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• • 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
  • A61K31/185—Acids; Anhydrides, halides or salts thereof, e.g. sulfur acids, imidic, hydrazonic or hydroximic acids
• • 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
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
  • A61K31/415—1,2-Diazoles
• • 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
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
  • A61K31/433—Thidiazoles
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• • A—HUMAN NECESSITIES
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  • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
  • A61P1/16—Drugs for disorders of the alimentary tract or the digestive system for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
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  • A61P17/00—Drugs for dermatological disorders
  • A61P17/02—Drugs for dermatological disorders for treating wounds, ulcers, burns, scars, keloids, or the like
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P19/00—Drugs for skeletal disorders
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P19/00—Drugs for skeletal disorders
  • A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P27/00—Drugs for disorders of the senses
  • A61P27/02—Ophthalmic agents
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P27/00—Drugs for disorders of the senses
  • A61P27/02—Ophthalmic agents
  • A61P27/06—Antiglaucoma agents or miotics
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P9/00—Drugs for disorders of the cardiovascular system
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P9/00—Drugs for disorders of the cardiovascular system
  • A61P9/02—Non-specific cardiovascular stimulants, e.g. drugs for syncope, antihypotensives
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P9/00—Drugs for disorders of the cardiovascular system
  • A61P9/12—Antihypertensives

Definitions

• the present invention relates to methods of treating pulmonary hypertension, facilitating joint fusion, facilitating tendon and ligament repair, reducing the occurrence of secondary fracture, treating avascular necrosis, facilitating cartilage repair, facilitating bone healing after limb transplantation, facilitating liver regeneration, facilitating wound healing, reducing the occurrence of gastric ulceration, treating hypertension, facilitating the growth of tooth enamel or finger or toe nails, treating glaucoma, treating ocular hypertension, and repairing damage caused by metastatic bone disease using an EP 2 selective receptor agonist.
• the present invention provides methods of treating pulmonary hypertension, facilitating joint fusion, facilitating tendon and ligament repair, reducing the occurrence of secondary fracture, treating avascular necrosis, facilitating cartilage repair, facilitating bone healing after limb transplantation, facilitating liver regeneration, facilitating wound healing, reducing the occurrence of gastric ulceration, treating hypertension, facilitating the growth of tooth enamel or finger or toe nails, treating glaucoma, treating ocular hypertension, and repairing damage caused by metastatic bone disease using an EP 2 selective receptor agonist.
• Certain EP 2 selective receptor agonists are known in the art. See, for example, U.S. Patent Number 6,498,172.
• the present invention provides methods of treating pulmonary hypertension, facilitating joint fusion, facilitating tendon and ligament repair, reducing the occurrence of secondary fracture, treating avascular necrosis, facilitating cartilage repair, facilitating bone healing after limb transplantation, facilitating liver regeneration, facilitating wound healing, reducing the occurrence of gastric ulceration, treating hypertension, facilitating the growth of tooth enamel or finger or toe nails, treating glaucoma, treating ocular hypertension, and repairing damage caused by metastatic bone disease, the methods comprising administering to a patient in need thereof a therapeutically effective amount of an EP 2 selective receptor agonist.
• the present invention also provides such methods wherein the EP 2 selective receptor agonist is a compound of Formula I
• %l Formula I or a prodrug thereof, or a pharmaceutically acceptable salt thereof wherein A is SO 2 or CO; G is Ar, Ar'-V-Ar 2 , Ar-(C C 6 )alkylene, Ar-CONH-(C ⁇ -C 6 )alkylene, R 1 R 2 -amino, oxy(CrC 6 )alkylene, amino substituted with Ar, or amino substituted with Ar(C C 4 )alkylene and R 11 , wherein R 11 is H or (C C 8 )alkyl, R 1 and R 2 may be taken separately and are independently selected from H and (C C 8 )alkyl, or R 1 and R 2 are taken together with the nitrogen atom ofthe amino group to form a five- or six- membered azacycloalkyl, said azacycloalkyl optionally containing an oxygen atom and optionally mono-, di- or tri-substituted independently with up to two oxo, hydroxy, (C C 4 )
• alkylenes each optionally substituted with up to four substituents independently selected from fluoro or (C C 4 )alkyl,
• alkylenes each optionally substituted with up to four substituents each independently selected from fluoro or (C C 4 )alkyl,
• alkylenes each optionally substituted with up to four substituents each independently selected from fluoro or (C C 4 )alkyl,
• alkylenes and said ethenylene each optionally substituted with up to four substituents each independently selected from fluoro or (C C )alkyl,
• alkylenes and said ethenylene each optionally substituted with up to four substituents each independently selected from fluoro or (C r C )alkyl,
• Z is carboxyl, (C ⁇ -C 6 )alkoxycarbonyl, tetrazolyl, 1 ,2,4-oxadiazolyl, 5-oxo-1,2,4- oxadiazolyl, 5-oxo-1 ,2,4-thiadiazolyl, (C r C 4 )alkylsulfonylcarbamoyl or phenylsulfonylcarbamoyl; K is a bond, (C C 9 )alkylene, thio(C C )alkylene, (C C 4 )alkylenethio(C C 4 )alkylene, (C r C 4 )alkyleneoxy(CrC 4 )alkylene or oxy(C C 4 )alkylene, said (C C 9 )alkylene optionally mono-unsaturated and wherein, when K is not a bond, K is optionally mono-, di- or tri-substituted independently with chloro, fluor
• M is -Ar 3 , -Ar 4 -V 1 -Ar 5 , -Ar 4 -S-Ar 5 , -Ar 4 -SO-Ar 5 , -Ar 4 -SO 2 -Ar 5 or -Ar 4 -O-Ar 5 ;
• Ar is a partially saturated or fully unsaturated five to eight membered ring optionally having one to four heteroatoms selected independently from oxygen, sulfur and nitrogen, or a bicyclic ring consisting of two fused independently partially saturated, fully saturated or fully unsaturated five or six membered rings, taken independently, optionally having one to four heteroatoms selected independently from nitrogen, sulfur and oxygen, or a tricyclic ring consisting of three fused independently partially saturated, fully saturated or fully unsaturated five or six membered rings, optionally having one to four heteroatoms selected independently from nitrogen, sulfur and oxygen, said partially or fully saturated ring, bicyclic ring or tricyclic ring optionally having one or two ox
• C 4 alkoxycarbonylamino, hydroxysulfonyl, aminocarbonylamino or mono-N-, di-N,N-, di-N,N'- ortri-N,N,N'-(C C 4 )alkyl substituted aminocarbonylamino, sulfonamido, (C C 4 )alkylsulfonamido, amino, mono-N- or di-N,N-(C C 4 )alkylamino, carbamoyl, mono- N- or di-N,N-(C C 4 )alkylcarbamoyl, cyano, thiol, (CrC 6 )alkylthio, (C C 6 )alkylsulfinyl, (C C 4 )alkylsulfonyl or mono-N- or di-N,N-(C C 4 )alkylaminosulfinyl;
• Ar 3 , Ar 4 and Ar 5 are each independently a partially saturated, fully saturated or fully unsaturated five to eight membered ring optionally having one to four heteroatoms selected independently from oxygen, sulfur and nitrogen, or a bicyclic ring consisting of two fused independently partially saturated, fully saturated or fully unsaturated five or six membered rings, taken independently, optionally having one to four heteroatoms selected independently from nitrogen, sulfur and oxygen, or a tricyclic ring consisting of three fused independently partially saturated, fully saturated or fully unsaturated five or six membered rings, optionally having one to four heteroatoms selected independently from nitrogen, sulfur and oxygen, said partially or fully saturated ring, bicyclic ring or tricyclic ring optionally having one or two oxo groups substituted on carbon or one or two oxo groups substituted on sulfur; said Ar 3 , Ar 4 and Ar 5 moieties are optionally substituted on carbon or nitrogen, on one ring if the moiety is monocyclic, on one or both rings if the moiety is bicyclic
• W is oxy, thio, sulfino, sulfonyl, aminosulfonyl-, -mono-N-(C r C 4 )alkyleneaminosulfonyl-, sulfonylamino, N-(C C )alkylenesulfonylamino, carboxamido, N-(C r C 4 )alkylenecarboxamido, carboxamidooxy, N-(C
• X is a five or six membered aromatic ring optionally having one or two heteroatoms selected independently from oxygen, nitrogen, and sulfur; said ring optionally mono-, di- or tri-substituted independently with halo, (C C 3 )alkyl, trifluoromethyl, trifluoromethyloxy, difluoromethyloxy, hydroxyl, (C C 4 )alkoxy, or carbamoyl;
• R ⁇ R 2 , R 3 , R 4 R 5 , R 11 , R 31 , R 41 and R 51 when containing an alkyl, alkylene, alkenylene or alkynylene moiety, are optionally mono-, di- or tri-substituted on carbon independently with halo or hydroxy;
• V and V 1 are each independently a bond, thio(CrC )alkylene, (C C )alkylenethio, (CrC )alkyleneoxy, oxy(CrC 4 )alkylene or (C r C 3 )alkylene optionally mono- or disubstituted independently with hydroxy or fluoro; with the provisos that: a.
• the present invention also provides methods of treating pulmonary hypertension, facilitating joint fusion, facilitating tendon and ligament repair, reducing the occurrence of secondary fracture, treating avascular necrosis, facilitating cartilage repair, facilitating bone healing after limb transplantation, facilitating liver regeneration, facilitating wound healing, reducing the occurrence of gastric ulceration, treating hypertension, facilitating the growth of tooth enamel or finger or toe nails, treating glaucoma, treating ocular hypertension, and repairing damage caused by metastatic bone disease, the methods comprising administering to a patient in need thereof a therapeutically effective amount of (3-(((4-tert-butyl-benzyl)-(pyridine-3- sulfonyl)-amino)-methyl)-phenoxy)-acetic acid or a pharmaceutically acceptable salt thereof.
• EP 2 selective receptor agonists examples include EP 2 selective receptor agonists, and examples of EP 2 selective receptor agonists.
• a preferred EP 2 selective receptor agonist that can be used in the present methods is a compound of Formula I as defined above.
• a preferred group of compounds designated the A Group comprises those compounds having the Formula I as shown above, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein B is N; Z is carboxyl, (C C 6 )alkoxycarbonyl or tetrazolyl; Ar is phenyl, furyl, thienyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, 2H-pyrrolyl, 3H-pyrrolyl, pyrrolyl, 2-pyrrolinyl, 3- pyrrolinyl, pyrrolidinyl, 1 ,3-dioxolanyl, 2H-imidazolyl, 2-imidazolinyl, imidazolidinyl, 2- pyrazolin
• X is phenyl, thienyl, furanyl or thiazolyl, wherein X is optionally mono-, di- or tri-substituted with chloro, fluoro, methoxy, difluoromethoxy, trifluoromethoxy, trifluoromethyl or methyl.
• Another group of compounds which is preferred within the A Group, designated the C Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein A is CO; G is Ar; Q is -(C 2 -C 6 )alkylene-O-(C ⁇ -C 3 )alkylene-,
• X is phenyl, thienyl, furanyl or thiazolyl, wherein X is optionally mono-, di- or tri-substituted with chloro, fluoro, methoxy, difluoromethoxy, trifluoromethoxy, trifluoromethyl or methyl.
