 2: Anticuerpos antinucleolina disponibles comercialmente
Anticuerpo
Fuente Fuente de antígeno
Anticuerpo monoclonal (mAb) de ratón p7-1A4
Developmental Studies Hybridoma Bank Oocitos de Xehopus laevis
mAb de ratón Sc-8031
Santa Cruz Biotech Humana
Ab policlonal (pAb) de cabra Sc-9893
Santa Cruz Biotech Humana
pAb de cabra Sc-9892
Santa Cruz Biotech Humana
mAb de ratón clon 4E2
MBL International Humana
mAb de ratón clon 3G4B2
Upstate Biotechnology Perro (células MDCK)
Nucleolina, humana (mAb de ratón)
MyBioSource Humana
Antinucleolina-Fosfo purificado, Thr76/Thr84 (mAb de ratón)
BioLegend Humana
Anticuerpo policlonal de Nucleolina de Conejo
Novus Biologicals Humana
Nucleolina (NCL, C23, FLJ45706)
US Biological Humana
FLJ59041, Proteína C23, mAb Mo xHu
Humana
Nucleolina (NCL, Nucl, C23, FLJ45706, Proteína C23) Pab Rb xHu
US Biological Humana
mAb antinucleolina humana Fosfo-Thr76/Thr84 de ratón, clon 10C7
Cell Sciences Humana
Anti-NCL / Nucleolina (pAb)
LifeSpan Biosciences Humana
Anticuerpo policlonal de ratón MaxPab purificado de NCL (B02P)
Abnova Humana
Anticuerpo policlonal de conejo MaxPab purificado de NCL (D01 P)
Abnova Humana
Anticuerpo monoclonal de NCL, clon 10C7 (mAb de ratón)
Abnova Humana
Anticuerpo monoclonal de nucleolina (4E2) (mAb de ratón)
Enzo Life Sciences Humana
Nucleolina, Anticuerpo Monoclonal de Ratón
Life Technologies Corporation Humana
Anticuerpo de NCL (Centro E443) (pAb de conejo)
Abgent Humana
Antinucleolina, clon 3G4B2 (mAb de ratón)
EMD Milipore Humana
NCL (pAb de conejo)
Proteintech Group Humana
Anticuerpo Monoclonal Antinucleolina de Ratón, sin conjugar, clon 3G4B20
Active Motif Humana
Nsr1p – monoclonal de ratón
EnCor Biotechnology Humana
Nucleolina (mAb de ratón)
Thermo Scientific Pierce Products Humana
Anticuerpo [4E2] de nucleolina (mAb de ratón)
GeneTex Humana
10 [0030] Las proteínas de reconocimiento de nucleolina son proteínas, diferentes de anticuerpos, que se unen específica y selectivamente a nucleolina. Los ejemplos incluyen la proteína ribosomal S3, péptidos F3 de migración tumoral [26, 27] y la miosina H9 (una miosina no muscular que se une la nucleolina de la superficie celular de las células endoteliales en vasos angiogénicos durante la tumorigénesis).
15 [0031] Los agentes antinucleolina se pueden conjugar a partículas fabricadas de una variedad de materiales, materiales sólidos, incluyendo (1) metales y elementos; (2) óxidos; (3) semiconductores; y (4) polímeros. Los
6
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Claims (1)

  1. imagen1
ES12730288.3T 2011-06-02 2012-06-01 Nanopartículas conjugadas a un agente antinucleolina Active ES2572915T3 (es)

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US201161492683P 2011-06-02 2011-06-02
US201161492683P 2011-06-02
PCT/US2012/040577 WO2012167173A1 (en) 2011-06-02 2012-06-01 Anti-nucleolin agent-conjugated nanoparticles

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PL (1) PL3446714T3 (es)
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