• Another group of compounds which is preferred within the A Group, designated the D Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein A is CO; G is R 1 R 2 -amino or amino substituted with Ar, or amino substituted with Ar(d- C 4 )alkylene and R 11 , wherein R 11 is H; Q is
• X is phenyl, thienyl, furanyl or thiazolyl, wherein X is optionally mono-, di- or tri-substituted with chloro, fluoro, methoxy, difluoromethoxy, trifluoromethoxy, trifluoromethyl or methyljand wherein R 1 and R 2 may be taken separately and are independently selected from H and (CrC 8 )alkyl, or R 1 and R 2 are taken together to form a five- or six- membered azacycloalkyl, said azacycloalkyl optionally containing an oxygen atom.
• Another group of compounds which is preferred within the G Group, designated the E Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein A is SO 2 ; G is R 1 R 2 -amino, or amino substituted with Ar and R 11 ; Q is
• X is phenyl, thienyl, furanyl or thiazolyl, wherein X is optionally mono-, di- or tri-substituted with chloro, fluoro, methoxy, difluoromethoxy, trifluoromethoxy, trifluoromethyl or methyl; and wherein R 1 and R 2 may be taken separately and are independently selected from H and (C C 8 )alkyl, or R 1 and R 2 are taken together to form a five- or six- membered azacycloalkyl, said azacycloalkyl optionally containing an oxygen atom.
• Another group of compounds which is preferred within the A Group, designated the F Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein A is SO 2 ; G is Ar, Ar(C r C 2 )alkylene or A ⁇ -V-Ar 2 ; Q is
• X is phenyl,pyrimidyi, pyridyl, thienyl, tetrahydrofuranyl, furanyl or thiazolyl, wherein X is optionally mono-, di- or trisubstituted with chloro, fluoro, methoxy, difluoromethoxy, trifluoromethoxy, trifluoromethyl or methyl.
• a particularly preferred group of compounds within the F Group, designated the FA Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein G is Ar or Ar-(C C 2 )- alkylene; Ar is phenyl, furyl, thienyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, isothiazolyl, 1 ,2,3-triazolyl, 1 ,2,4- triazolyl or 1,3,4-thiadiazolyl wherein each of said Ar groups is optionally substituted on carbon or nitrogen with R 1 , R 2 or R 3 ; Ar 4 is cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, phenyl, furyl, thieny
• K is methylene
• M is Ar 4 -Ar 5 , Ar 4 -O-Ar 5 or Ar 4 -S-Ar 5
• Ar is phenyl, pyridyl, pyrazolyl, imidazolyl, pyrimidyl, thienyl or thiazolyl
• Ar is optionally mono-, di- or tri-substituted on carbon or nitrogen with R 3 , R 4 or R 5 .
• a preferred group of compounds within the FB Group, designated the FC Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein M is Ar 4 -Ar 5 ; Ar is phenyl, pyridyl or imidazolyl; Ar 4 is phenyl, furanyl or pyridyl; and Ar 5 is cyclopentyl, cyclohexyl, cycloheptyl, phenyl, pyridyl, imidazolyl, pyrimidyl, thienyl, pyridazinyl, pyrazinyl, imidazolyl, pyrazolyl or thiazolyl, wherein Ar, Ar 4 and Ar 5 are optionally mono, -di- or tri-substituted on carbon or nitrogen independently with chloro, fluoro, methyl, methoxy, difluoromethoxy, trifluoromethyl or trifluoromethoxy.
• An especially preferred group of compounds within the FC Group designated the FD Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(C 5 - C 7 )alkylene-.
• Another especially preferred group of compounds within the FC Group designated the FE Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is CH 2 -X-CH 2 - and X is metaphenylene optionally mono- or di- substituted with chloro, fluoro, methoxy, difluoromethoxy, trifluoromethoxy, trifluoromethyl or methyl.
• a preferred group of compounds within the FE Group are those compounds, and pharmaceutically acceptable salts and prodrugs thereof, selected from (3- (((pyridine-3-sulfonyl)-(4-pyrimidin-5-yl-benzyl)-amino)-methyl)-phenyl)-acetic acid; (3-(((5-phenyl-furan-2-ylmethyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)-acetic acid; (3-((pyridine-3-sulfonyl)-(4-pyrimidin-2-yl-benzyl)-amino)-methyl)-phenyl)-acetic acid; (3-((pyridine-3-sulfonyl)-(4-thiazol-2-yl-benzyl)-amino)-methyl)-phenyl)-acetic acid; and (3-((4-pyrazin-2-yl-benzyl)-(pyridine-3-s)-
• An especially preferred compound within the FE Group is the compound wherein Ar is pyrid-3-yl; Z is carboxy; M is Ar 4 -Ar 5 wherein Ar 4 is a furanyl ring and Ar 5 is phenyl wherein said phenyl moiety is substituted at the 5-position of said furanyl ring; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• Another especially preferred compound within the FE Group is the compund wherein Ar is pyrid-3-yl; Z is carboxy; M is Ar 4 -Ar 5 wherein Ar 4 is phenyl and Ar 5 is pyrimid-2-yl and said pyrimid-2-yl moiety is substituted at the 4-position of said phenyl ring; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• Yet another especially preferred compound within th FE Group is the compound wherein Ar is pyrid-3-yl; Z is carboxy; M is Ar 4 -Ar 5 wherein Ar 4 is phenyl and Ar 5 is thiazol-2-yl and said thiazol-2-yl moiety is substituted at the 4-position of said phenyl ring; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• Yet another especially preferred compound within the FE Group is the compound wherein Ar is pyrid-3-yl; Z is carboxy; M is Ar 4 -Ar 5 wherein Ar 4 is phenyl and Ar 5 is pyrimid-5-yl and said pyrimid-5-yI moiety is substituted at the 4-position of said phenyl ring; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• Yet another especially preferred compound within the FE Group is the compound wherein Ar is pyrid-3-yl; Z is carboxy; M is Ar 4 -Ar 5 wherein Ar 4 is phenyl and Ar 5 is pyrazin-2-yl and said pyrazin-2-yl is substituted at the 4-position of said phenyl ring; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• a preferred group of compounds within the FC Group, designated the G Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(C 2 -C 4 )- alkylene-thienyl-, -(C 2 -C 4 )-alkylene-furanyl- or -(C 2 -C 4 )-alkylene-thiazolyk
• An especially preferred compound within the G Group is 5-(3-((pyridine-3- sulfonyl)-(4-thiazol-2-yl-benzyl)-amino)-propyl)-thiophene-2-carboxylic acid.
• An especially preferred compound within the G Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is n-propylenyl; X is thienyl; Z is carboxy; Ar is 3-pyridyl; Ar 4 is phenyl; and Ar 5 is 2-thiazolyl; said 2-thiazolyl being substituted at the 4-position of said phenyl.
• Another especially preferred group of compounds within the FC Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -CH 2 -X-O-CH 2 -; Ar 4 is phenyl or pyridyl; said phenyl and pyridyl are optionally substituted with chloro, fluoro, methoxy, difluoromethoxy, trifluoromethoxy, trifluoromethyl and methyl; and X is metaphenylene.
• a preferred group of compounds within the H Group are (3-(((4-cyclohexyl- benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenoxy)-acetic acid; (3-(((pyridine-3- sulfonyl)-(4-pyridin-2-yl-benzyl)-amino)-methyl)-phenoxy)-acetic acid; (3-((pyridine-3- sulfonyl)-(4- pyridin-3-yl-benzyl)-amino)-methyl)-phenoxy)-acetic acid; (3-(((pyridine- 3-sulfonyl)-(4-pyridin-4-yl-benzyl)-amino)-methyl)-phenoxy)-acetic acid; and (3- (((pyridine-3-sulfonyl)-(4-thiazol-2-yl-benzyl)-amino)-methyl)-phenoxy)-
• An especially preferred compound within the H Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar is pyrid-3-yl; Z is carboxy; Ar 4 is phenyl; Ar 5 is cyclohexyl; and said cyclohexyl moiety is substituted at the 4-position of said phenyl ring.
• Another especially preferred compound within the H Group is the compound wherein Ar is pyrid-3-yl; Z is carboxy; Ar 4 is phenyl; Ar 5 is thiazol-2-yl; and said thiazol-2-yl moiety is substituted at the 4-position of said phenyl ring.
• Yet another especially preferred compound within the H Group is the compound wherein Ar is pyrid-3-yl; Z is carboxy; Ar 4 is phenyl; Ar 5 is 2-pyridyl; and said 2-pyridyl moiety is substituted at the 4-position of said phenyl ring.
• Yet another especially preferred compound within the H Group is the compound wherein Ar is pyrid-3-yl; Z is carboxy; Ar 4 is phenyl; Ar 5 is 3-pyridyl; and said 3-pyridyl moiety is substituted at the 4-position of said phenyl ring.
• Yet another especially preferred compound within the H Group is the compound wherein Ar is pyrid-3-yl; Z is carboxy; Ar 4 is phenyl; Ar 5 is 4-pyridyl; and said 4-pyridyl moiety is substituted at the 4-position of said phenyl ring.
• a preferred group of compounds within the FA Group, designated the I Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein K is methylene, G is Ar; Ar is phenyl, pyridazinyl, pyrazolyl, pyrazinyl, pyridyl, imidazolyl, pyrimidyl, thienyl or thiazolyl, Ar is optionally mono-, di- or tri-substituted with R 3 , R 4 or R 5 , and M is Ar 3 , wherein said Ar 3 is cyclopentyl, cyclohexyl, phenyl, thienyl, pyridazinyl, pyrimidinyl, pyrazinyl, indolyl, benzofuryl, benzo(b)thienyl, benzoxazolyl, benzthiazolyl, quinolinyl, isoquinolinyl, naphth
• An especially preferred group of compounds within the I Group are (3-(((2,3- dihydro-benzo[1 ,4]dioxin-6-ylmethyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)- acetic acid; and (3-((benzofuran-2-ylmethyl-(pyridine-3-sulfonyl)-amino)-methyl)- phenyl)-acetic acid.
• An especially preferred compound within the I Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compound and prodrugs, wherein Ar is pyrid-3-yl; Z is carboxy; M is 6-(1 ,4-benzodioxan); and Q is - CH 2 -X-CH 2 - wherein X is metaphenylene.
• Another especially preferred compound within the I Group is the compound wherein Ar is pyrid-3-yl; Z is carboxy; M is 2-benzofuryl; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• Another especially preferred group of compounds within the I Group, designated the J Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar is phenyl, pyridyl or imidazolyl, said phenyl, pyridyl and imidazolyl optionally substituted independently with chloro, fluoro, methyl, methoxy, difluoromethoxy, trifluoromethyl or trifluoromethoxy; Ar 3 is phenyl substituted with R 31 , wherein R 31 is (C C 7 )alkyl, mono-N- or di-N, N-(C C 4 )alkylamine, or (C r C 5 )alkoxy, said (C C 7 )al
• a preferred group of compounds within the J Group, designated the K Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(C 5 -C 7 )alkylene-.
• Another preferred group of compounds within the J Group, designated the L Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -CH 2 -X-CH 2 - and X is phenyl optionally mono-, di- or tri- substituted with chloro, fluoro, methoxy, difluoromethoxy, trifluoromethoxy, trifluoromethyl or methyl.
• An especially preferred group of compounds within the L Group are (3-(((4- butyl-benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)-acetic acid; (3-
• An especially preferred compound within the L Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar is pyrid-3-yl; Z is carboxy; M is phenyl substituted at the 4- position with n-butyl; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• Another especially preferred compound within the L Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar is phenyl; Z is carboxy; M is phenyl substituted at the 4-position with n-butyl; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• Yet another especially preferred compound within the L Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar is 4-(1-methyl-imidazolyl); Z is carboxy; M is phenyl substituted at the 4-position with n-butyl; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• Yet another especially preferred compound within the L Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar is pyrid-3-yl; Z is oarboxy; M is phenyl substituted at the 4-position with dimethylamino; and Q is -CH 2 -X-CH 2 - wherein X is metaphenylene.
• Another preferred group of compounds within the J Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(C 2 -C )alkylene-thienyl, -(C 2 - C 4 )alkylene-furanyl or -(C 2 -C )alkylene-thiazolyl.
• a preferred group of compounds within the J Group, designated the M Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(CrC 2 )-X-O-(C 1 -C 2 )alkylene- and X is metaphenylene, said X being optionally mono-, di- or tri-substituted with chloro, fluoro, methoxy, difluoromethoxy, trifluoromethoxy, trifluoromethyl or methyl.
• An especially preferred group of compounds within the M Group are (3-(((4- dimethylamino-benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenoxy)-acetic acid and (3-(((4-tert-butyl-benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenoxy)-acetic acid.
• An especially preferred compound within the M Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar is pyrid-3-yl; Z is carboxy; M is phenyl substituted at the 4- position with dimethylamino; and Q is -CH 2 -X-0-CH 2 - wherein X is metaphenylene.
• Another especially preferred compound within the M Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar is pyrid-3-yl; Z is carboxy; M is phenyl substituted at the 4- position with tert-butyl; and Q is -CH 2 -X-O-CH 2 - wherein X is metaphenylene.
• Another preferred group of compounds within the FA Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein G is Ar; K is (C 2 -C 4 ) alkylene or n-propenylene; Ar is phenyl, pyrazolyl, pyridazinyl, pyrazinyl, pyridyl, imidazolyl, pyrimidyl, thienyl or thiazolyl, wherein Ar is optionally mono-, di- or trisubstituted with R 3 , R 4 or R 5 ; and M is Ar 3 , optionally mono-, di- or tri-substituted with chloro, fluoro, methyl, methoxy, difluoromethoxy, trifluoromethoxy and trifluoromethyl.
• An especially preferred compound within the N Group is f.rat?s-(3-(((3-(3,5- dichloro-phenyl)-allyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)-acetic acid.
• An especially preferred compound within the N Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein K is frans-n-propenylene, said M group being attached to the 1- position of the n-propenylene and said N atom being attached to the 3-position of the n-propenylene; Ar is pyrid-3-yl; M is phenyl 3,5-disubstituted with chloro; Z is carboxy; and Q is CH 2 -X-CH 2 - wherein X is metaphenylene.
• a preferred group of compounds within the N Group, designated the O Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar 3 is phenyl optionally substituted with chloro, fluoro, methyl, methoxy, difluoromethoxy, trifluoromethoxy or trifluoromethyl.
• a preferred group of compounds within the O Group, designated the P Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(C 5 -C 7 )alkylene-.
• Q Group Another group of compounds within the O Group, designated the Q Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -CH 2 -X-CH 2 - and X is metaphenylene.
• R Yet another group of compounds within the O Group, designated the R
• Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(C 2 - C 4 )alkylene-X- and X is furanyl, thienyl or thiazolyl.
• Yet another preferred group of compounds within the O Group designated the S Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(C C 2 )-X-O- (CrC 2 )alkylene- and X is metaphenylene.
• T Group Another preferred group of compounds within the FA Group, designated the T Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein G is Ar; K is thioethylene or oxyethylene, Ar is phenyl, pyrazolyl, pyridazinyl, pyrazinyl, pyridyl, imidazolyl, pyrimidyl, thienyl or thiazolyl, wherein Ar is optionally substituted with up to three R 3 , R 4 or R 5 ; and M is Ar 3 , optionally mono-, di- or tri-substituted with chloro, fluoro, methyl, difluoromethoxy, trifluoromethoxy or trifluoromethyl.
• a preferred group of compounds within the T Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Ar 3 is phenyl.
• a preferred group of compounds within the U Group, designated the V Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(C 5 -C 7 )alkylene-.
• Another preferred group of compounds within the U Group designated the W Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -CH 2 -X-CH 2 - and X is metaphenylene.
• Another preferred group of compounds within the U Group, designated the X Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(C 2 - C 4 )alkylene-X- and X is furanyl, thienyl or thiazolyl.
• Another preferred group of compounds within the U Group designated the Y Group, comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein Q is -(CrC 2 )-X-O- (C ⁇ -C 2 )alkylene- and X is metaphenylene.
• An especially preferred compound within the Y Group is (3-(((2-(3,5-dichloro- phenoxy)-ethyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenoxy)-acetic acid.
• An especially preferred compound within the Y Group is the compound, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein K is ethylenyloxy; said M group being attached to the oxygen atom of the ethylenyloxy group and said N atom being attached to the 2-position of the ethylenyloxy group; Ar is pyrid-3-yl; M is phenyl 3,5-disubstituted with chloro; Z is carboxy and Q is -CH 2 -X-O-CH 2 - wherein X is a second phenyl ring and said CH 2 and OCH 2 substituents are situated in a meta substitution pattern on said second phenyl ring.
• Another preferred group of compounds designated the Z Group, comprises those compounds of Formula I, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein B is CH.
• a preferred group of compounds within the Z Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein A is CO; G is Ar, K is methylenyl, propylenyl, propenylenyl or oxyethylenyl; M is Ar 3 or Ar 4 -Ar 5 ; Ar 3 is phenyl or pyridyl; Ar 4 is phenyl, thienyl, pyridyl or furanyl; Ar 5 is (C 5 -C 7 ) cycloalkyl, phenyl, pyridyl, imidazolyl, pyrimidyl, thienyl, pyridazinyl, pyrazinyl, imidazolyl, pyrazolyl or thiazolyl ; Ar is phenyl, pyrazolyl, pyridazinyl, pyrazinyl, pyridyl, imidazoly
• Another especially preferred group of compounds within the Z Group comprises those compounds, prodrugs thereof and pharmaceutically acceptable salts of said compounds and said prodrugs, wherein A is CO; G is Ar, K is methylenyl, propylenyl, propenylenyl or oxyethylenyl; M is Ar 3 or Ar 4 -Ar 5 ; Ar 3 is phenyl or pyridyl; Ar 4 is phenyl, thienyl, pyridyl or furanyl; Ar 5 is (C 5 -C 7 ) cycloalkyl, phenyl, pyridyl, imidazolyl, pyrimidyl, thienyl, pyridazinyl, pyrazinyl, imidazolyl, pyrazolyl or thiazolyl ; Ar is phenyl, pyrazolyl, pyridazinyl, pyrazinyl, pyridyl, imidazo
• Exemplary five to six membered aromatic rings optionally having one or two heteroatoms selected independently from oxygen, nitrogen and sulfur are phenyl, furyl, thienyl, pyrrolyl, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, isoxazolyl, isothiazolyl, pyridyl, pyridiazinyl, pyrimidinyl and pyrazinyl.
• Exemplary partially saturated, fully saturated or fully unsaturated five to eight membered rings optionally having one to four heteroatoms selected independently from oxygen, sulfur and nitrogen (i.e., Ar, Ar 1 and Ar 2 ) are cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl and phenyl.
• FIG. 1 Further exemplary five membered rings are furyl, thienyl, 2H-pyrrolyl, 3H-pyrrolyl, pyrrolyl, 2-pyrrolinyl, 3-pyrrolinyl, pyrrolidinyl, 1,3- dioxolanyl, oxazolyl, thiazolyl, imidazolyl, 2H-imidazolyl, 2-imidazolinyl, imidazolidinyl, pyrazolyl, 2-pyrazolinyl, pyrazolidinyl, isoxazolyl, isothiazolyl, 1 ,2-dithiolyl, 1,3- dithiolyl, 3H-1,2-oxathiolyl, 1,2,3-oxadiazolyl, 1 ,2,4-oxadiazolyl, 1,2,5-oxadiazolyl, 1,3,4-oxadiazolyl, 1 ,2,3-triazolyl, 1 ,2,4-tri
• FIG. 1 For exemplary six membered rings are 2H-pyranyl, 4H-pyranyl, pyridyl, piperidinyl, 1,2-dioxinyl, 1,3-dioxinyl, 1 ,4-dioxanyl, morpholinyl, 1 ,4-dithianyl, thiomorpholinyl, pyridazinyl, pyrimidinyl, pyrazinyl, piperazinyl, 1,3,5-triazinyl, 1,2,4- triazinyl, 1,2,3-triazinyl, 1,3,5-trithianyl, 4H-1 ,2-oxazinyl, 2H-1,3-oxazinyl, 6H-1.3- oxazinyl, 6H-1 ,2-oxazinyl, 1,4-oxazinyl, 2 H-1 ,2-oxazinyl, 4H-1 ,4-oxazinyl, 1,2,
• Further exemplary seven membered rings are azepinyl, oxepinyl, thiepinyl and 1 ,2,4-diazepinyl. Further exemplary eight membered rings are cyclooctyl, cyclooctenyl and cyclooctadienyl.
• Exemplary bicyclic rings consisting of two fused independently partially saturated, fully saturated or fully unsaturated five and/or six membered rings, taken independently, optionally having one to four heteroatoms selected independently from nitrogen, sulfur and oxygen are indolizinyl, indolyl, isoindolyl, 3H-indolyl, 1 H- isoindolyl, indolinyl, cyclopenta(b)pyridinyl, pyrano(3,4-b)pyrrolyl, benzofuryl, isobenzofuryl, benzo(b)thienyl, benzo(c)thienyl, 1H-indazolyl, indoxazinyl, benzoxazolyl, anthranilyl, benzimidazolyl, benzthiazolyl, purinyl, 4H-quinolizinyl, quinolinyl, isoquinolinyl, cinnolinyl, phthalazinyl, quinazol
• Exemplary tricyclic rings consisting of three fused independently partially saturated, fully saturated or fully unsaturated five and/or six membered rings, taken independently, optionally having one to four heteroatoms selected independently from nitrogen, sulfur and oxygen are indacenyl, biphenylenyl, acenaphthylenyl, fluorenyl, phenalenyl, phenanthrenyl, anthracenyl, naphthothienyl, thianthrenyl, xanthenyl, phenoxathiinyl, carbazolyl, carbolinyl, phenanthridinyl, acridinyl, perimidinyl, phenanthrolinyl, phenazinyl, phenothiazinyl and phenoxazinyl.
• alkylene saturated hydrocarbon (straight chain or branched) wherein a hydrogen atom is removed from each of the terminal carbons.
• alkylene groups (assuming the designated length encompases the particular example) are methylene, ethylene, propylene, butylene, pentylene, hexylene and heptylene.
• alkenylene is meant a hydrocarbon containing monounsaturation in the form of one double bond wherein said hydrocarbon is straight chain or branched and wherein a hydrogen atom is removed from each of the terminal carbons.
• alkenylene is meant a hydrocarbon containing monounsaturation in the form of one double bond wherein said hydrocarbon is straight chain or branched and wherein a hydrogen atom is removed from each of the terminal carbons.
• ethenylene (or vinylene) propenylene, butenylene, pentenylene, hexenylene and heptenylene.
• alkynylene is meant a hydrocarbon containing di-unsaturation in the form of one triple bond wherein said hydrocarbon is straight chain or branched and wherein a hydrogen atom is removed from each of the terminal carbons.
• alkynylene a hydrocarbon containing di-unsaturation in the form of one triple bond wherein said hydrocarbon is straight chain or branched and wherein a hydrogen atom is removed from each of the terminal carbons.
• exemplary of such groups are ethynylene, propynylene, butynylene, pentynylene, hexynylene and heptynylene.
• halo is meant chloro, bromo, iodo, or fluoro.
• alkyl straight chain saturated hydrocarbon or branched saturated hydrocarbon.
• alkyl groups (assuming the designated length encompasses the particular example) are methyl, ethyl, propyl, isopropyl, butyl, sec- butyl, tertiary butyl, pentyl, isopentyl, neopentyl, tertiary pentyl, 1-methylbutyl, 2- methylbutyl, 3-methylbutyl, hexyl, isohexyl, heptyl and octyl.
• alkoxy straight chain saturated alkyl or branched saturated alkyl bonded through an oxy.
• alkoxy groups are methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tertiary butoxy, pentoxy, isopentoxy, neopentoxy, tertiary pentoxy, hexoxy, isohexoxy, heptoxy and octoxy.
• the term mono-N- or di-N,N-(C C x )alkyl refers to the (C C x )alkyl moiety taken independently when it is di-N,N-(C C x )alkyl...(x refers to integers and is taken independently when two (C r C x )alkyl groups are present, e.g., methylethylamino is within the scope of di-N,N-(C ⁇ -C x )alkyl).
• M moieties defined above are optionally substituted (e.g., the mere listing of a substituent such as R 1 in a subgenus or dependent claim does not mean that M is always substituted with the R 1 moiety unless it is stated that the M moiety is substituted with R 1 ).
• K is a bond and M is phenyl
• said phenyl group is substituted with one to three substituents.
• Ar or Ar 1 is a fully saturated five to eight membered ring, said ring is unsubstituted.
• a carbocyclic or heterocyclic moiety may be bonded or otherwise attached to a designated substrate, through differing ring atoms without denoting a specific point of attachment, then all possible points are intended, whether through a carbon atom or, for example, a trivalent nitrogen atom.
• pyridyl means 2-, 3-, or 4-pyridyl
• thienyl means 2-, or 3-thienyl, and so forth.
• a particularly preferred compound of Formula I is (3-(((4-tert-butyl-benzyl)- (pyridine-3-sulfonyl)-amino)-methyl)-phenoxy)-acetic acid, or a pharmaceitcally acceptable salt or prodrug thereof, or a salt of a prodrug.
• a particularly preferred salt is the sodium salt.
• Other EP 2 selective receptor agonists that can be used in the present invention include the prostaglandin receptor agonists disclosed in U.S. patent numbers 6,288,120; and 6,124,314; and PCT published patent application WO 98/58911 (PCT/IB98/00866).
• 6,288,120 is 7-[(4-butyl-benzyl)-methanesulfonyl-amino]-heptanoic acid or a pharmaceutically acceptable salt or prodrug thereof, or a salt of a prodrug.
• a preferred salt of 7-[ 4- butyl-benzyl)-methanesulfonyl-amino]-heptanoic acid is the monosodium salt.
• Other EP 2 selective receptor agonists that can be used in the present invention include the compounds disclosed in the following: Burk, Robert M.; Holoboski, Mark; Posner, Mari F., Preparation of prostaglandin E2 analogs as EP2- receptor agonists-US patent application no.
• Pulmonary hypertension also known as primary pulmonary hypertension
• Pulmonary hypertension is a disease of unknown origin that involves the medium and small pulmonary arteries and results in right ventricular failure or fatal syncope, typically 2 to 5 years after detection.
• Intimal hyperplasia and consequent narrowing of the vessel lumen are always present. Areas of medial hypertrophy and hyperplasia, irreversible plexiform lesions, and necrotizing arteritis (plexogenic arteriopathy) occur in more advanced cases.
• Those skilled in the art are familiar with the identification of patients having pulmonary hypertension.
• the present methods can also be used in facilitating joint fusion.
• joint fusions include fusion of bones in the wrist or ankle as well as other joints. In a joint fusion, two or more bones are fused together.
• the present methods can also be used to facilitate tendon and/or ligament repair.
• the repair can comprise the strengthening of a tendon or ligament or can comprise the reconstruction of a damaged portion of a tendon and/or ligament.
• Another aspect of tendon and ligament repair is to strengthen or repair the attachment of a tendon or ligament to a bone.
• the present methods can also be used to reduce the occurrence of secondary fractures.
• a secondary fracture is a fracture subsequent to a primary fracture. Once a fracture has occurred, the present methods can be used to prevent another fracture from occurring or reduce the magnitude or complexity of any secondary fracture.
• the present method may be used to help avoid or ameliorate the extent of a second fracture in the hip either on the same side of the hip as the first fracture or on the other side of the hip.
• the prevention or amelioration of damage caused by secondary fractures is also important in cases where the risk of secondary fractures is enhanced, such as in response to chemotherapy. Also, the prevention of secondary fractures in the spine and spinal stabilization are important.
• the present invention also provides methods of treating avascular necrosis.
• Avascular necrosis is characterized by cell death in bone resulting from a compromised blood supply.
• the hip, femur and shoulder are commonly affected bones.
• Various conditions have been associated with avascular necrosis including fracture of the femoral neck, hip dislocation, decompression sickness, sickle cell disease, radiotherapy, Gaucher's disease, and corticosteroid high-dose therapy.
• avascular necrosis Other conditions that have been associated with avascular necrosis include systemic lupus erythrmatosus, renal transplantation, polycythemia versa, Cushing's syndrome, Diabetes mellitus, atherosclerosis, cytotoxic chemotherapy, alcohol abuse, fatty liver, psoriasis, pancreatitis, pancreatic cancer and gout.
• the presence of a condition associated with avascular necrosis can be an indicator to apply the methods of the present invention to prevent or ameliorate the occurrence of avascualr necrosis in a patient.
• the present invention also be used to facilitate cartilage repair, facilitate bone healing after limb transplantation, facilitate liver regeneration, facilitate wound healing, reduce the occurrence of gastric ulceration, treat hypertension, facilitate the growth of tooth enamel or finger or toe nails, treat glaucoma, treat ocular hypertension, or repair damage caused by metastatic bone disease, which conditions are well known to those skilled in the art. Patients having such conditions are easily identified by those skilled in the art.
• facilitating means to make the methods less difficult or improve the speed of the methods.
• facilitating bone fusion means to make the procedure less difficult to accomplish or proceed more rapidly in the presence of an EP 2 selective receptor agonist than in the absence of an EP 2 selective receptor agonist.
• a preferred dosage is about 0.001 to 100 mg/kg/day of an EP 2 selective receptor agonist, such as a compound of Formula I.
• An especially preferred dosage is about 0.01 to 10 mg/kg/day of an EP 2 selective receptor agonist, such as a compound of Formula I.
• the present invention is also concerned with pharmaceutical compositions comprising an EP 2 selective receptor agonist, such as a compound of Formula I, and a carrier, solvent, diluent and the like.
• Another aspect of this invention is directed to combinations of an EP 2 selective receptor agonist, such as a compound of Formula I, and other therapeutically useful compounds.
• the combinations of this invention comprise a therapeutically effective amount of a first compound, said first compound being an EP 2 selective receptor agonist, such as a compound of Formula I; and a therapeutically effective amount of a second compound, the second compound being an anti-resorptive agent such as an estrogen agonist antagonist or a bisphosphonate.
• An estrogen agonist/antagonist is also called a selective estrogen receptor modulator (SERM).
• kits comprising: a. an amount of an EP 2 selective receptor agonist, such as a compound of Formula I, and a pharmaceutically acceptable carrier or diluent in a first unit dosage form; b. an amount of an anti-resorptive agent, and a pharmaceutically acceptable carrier or diluent in a second unit dosage form; and c. a container.
• compositions or kits comprising an EP 2 selective receptor agonist, such as a compound of Formula I, and another bone anabolic agent (although the other bone anabolic agent may be another EP 2 selective receptor agonist, such as a different compound of Formula.!).
• Such compositions comprise a therapeutically effective amount of a first compound, said first compound being an EP 2 selective receptor agonist, such as a compound of Formula I; and a therapeutically effective amount of a second compound, said second compound being another bone anabolic agent.
• the first compound can administered at the same time as the second compound in the same dosage form or in different dosage forms.
• the first compound and second compound can be administered at different times.
• the combinations of the present invention can comprise more than two compounds.
• two EP 2 selective receptor agonists and an anti- repsorptive or bone anabolic compound can be administered to a patient.
• kits comprising: a. an amount of an EP 2 receptor selective agonist, such as a compound of Formula I, in a first unit dosage form; b. an amount of a second compound, said second compound being another bone anabolic agent in a second unit dosage form; and c. a container.
• an EP 2 receptor selective agonist such as a compound of Formula I
• Preferred bone anabolic agents include IGF-1, prostaglandins, prostaglandin agonists/antagonists, sodium fluoride, parathyroid hormone (PTH), active fragments of parathyroid hormone, parathyroid hormone related peptides and active fragments and analogues of parathyroid hormone related peptides, growth hormones 'or growth hormone secretagogues and the pharmaceutically acceptable salts or prodrugs thereof or a salt of a prodrug.
• the present invention has an aspect that relates to a combination of active ingredients, which may be administered separately, the invention also relates to combining separate pharmaceutical compositions in kit form.
• the kit comprises two separate pharmaceutical compositions: an EP 2 selective receptor agonist, such as a compound of Formula I and a second compound as described above.
• the kit comprises container means for containing the separate compositions such as a divided bottle or a divided foil packet, however, the separate compositions may also be contained within a single, undivided container.
• the kit comprises directions for the administration of the separate components.
• the kit form is particularly advantageous when the separate components are preferably administered in different dosage forms (e.g., oral and parenteral), are administered at different dosage intervals, or when titration of the individual components of the combination is desired by the prescribing physician.
• An example of such a kit is a so-called blister pack.
• Blister packs are well known in the packaging industry and are being widely used for the packaging of pharmaceutical unit dosage forms (tablets, capsules, and the like).
• Blister packs generally consist of a sheet of relatively stiff material covered with a foil of a preferably transparent plastic material.
• the recesses have the size and shape of the tablets or capsules to be packed.
• the tablets or capsules are placed in the recesses and the sheet of relatively stiff material is sealed against the plastic foil at the face of the foil which is opposite from the direction in which the recesses were formed.
• the tablets or capsules are sealed in the recesses between the plastic foil and the sheet.
• the strength of the sheet is such that the tablets or capsules can be removed from the blister pack by manually applying pressure on the recesses whereby an opening is formed in the sheet at the place of the recess. The tablet or capsule can then be removed via said opening.
• a memory aid on the kit, e.g., in the form of numbers next to the tablets or capsules whereby the numbers correspond with the days of the regimen which the dosage form so specified should be ingested.
• a memory aid is a calendar printed on the card e.g., as follows "First Week, Monday, Tuesday, ...etc.... Second Week, Monday, Tuesday, etc.
• a "daily dose” can be a single tablet or capsule or several tablets or capsules to be taken on a given day.
• a daily dose of a Formula I compound, a prodrug thereof or a pharmaceutically acceptable salt of said compound or said prodrug can consist of one tablet or capsule while a daily dose of the second compound can consist of several tablets or capsules and vice versa.
• the memory aid should reflect this.
• a dispenser designed to dispense the daily doses one at a time in the order of their intended use is provided.
• the dispenser is equipped with a memory-aid, so as to further facilitate compliance with the regimen.
• An example of such a memory-aid is a mechanical counter which indicates the number of daily doses that has been dispensed.
• a memory-aid is a battery-powered micro-chip memory coupled with a liquid crystal readout, or audible reminder signal which, for example, reads out the date that the last daily dose has been taken and/or reminds one when the next dose is to be taken.
• Preferred estrogen agonists / antagonists of the present invention include the compounds described in U.S. patent no. 5,552,412. Those compounds are described by the formula designated herein as formula (I) given below:
• A is selected from CH 2 and NR;
• B, D and E are independently selected from CH and N;
• a bicyclic ring system consisting of a five or six membered heterocyclic ring fused to a phenyl ring, said heterocyclic ring containing up to two heteroatoms selected from the group consisting of -O-, -NR 2 - and -S(O) n -, optionally substituted with 1-3 substituents independently selected from R 4 ;
• Z 1 is
• R 7 and R 8 in either linear or ring form may optionally be substituted with up to three substituents independently selected from C C 6 alkyl, halogen, alkoxy, hydroxy and carboxy; a ring formed by R 7 and R 8 may be optionally fused to a phenyl ring; e is 0, 1 or 2; m is 1 , 2 or 3; n is 0, 1 or 2; p is 0, 1 , 2 or 3; q is 0, 1 , 2 or 3; and optical and geometric isomers thereof; and nontoxic pharmaceutically acceptable acid addition salts, N-oxides, esters, quaternary ammonium salts and prodrugs thereof. Additional preferred estrogen agonists/antagonists are disclosed in U.S. patent no. 5,552,412 and are described by the formula designated herein as formula (IA): 04/0781
• R 4 is H, OH, F, or CI; and B and E are independently selected from CH and N, and optical and geometric isomers thereof; and nontoxic pharmaceutically acceptable acid addition salts, N-oxides, esters, quaternary ammonium salts and prodrugs thereof.
• Especially preferred estrogen agonists/antagonists for the methods of the invention are: cis-6-(4-fluoro-phenyl)-5-[4-(2-piperidin-1-yl-ethoxy)-phenyl]-5,6,7,8- tetrahydro-naphthalene-2-ol; (-)-cis-6-phenyl-5-[4-(2-pyrrolidin-1 -yl-ethoxy)-phenyl]-5,6,7,8-tetrahydro- naphthalene-2-ol; cis-6-phenyl-5-[4-(2-pyrrolidin-1-yl-ethoxy)-phenyl]-5,6,7,8-tetrahydro- naphthalene-2-ol; cis-1-[6'-pyrrolidinoethoxy-3'-pyridyl]-2-phenyl-6-hydroxy-1 ,2,3,4- tetrahydronaphthalene;
• An especially preferred salt of (-)-cis-6-phenyl-5-[4-(2-pyrrolidin-1-yl-ethoxy)- phenyl]-5,6,7,8-tetrahydro-naphthalene-2-ol is the D-tartrate salt.
• R 1A and R 2A may be the same or different and are either H, methyl, ethyl or a benzyl group; and optical or geometric isomers thereof; and pharmaceutically acceptable salts, N-oxides, esters, quaternary ammonium salts, and prodrugs thereof.
• Additional preferred estrogen agonists / antagonists are the compounds disclosed in U.S. Patent No. 4,536,516; 4-hydroxy tamoxifen ⁇ i.e., tamoxifen wherein the 2-phenyl moiety has a hydroxy group at the 4 position) and other compounds as disclosed in U.S. Patent No. 4,623,660; raloxifene: (methanone, [6-hydroxy-2-(4- hydroxyphenyl)benzo[b]thien-3-yl][4-[2-(1-piperidinyl)ethoxy]phenyl]-,hydrochloride) and other compounds as disclosed in U.S.
• centchroman 1-[2-[[4-(-methoxy-2,2, dimethyl-3- phenyl-chroman-4-yl)-phenoxy]-ethyl]-pyrrolidine and other compounds as disclosed in U.S. Patent No. 3,822,287; idoxifene: pyrrolidine, 1-[-[4-[[1-(4-iodophenyl)-2- phenyl-1-butenyl]phenoxy]ethyl] and other compounds as disclosed in U.S. Patent No.
• estrogen agonists / antagonists include EM-652 (as shown in the formula designated herein as formula (III) and EM-800 (as shown in the formula designated herein as formula (IV)).
• EM-652 as shown in the formula designated herein as formula (III)
• EM-800 as shown in the formula designated herein as formula (IV)
• the synthesis of EM-652 and EM-800 and the activity of various enantiomers is described in Gauthier et al., J. Med. Chem..
• estrogen agonists / antagonists include TSE 424 and other compounds disclosed in U.S. Patent No. 5,998,402, U.S. Patent No. 5,985,910, U.S. Patent No. 5,780,497, U.S. Patent No. 5,880,137, and European Patent Application EP 0802183 A1 including the compounds described by the formulae designated herein as formulae V and VI, below:
• R IB is selected from H, OH or the C C ⁇ 2 esters (straight chain or branched) or d-C 12 (straight chain or branched or cyclic) alkyl ethers thereof, or halogens; or d-d halogenated ethers including trifluoromethyl ether and trichloromethyl ether. R 2B .
• R 3B , R4B, RSB, and R 6B are independently selected from H, OH or the C r C 12 esters (straight chain or branched) or C C 12 alkyl ethers (straight chain or branched or cyclic) thereof, halogens, or C C halogenated ethers including trifluoromethyl ether and trichloromethyl ether, cyano, d-C 6 alkyl (straight chain or branched), or trifluoromethyl;
• X A is selected from H, C r C 6 alkyl, cyano, nitro, trifluoromethyl, and halogen; s is 2 or 3; Y A is selected from: a) the moiety:
• R 7B and R 8B are independently selected from the group of H, C r C 6 alkyl, or phenyl optionally substituted by CN, d-C 6 alkyl (straight chain or branched), C C 6 alkoxy (straight chain or branched), halogen, -OH, -CF 3 , or -OCF 3 ;
• a five-membered saturated, unsaturated or partially unsaturated heterocycle containing up to two heteroatoms selected from the group consisting of - O-, -NH-, -N(d-C 4 alkyl)-, -N , and -S(O) u -, wherein u is an integer of from 0-2, optionally substituted with 1-3 substituents independently selected from the group consisting of hydrogen, hydroxyl, halo, C d alkyl, trihalomethyl, C d alkoxy, trihalomethoxy, C r C 4 acyloxy, d-d alkylthio, C r C 4 alkylsulfinyl, C ⁇ -C 4 alkylsulfonyl, hydroxy (C C 4 )alkyl, -CO 2 H, -CN, -CONHR 1B , -NH 2 , C r C 4 alkylamino, di(C C 4 )alkylamino, -
• a six-membered saturated, unsaturated or partially unsaturated heterocycle containing up to two heteroatoms selected from the group consisting of -O-, -NH-, -N(d-C 4 alkyl)-, -N , and -S(O) u -, wherein u is an integer of from 0-2, optionally substituted with 1-3 substituents independently selected from the group consisting of hydrogen, hydroxyl, halo, d-d alkyl, trihalomethyl, d-d alkoxy, trihalomethoxy, C C 4 acyloxy, d-d alkylthio, C d alkylsulfinyl, d-d alkylsulfonyl, hydroxy (d-d)alkyl, -CO 2 H, -CN, -CONHRL -NH 2 , d-C 4 alkylamino, di(C C 4 )alkylamino, -NHSO 2 R 1B
• a seven-membered saturated, unsaturated or partially unsaturated heterocycle containing up to two heteroatoms selected from the group consisting of -O-, -NH-, -N(C C 4 alkyl)-, -N , and -S(O) u -, wherein u is an integer of from 0-2, optionally substituted with 1-3 substituents independently selected from the group consisting of hydrogen, hydroxyl, halo, C r C 4 alkyl, trihalomethyl, C C 4 alkoxy, trihalomethoxy, C C 4 acyloxy, C ⁇ -C 4 alkylthio, d-C 4 alkylsulfinyl, C d alkylsulfonyl, hydroxy (C C 4 )alkyl, -CO 2 H, -CN, -CONHR 1B , -NH 2 , C r C alkylamino, di(C ⁇ -C 4 )alkylamino, -NH
• Preferred compounds of this invention are those having the general structures V or VI, above, wherein:
• R 1B is selected from H, OH or the C C 12 esters or alkyl ethers thereof, and halogen;
• R 2B , R 3B , R B, R 5B , and R 6B are independently selected from H, OH or the C
• X A is selected from H, C C 6 alkyl, cyano, nitro, trifluoromethyl, and halogen;
• Y A is the moiety:
• R 7B and R 8B are selected independently from H, C C 6 alkyl, or combined by -(CH 2 ) W -, wherein w is an integer of from 2 to 6, so as to form a ring, the ring being optionally substituted by up to three substituents selected from the group of hydrogen, hydroxyl, halo, d-C 4 alkyl, trihalomethyl, C C 4 alkoxy, trihalomethoxy, d-d alkylthio, C C 4 alkylsulfinyl, C C 4 alkylsulfonyl, hydroxy (C C 4 )alkyl, -CO 2 H, -CN, -CONH(C C 4 alkyl), -NH 2 , d-C 4 alkylamino, d-C 4 dialkylamino, -NHSO 2 (C C 4 alkyl), -CO(CrC 4 alkyl), and -NO 2 ; and optical and geometric isomers thereof; and nontoxic
• the rings formed by a concatenated R B and R 8B may include, but are not limited to, aziridine, azetidine, pyrrolidine, piperidine, hexamethyleneamine or heptamethyleneamine rings.
• Preferred compounds of structural formulas V and VI, above, are those wherein R-
• R 7B and R 8B are concatenated together as -(CH 2 ) r , wherein t is an integer of from 4 to 6, to form a ring optionally substituted by up to three subsituents selected from the group of hydrogen, hydroxyl, halo, d-d alkyl, trihalomethyl, C C 4 alkoxy, trihalomethoxy, d-d alkylthio, C C 4 alkylsulfinyl, C C 4 alkylsulfonyl, hydroxy (C C 4 )alkyl, -CO 2 H, -CN, -CONH(C r C 4 )alkyl, -NH 2 , C C 4 alkylamino, di(C C 4 )alkylamino, -NHSO 2 (C C 4 )alkyl, -NHCO(C C 4 )alkyl, and -NO 2 ; and optical and geometric isomers thereof; and nontoxic pharmaceutically acceptable acid addition salt
• TSE-424 As described by the formula designated herein as formula (Va) below:
• Another estrogen agonist/antagonist that can be used in the combination aspect of the present invention is arzoxifene, which is disclosed in U.S. patent no. 5,723,474.
• a particularly preferred combination of an EP 2 selective receptor agonist and an estrogen agonist antagonist is (3-(((4-tert-butyl-benzyl)-(pyridine-3-sulfonyl)- amino)-methyl)-phenoxy)-acetic acid and (-)-c/ ' s-6-phenyl-5-(4-(2-pyrrolidin-1-yl- ethoxy)-phenyl)-5,6,7,8-tetrahydro-naphthalene-2-ol.
• the (3-(((4-tert-butyl-benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)- phenoxy)-acetic acid is in the form of the sodium salt
• the (-)-c/s-6-phenyl-5-(4-(2- pyrrolidin-1-yl-ethoxy)-phenyl)-5,6,7,8-tetrahydro-naphthalene-2-ol is in the form of the D-tartrate salt.
• Preferred bisphosphonates include, tiludronic acid, alendronic acid, zoledronic acid, ibandronic acid, risedronic acid, etidronic acid, clodronic acid, and pamidronic acid and their pharmaceutically acceptable salts or prodrugs or salts of the prodrugs.
• prodrugs and pharmaceutically acceptable salts may be formed from the compounds of this invention. All of such prodrugs and pharmaceutically acceptable salts so formed are within the scope of this invention.
• Particularly preferred salt forms of the estrogen agonists/antagonists include, raloxifene hydrochloride, tamoxifen citrate, toremifene citrate, and lasofoxifene tartrate.
• anti-resorptive agents such as progestins, polyphosphonates, bisphosphonate(s), estrogen agonists/antagonists, estrogen, estrogen/progestin combinations, Premarin ® (conjugated estrogens), estrone, estriol or 17 ⁇ - or 17 ⁇ -ethynyl estradiol may be used in conjunction with the EP 2 selective receptor agonists in the present methods.
• progestins are available from commercial sources and include: algestone acetophenide, altrenogest, amadinone acetate, anagestone acetate, chlormadinone acetate, cingestol, clogestone acetate, clomegestone acetate, delmadinone acetate, desogestrel, dimethisterone, dydrogesterone, ethynerone, ethynodiol diacetate, etonogestrel, flurogestone acetate, gestaclone, gestodene, gestonorone caproate, gestrinone, haloprogesterone, hydroxyprogesterone caproate, levonorgestrel, lynestrenol, medrogestone, medroxyprogesterone acetate, melengestrol acetate, methynodiol diacetate, norethindrone, norethindrone
• Preferred progestins are medroxyprogestrone, norethindrone and norethynodrel.
• Exemplary polyphosphonates include polyphosphonates of the type disclosed in U.S. Patent 3,683,080.
• Preferred polyphosphonates are geminal diphosphonates (also referred to as bis-phosphonates).
• Tiludronate disodium is an especially preferred polyphosphonate.
• Ibandronic acid is an especially preferred polyphosphonate.
• Alendronate is an especially preferred polyphosphonate.
• Zoledronic acid is an especially preferred polyphosphonate.
• Other preferred polyphosphonates are 6-amino-1-hydroxy-hexylidene-bisphosphonic acid and 1- hydroxy-3(methylpentylamino)-propylidene-bisphosphonic acid.
• the polyphosphonates may be administered in the form of the acid, or of a soluble alkali metal salt or alkaline earth metal salt. Hydrolyzable esters of the polyphosphonates are likewise included. Specific examples include ethane-1 -hydroxy 1 ,1 -diphosphonic acid, methane diphosphonic acid, pentane-1-hydroxy-1 ,1 -diphosphonic acid, methane dichloro diphosphonic acid, methane hydroxy diphosphonic acid, ethane-1 - amino-1 ,1 -diphosphonic acid, ethane-2-amino-1 ,1 -diphosphonic acid, propane-3- amino-1-hydroxy-1,1 -diphosphonic acid, propane-N,N-dimethyl-3-amino-1-hydroxy- 1 , 1 -diphosphonic acid, propane-3,3-dimethyl-3-amino-1 -hydroxy-1 , 1 -diphosphonic acid, phenyl amino methane diphosphonic acid,N
• prostaglandin refers to compounds that are analogs of the natural prostaglandins PGD ⁇ PGD 2 , PGE 2 , PGE., and PGF 2 . These compounds bind to the prostaglandin receptors. Such binding is readily determined by those skilled in the art of standard assays (e.g., An S. et al., Cloning and Expression of the EP 2 Subtype of Human Receptors for Prostaglandin E 2 , Biochemical and Biophvsical Research Communications. 1993, 197(1):263-270).
• Prostaglandins are alicyclic compounds related to the basic compound prostanoic acid.
• the carbon atoms of the basic prostaglandin are numbered sequentially from the carboxylic carbon atom through the cyclopentyl ring to the terminal carbon atom on the adjacent side chain. Normally the adjacent side chains are in the trans orientation.
• the presence of an oxo group at C-9 of the cyclopentyl moiety is indicative of a prostaglandin within the E class while PGE 2 contains a trans unsaturated double bond at the C ⁇ 3 -C ⁇ 4 and a cis double bond at the C 5 -C 6 position.
• a variety of prostaglandins are described and referenced below. However, other prostaglandins will be known to those skilled in the art. Exemplary prostaglandins are disclosed in U.S. patents 4,171 ,331 and 3,927,197.
• prostaglandin agonist/antagonist refers to compounds which bind to prostaglandin receptors ⁇ e.g., An S. et al., Cloning and Expression of the EP 2 Subtype of Human Receptors for Prostaglandin E 2 , Biochemical and Biophvsical Research Communications. 1993, 197(1):263-270) and mimic the action of prostaglandin in vivo. Such actions are readily determined by those skilled in the art of standard assays. Eriksen E.F. et al., Bone Histomorphometrv. Raven Press, New York, 1994, pages 1-74; Grier SJ. et.
• patent 3,932,389 discloses 2-descarboxy-2-(tetrazol-5-yl)-11-desoxy-15-substituted-omega- pentanorprostaglandins;
• U.S. patent 4,018,892 discloses 16-aryl-13,14-dihydro-PGE 2 p-biphenyl esters;
• U.S. patent 4,219,483 and 4,132,847 discloses 2,3,6-substituted-4- pyrones;
• U.S. patent 4,000,309 and 3,982,016 discloses 16-aryl-13,14-dihydro-PGE 2 p-biphenyl esters;
• U.S. patent 4,621 ,100 discloses substituted cyclopentanes; and
• U.S. patent 5,216,183 discloses cyclopentanones.
• sodium fluoride may be used as the second compound in certain aspects of this invention.
• the term sodium fluoride refers to sodium fluoride in all its forms (e.g., slow release sodium fluoride, sustained release sodium fluoride). Sustained release sodium fluoride is disclosed in U.S. patent 4,904,478.
• the activity of sodium fluoride is readily determined by those skilled in the art of biological protocols (e.g., see Eriksen E.F. et al., Bone Histomorphometrv, Raven Press, New York, 1994, pages 1- 74; Grier SJ. et. al., The Use of Dual-Energy X-Ray Absorptiometry In Animals, !Q ⁇ Radiol.. 1996, 31(1 ):50-62; Wahner H.W. and Fogelman I., The Evaluation of
• Osteoporosis Dual Energy X-Ray Absorptiometry in Clinical Practice., Martin Dunitz Ltd., London 1994, pages 1-296).
• parathyroid hormone refers to parathyroid hormone, fragments or metabolites thereof and structural analogs thereof which can stimulate bone formation and increase bone mass. Also included are parathyroid hormone related peptides and active fragments and analogs of parathyroid related peptides (see PCT publication no. WO 94/01460). A variety of these compounds are described and referenced below. However, other parathyroid hormones will be known to those skilled in the art. Exemplary parathyroid hormones are disclosed in the following references.
• growth hormone secretagogue refers to a compound which stimulates the release of growth hormone or mimics the action of growth hormone (e.g., increases bone formation leading to increased bone mass). Such actions are readily determined by those skilled in the art of standard assays well known to those of skill in the art. A variety of these compounds are disclosed in the following published PCT patent applications: WO 95/14666; WO 95/13069; WO 94/19367; WO 94/13696; and WO 95/34311. However, other growth hormones or growth hormone secretagogues will be known to those skilled in the art.
• a preferred growth hormone secretagogue is N-[1 (R)-[1 ,2- Dihydro-1-methanesulfonylspiro[3H-indole-3,4'-piperidin]-1'-yl)carbonyl]-2- (phenylmethyloxy)ethyl]-2-amino-2-methylpropanamide:MK-677.
• treating includes preventative (e.g., prophylactic), palliative and curative treatment.
• pharmaceutically acceptable it is meant the carrier, diluent, excipients, and/or salts or prodrugs must be compatible with the other ingredients of the formulation, and not deleterious to the recipient thereof.
• prodrug means a compound that is transformed in vivo to yield a compound of the present invention. The transformation may occur by various mechanisms, such as through hydrolysis in blood.
• T. Higuchi and W. Stella "Pro-drugs as Novel Delivery Systems," Vol. 14 of the A.C.S. Svmposium Series, and in Bioreversible Carriers in Drug Design, ed. Edward B. Roche, American Pharmaceutical Association and Pergamon Press, 1987.
• a prodrug can comprise an ester formed by the replacement of the hydrogen atom of the acid group with a group such as (d-
• C 8 )alkyl (C 2 -C 2 )alkanoyloxymethyl, 1-(alkanoyloxy)ethyl having from 4 to 9 carbon atoms, 1-methyl-1-(alkanoyloxy)-ethyl having from 5 to 10 carbon atoms, alkoxycarbonyloxymethyl having from 3 to 6 carbon atoms, 1- (alkoxycarbonyloxy)ethyl having from 4 to 7 carbon atoms, 1-methyl-1- (alkoxycarbonyloxy)ethyl having from 5 to 8 carbon atoms, N-
• a prodrug can be formed by the replacement of the hydrogen atom of the alcohol group with a group such as (C ⁇ -C 6 )alkanoyloxymethyl, 1-((C C 6 )alkanoyloxy)ethyl, 1-methyl-1-((d-C 6 )alkanoyloxy)ethyl, (C
• each ⁇ -aminoacyl group is independently selected from the naturally occurring L-amino acids, P(O)(OH) 2 , -P(O)(O(C C 6 )alkyl) 2 or glycosyl (the radical resulting from the removal of a hydroxyl group of the hemiacetal form of a carbohydrate).
• a prodrug can be formed by the replacement of a hydrogen atom in the amine group with a group such as R x -carbonyl, R x O-carbonyl, NR x R x '-carbonyl where R x and R x ' are each independently (d-C ⁇ o)alkyl, (C 3 -C 7 )cycloalkyl, benzyl, or R x -carbonyl is a natural ⁇ -aminoacyl or natural ⁇ -aminoacyl-natural ⁇ -aminoacyl, -C(OH)C(O)OY x wherein Y x is H, (C C 6 )alkyl or benzyl), -C(OY xo ) Y x1 wherein Y xo is (C C ) alkyl and Y X1 is (C C 6 )alkyl, carboxy(C 1 -C
• pharmaceutically acceptable salt refers to nontoxic anionic salts containing anions such as (but not limited to) chloride, bromide, iodide, sulfate, bisulfate, phosphate, acetate, maleate, fumarate, oxalate, lactate, tartrate, citrate, gluconate, methanesulfonate and 4-toluene-sulfonate.
• anions such as (but not limited to) chloride, bromide, iodide, sulfate, bisulfate, phosphate, acetate, maleate, fumarate, oxalate, lactate, tartrate, citrate, gluconate, methanesulfonate and 4-toluene-sulfonate.
• nontoxic cationic salts such as (but not limited to) sodium, potassium, calcium, magnesium, ammonium or protonated benzathine (N,N'-dibenzylethylenediamine), choline, ethanolamine, diethanolamine, ethylenediamine, meglamine (N-methyl- glucamine), benethamine (N-benzylphenethylamine), piperazine or tromethamine (2- amino-2-hydroxymethyl-1 ,3-propanediol).
• nontoxic cationic salts such as (but not limited to) sodium, potassium, calcium, magnesium, ammonium or protonated benzathine (N,N'-dibenzylethylenediamine), choline, ethanolamine, diethanolamine, ethylenediamine, meglamine (N-methyl- glucamine), benethamine (N-benzylphenethylamine), piperazine or tromethamine (2- amino-2-hydroxymethyl-1 ,3-propaned
• reaction inert solvent and “inert solvent” refers to a solvent that does not interact with starting materials, reagents, intermediates or products in a manner which adversely affects the yield of the desired product.
• the compounds of this invention can exist in radiolabelled form, i.e., said compounds may contain one or more atoms containing an atomic mass or mass number different from the atomic mass or mass number ordinarily found in nature.
• Radioisotopes of hydrogen, carbon, phosphorous, fluorine and chlorine include 3 H, 14 C, 32 P, 35 S, 18 F and 36 CI, respectively.
• Compounds of this invention which contain those radioisotopes and/or other radioisotopes of other atoms are within the scope of this invention. Tritiated, i.e., 3 H, and carbon-14, i.e., 14 C, radioisotopes are particularly preferred for their ease of preparation and detectability.
• Radiolabelled compounds of this invention can generally be prepared of methods well known to those skilled in the art. Conveniently, such radiolabelled compounds can be prepared by carrying out the procedures disclosed in the above Schemes and/or in the Examples and Preparations below by substituting a readily available radiolabelled reagent for a non-radiolabelled reagent.
• Diasteromeric mixtures can be separated into their individual diastereomers on the basis of their physicochemical differences by methods known perse as, for example, by chromatography and/or fractional crystallization.
• Enantiomers can be separated by converting the enantiomeric mixture into a diasteromeric mixture by reaction with an appropriate optically active compound (e.g., alcohol), separating the diastereomers and converting (e.g., hydrolyzing, including both chemical hydrolysis methods and microbial lipase hydrolysis methods, e.g., enzyme catalyzed hydrolysis) the individual diastereomers to the corresponding pure enantiomers. All such isomers, including diastereomers, enantiomers and mixtures thereof are considered as part of this invention. Also, some of the compounds of this invention are atropisomers (e.g., substituted biaryls) and are considered as part of this invention.
• an appropriate optically active compound e.g., alcohol
• converting e.g., hydrolyzing, including both chemical hydrolysis methods and microbial lipase hydrolysis methods, e.g., enzyme catalyzed hydrolysis
• the anti-resorptive agents including the compounds of Formula I, the anti-resorptive agents, bone anabolic agents, prostaglandin agonists/antagonists, parathyroid hormones, growth hormones and growth hormone secretagogues, form hydrates or solvates, they are also within the scope of the invention.
• Administration of the compounds of this invention can be via any method that delivers a compound of this invention systemically and/or locally. These methods include oral routes, parenteral, intraduodenal routes, etc. Generally, the compounds of this invention are administered orally, but parenteral administration (e.g., intravenous, intramuscular, transdermal, subcutaneous, rectal or intramedullary) may be utilized, for example, where oral administration is inappropriate for the target or where the patient is unable to ingest the drug.
• parenteral administration e.g., intravenous, intramuscular, transdermal, subcutaneous, rectal or intramedullary
• the compounds of this invention may also be applied locally to a site in or on a patient in a suitable carrier or diluent, optionally in combination with one or more of the anabolic agents or anti-resorptive agents described above.
• the amount and timing of compounds administered will, of course, be dependent on the subject being treated, on the severity of the affliction, on the manner of administration and on the judgment of the prescribing physician.
• the dosages given herein are guidelines and the physician may titrate doses ofthe drug to achieve the treatment that the physician considers appropriate for the patient.
• the physician must balance a variety of factors such age of the patient, presence of preexisting disease, as well as presence of other diseases (e.g., cardiovascular disease).
• an effective dosage for the anabolic agents used in this invention described above is in the range of 0.001 to 100 mg/kg/day, preferably 0.01 to 50 mg/kg/day.
• an effective dosage for an anti-resorptive agent is about 0.001 mg/kg/day to about 20 mg/kg/day.
• an effective dosage for progestins is about 0.1 to 10 mg per day; the preferred dose is about 0.25 to 5 mg per day.
• an effective dosage for polyphosphonates is determined by its potency as a bone resorption inhibiting agent of standard assays.
• Ranges for the daily administration of some polyphosphonates are about 0.001 mg/kg/day to about 20 mg/kg/day.
• an effective dosage for the treatment of this invention for example the bone resorption treatment of this invention, for the estrogen agonists/antagonists of this invention is in the range of 0.01 to 200 mg/kg/day, preferably 0.5 to 100 mg/kg/day.
• an effective dosage for raloxifene is in the range of 0.1 to 100 mg/kg/day, preferably 0.1 to 10 mg/kg/day.
• an effective dosage for tamoxifen is in the range of 0.1 to 100 mg/kg/day, preferably 0.1 to 5 mg/kg/day.
• an effective dosage for 2-(4-methoxy-phenyl)-3-[4-(2-piperidin-1- yl-ethoxy)-phenoxy]- benzo[b]thiophen-6-ol is 0.001 to 1 mg/kg/day.
• 1-(4'-pyrrolidinolethoxyphenyl)-2-phenyl-6-hydroxy-1 ,2,3,4- tetrahydroisoquinoline is in the range of 0.0001 to 100 mg/kg/day, preferably 0.001 to 10 mg/kg/day.
• the compounds of the present invention are generally administered in the form of a pharmaceutical composition comprising at least one of the compounds of this invention together with a pharmaceutically acceptable vehicle or diluent.
• the compounds of this invention can be administered individually or together in any conventional oral, parenteral, rectal or transdermal dosage form.
• a pharmaceutical composition can take the form of solutions, suspensions, tablets, pills, capsules, powders, and the like.
• Tablets containing various excipients such as sodium citrate, calcium carbonate and calcium phosphate are employed along with various disintegrants such as starch and preferably potato or tapioca starch and certain complex silicates, together with binding agents such as polyvinylpyrrolidone, sucrose, gelatin and acacia.
• binding agents such as polyvinylpyrrolidone, sucrose, gelatin and acacia.
• lubricating agents such as magnesium stearate, sodium lauryl sulfate and talc are often very useful for tabletting purposes.
• compositions of a similar type are also employed as fillers in soft and hard-filled gelatin capsules; preferred materials in this connection also include lactose or milk sugar as well as high molecular weight polyethylene glycols.
• preferred materials in this connection also include lactose or milk sugar as well as high molecular weight polyethylene glycols.
• lactose or milk sugar as well as high molecular weight polyethylene glycols.
• the compounds of this invention can be combined with various sweetening agents, flavoring agents, coloring agents, emulsifying agents and/or suspending agents, as well as such diluents as water, ethanol, propylene glycol, glycerin and various like combinations thereof.
• solutions in sesame or peanut oil or in aqueous propylene glycol can be employed, as well as sterile aqueous solutions of the corresponding water-soluble salts.
• aqueous solutions may be suitably buffered, if necessary, and the liquid diluent first rendered isotonic with sufficient saline or glucose.
• aqueous solutions are especially suitable for intravenous, intramuscular, subcutaneous and intraperitoneal injection purposes.
• the sterile aqueous media employed are all readily obtainable by standard techniques well-known to those skilled in the art.
• dilute sterile, aqueous or partially aqueous solutions are prepared.
• compositions of the invention may contain 0.1%-95% of the compound(s) of this invention, preferably 1%-70%.
• the composition or formulation to be administered will contain a quantity of a compound(s) of this invention in an amount effective to treat the disease/condition of the subject being treated.
• the present invention can also be administered using an injectable, flowable composition that provides sustained release at the local site of the injection by forming a biodegradable solid or gel depot, matrix or implant.
• an administration system is an EP 2 selective receptor agonist compound in a slow- release biodegradable polymer based delivery system.
• the polymer based delivery system contains EP 2 selective receptor agonist compound, and optionally any additional therapeutically useful compounds, dissolved or dispersed in biodegradable, thermoplastic polymer solution or dispersion in an organic solvent.
• the organic solvent diffuses away from the injection site, causing the polymer to precipitate or gel; thereby entrapping the compound in a sustained-release depot.
• the compound is subsequently released by diffusion from, and erosion of, the polymeric matrix.
• the polymeric matrix slowly erodes by hydrolysis and eventually disappears from the site of administration.
• the molecular weight and concentration of the polymer can control the in vivo release of the compound as well as the degradation rate of the matrix.
• the polymer based delivery system provides sustained release of an EP 2 selective receptor agonist compound in vivo for a sustained period of time with minimum or reduced burst in a patient in need thereof.
• a large burst of compound would result in poor local toleration due to local effects of the compound (e.g., irritation) and would minimize the amount of compound available for efficacy.
• the advantages this administration method is that it minimizes or reduces the initial burst, but still delivers compound at efficacious levels for a sustained period of time upon a single local injection.
• the polymer system is prepared by contacting the flowable composition with a gelation medium to coagulate or gel the composition into a solid, microporous polymeric matrix or a gel polymeric matrix.
• the flowable composition contains a thermoplastic polymer or copolymer in combination with a suitable solvent.
• the polymers or copolymers, which form the body of the matrix are substantially insoluble, preferably essentially completely insoluble, in water and body fluids. The insolubility of the matrix body enables it to function as a single site for the controlled release of the EP 2 selective receptor agonist compound.
• the polymers or copolymers also are biocompatible and biodegradable and/or bioerodible within the body of an animal, e.g., mammal.
• the biodegradation enables the patient to metabolize the polymer matrix so that it can be excreted by the patient without the need for further surgery to remove it. Because the flowable composition and polymer system are biocompatible, the insertion process and the presence of the polymer system within the body do not cause substantial tissue irritation or necrosis at the implant site.
• the composition of the present invention is administered as a flowable composition directly into body tissues.
• thermoplastic polymers for incorporation into the solid matrix of the controlled release polymer system are solids, pharmaceutically compatible and biodegradable by cellular action and/or by the action of body fluids.
• suitable thermoplastic polymers include polyesters of diols and dicarboxylic acids or of hydroxycarboxylic acids, such as polylactides, polyglycolides and copolymers thereof. More preferably the polymer is the copolymer, poly-lactic-co- glycolic acid (abbreviated PLGH), which upon hydrolysis, produces lactic and glycolic acid. The burst of release of this copolymer can be minimized further by the addition of polyethylene glycol (PEG) to form the PEG end-capped PLGH.
• PEG polyethylene glycol
• Preferred materials for use in the present invention are the polylactides, polyglycolides and copolymers thereof. These polymers can be used to advantage in the polymer system in part because they show excellent biocompatibility. They produce little, if any, tissue irritation, inflammation, necrosis or toxicity. In the presence of water, these polymers produce lactic and glycolic acid, respectively, which are readily metabolized by the body.
• the polylactides can also incorporate glycolide monomer to enhance the resulting polymer's degradation. These polymers can also be used because they effectively control the rate of release of the EP 2 selective receptor agonist compound from the polymer system, and because they result in the local retention of the EP 2 selective receptor agonist compound at the site of the site of administration.
• thermoplastic polymer in the organic solvent of the composition will vary according to factors such as crystallinity, hydrophilicity, capacity for hydrogen bonding and molecular weight of the polymer. Consequently, the molecular weight and the concentration of the polymer in the solvent are adjusted to achieve desired miscibility, as well as a desired release rate for the incorporated EP 2 selective receptor agonist compound.
• the flowable composition of thermoplastic polymer, solvent and the EP 2 selective receptor agonist compound is a stable flowable substance.
• a homogenous solution of the EP 2 selective receptor agonist compound in an organic solvent preferably results.
• the thermoplastic polymer is substantially soluble in the organic solvent.
• the solvent Upon placement of the flowable composition into the body, the solvent will dissipate and the polymer will solidify or gel to form the polymer system having the EP 2 selective receptor agonist compound within a solid or gel polymeric matrix. It has been discovered that the molecular weight of the polymer used distinctly affects the rate of release of the EP 2 selective receptor agonist compound and the rate of degradation of the polymer from the site as long as the flowable composition has been used as an intermediate.
• the molecular weight of the polymer or copolymer is adjusted to be within a range of about 0.2 to about 0.4 inherent viscosity (IN. in deciliters/g) for effective sustained release of the bone growth promoting compound.
• the typical rate of release of the incorporated bone growth promoting compound occurs at an IN. of about 0.2
• the molecular weight of the polymer can vary depending on the particular components of the composition. For most systems, it is preferred to adjust the molecular weight of the polymer to about 0.2 IN. for an effective sustained release of the EP 2 selective receptor agonist compound.
• the unit of measure for the molecular weight is daltons.
• the desired molecular weight range is about 0.2 to about 0.4 IN., with an IN. of about 0.2 being most preferred.
• the molecular weight of a polymer can be varied by any of a variety of methods. The choice of method is typically determined by the type of polymer composition. The preferred polymers for use are commercially available.
• thermoplastic polymers for use in the present invention are the following: PLGH copolymer with 1 :1 ratio of lactic and glycolic acid with an inherent viscosity of about 0.2 dl/g (commercially available from Boehringer Ingelheim as Copolymer RESOMER® RG 502 H) (about 12,000 molecular weight); PLGH copolymer with 1 :1 ratio of lactic and glycolic acid with an inherent viscosity of about 0.3 dl/g (commercially available from Boehringer Ingelheim as Copolymer RESOMER® RG 503 H)(about 37,000 molecular weight); PLGH copolymer with 1 :1 ratio of lactic and glycolic acid with an inherent viscosity of about 0.4 dl/g (commercially available from Boehringer Ingelheim as Copolymer RESOMER® RG 504 H) (about 47,000 molecular weight); and polyethylene glycol (PEG) end- capped PLGH copolymer with 1 :1 ratio of
• the rate and extent of release of the EP 2 selective receptor agonist compound from the polymer system can be varied from very fast to very slow.
• the release rate of (3-(((4-tert-butyl-benzyl)-(pyridine-3- sulfonyl)-amino)-methyl)-phenoxy)-acetic acid, sodium salt can be slowed to produce substantially complete release of the compound within about seven days.
• the period of time can be increased to about fourteen days.
• the desired release rate of the EP 2 selective receptor agonist compound will depend on several factors, such as the species of animal being treated as well as the specific condition being treated.
• the concentration of the polymer in the system can also be varied to adjust the release rate of the incorporated EP 2 selective receptor agonist compound. It has been discovered that the more dilute the polymer concentration, the more readily the EP 2 selective receptor agonist compound compound will be released. This effect can be used in combination with other methods to more effectively control the release of the incorporated EP 2 selective receptor agonist compound as desired. For example, by adjusting the concentration of the polymer and EP 2 selective receptor agonist compound, if desired, a wide range of release rates can be obtained
• the solvents used in the thermoplastic compositions of the present invention are preferably pharmaceutically acceptable, biocompatible and will dissipate into body fluid in situ such that they may be classed as having a solubility in water ranging from highly soluble to insoluble. Preferably, they cause relatively little, if any, tissue irritation or necrosis at the site of the injection and implantation.
• the solvent may have at least a minimal degree of water solubility. When the organic solvent is water insoluble or is minimally soluble in water, the solvent will slowly disperse from the flowable polymeric composition. The result will be an implant that during the course of its life may contain a varying amount of residual solvent.
• the organic solvent has a moderate to high degree of water solubility so that it will facilely disperse from the polymeric composition into the body fluids.
• the solvent disperses rapidly from the polymeric composition so as to quickly form a solid implant.
• the thermoplastic polymer coagulates or gels into the solid polymer system.
• the solvent dispersion causes pore formation within the polymer system.
• the flowable composition containing thermoplastic polymer, solvent and EP 2 selective receptor agonist compound will form a porous solid polymer system.
• the solvent is slightly water soluble or is water insoluble, the solvent dispersion may result in the formation of a solid porous implant, or if some solvent remains with the implant, the result may be formation of a gel implant having few or no pores.
• Suitable solvents include those liquid organic compounds meeting the foregoing criteria.
• the preferred solvent for use in the present invention is N-methyl-2- pyrrolidone (NMP) due, at least in part, to its solvating ability and its biocompatibility.
• the solvents for the thermoplastic polymer flowable compositions are chosen for compatibility and appropriate solubility of the polymer and solvent.
• Lower molecular weight thermoplastic polymers will normally dissolve more readily in the solvents than high molecular weight polymers.
• concentration of a thermoplastic polymer dissolved in the various solvents differs depending upon type of polymer and its molecular weight.
• the higher molecular weight thermoplastic polymers will tend to coagulate, gel or solidify faster than the very low molecular weight thermoplastic polymers.
• the higher molecular weight polymers tend to give higher solution viscosities than the low molecular weight materials.
• the molecular weight and the concentration of the polymer in the solvent are controlled.
• the EP 2 selective receptor agonist compound Upon formation of the polymer system from the flowable composition, the EP 2 selective receptor agonist compound becomes incorporated into the polymer matrix. After insertion of the flowable composition to form in situ the polymer system, the EP 2 selective receptor agonist compound will be released from the matrix into the adjacent tissues or fluids by diffusion and polymer degradation mechanisms. Manipulation of these mechanisms also can influence the release of the EP 2 selective receptor agonist compound into the surroundings at a controlled rate.
• the polymer matrix can be formulated to degrade after an effective and/or substantial amount of the EP 2 selective receptor agonist compound is released from the matrix.
• the release of the EP 2 selective receptor agonist compound from the matrix can be varied by, for example, the solubility of the EP 2 selective receptor agonist compound in water, the distribution of the bone growth promoting compound within the matrix, or the size, shape, porosity, solubility and biodegradability of the polymer matrix, among other factors.
• the release of the EP 2 selective receptor agonist compound from the matrix is controlled relative to its inherent rate by varying the polymer molecular weight to provide a desired duration and rate of release.
• a preferred dosage form of the EP 2 selective receptor agonist compound (3-(((4-tert-butyl-benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenoxy)- acetic acid, is a lyophile of the sodium salt to be reconstituted with a solution of PLGH in NMP before administration.
• the dosage form consisting of the lyophilized compound in one syringe (syringe A) and a solution of PLGH in NMP in a second syringe (syringe B), is known as the A/B reconstitution system.
• the contents of both syringes are mixed together immediately prior to dose delivery at or near site.
• the administered dosage forms will be a solution and will result in the dispersion of the compound with PLGH in NMP at desired strengths of, for example, 5 and 50 mgA/ml (mgA/ml refers to the free acid equivalent of the sodium salt form of the compound).
• the dosage form is a parenteral (e.g., subcutaneous, intramuscular or intramedullary) sustained release injection for local administration. This compound in a slow-release polymer matrix (depot injection) is designed for administration at or near a site, and is not intended for intravenous administration.
• a two-syringe system (A/B), as described above, may be used, preferably with the sodium salt form of the compound.
• a Uniphase formulation preferably with the free acid form of the compound, is a preferred alternative formulation.
• sterile filtration of the compound and irradiation of the polymer solution may be preferred for manufacturing a stable sterile product.
• the dosage form can be manufactured and shipped as separate aluminum pouches containing syringes filled with the lyophile form of the compound in one pouch and the polymer solution in the other pouch. Delivery containers, systems and methods for the lyophilization of the bone growth promoting compounds of the present invention to prepare pharmaceutical compositions and kits are described in published International patent application WO 01/73363. EXAMPLE A
• Drug Syringe A contained 4 mgA of the sodium salt lyophile in 1.25 ml male syringe without graduations; and Vehicle Syringe B contained 0.8 ml 50% RG502H/50%NMP solution in 1.25 ml female syringe without graduations.
• Drug Syringe A contained 40 mgA of the sodium salt lyophile in 1.25 ml male (fat) B-D syringe without graduations;
• Vehicle Syringe B contained 0.8 ml 50% RG502H/50% NMP solution in 1.25 ml female (thin) syringe without graduations.
• MgA refers to free acid equivalent of the sodium salt form of the compound
• RG502H is a PLGH copolymer with 1 :1 ratio of lactic and glycolic acid with inherent viscosity of 0.2 dl/gm, which is commercially available such as from Boehringer Ingelheim as Copolymer RESOMER® RG 502 H.
• EXAMPLE 1 50% RG502H/50% NMP with 5 mgA/ml of sodium salt of (3-(((4-tert-butyl- benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenoxy)-acetic acid, mixed A/B (polymer solution autoclaved, compound lyophilized)
• EXAMPLE 2 50% RG502H/50% NMP with 5 mgA/ml of sodium salt of (3-(((4-tert-butyl- benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenoxy)-acetic acid, mixed A/B (poly
• syringe A contains the lyophile of the sodium salt of (3-(((4-tert-butyl-benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)- phenoxy)-acetic acid which in made to result in 4 mg per syrinige or 40 mg per syringe.
• Syringe B contains Resomer 502H, 50:50 Poly(D,L lactide-co-glycolide), (50,50 PLGH) and N-methyl-2-pyrroliddone (NMP).
• a preffered dose range includes O. ⁇ mgA up to about 100mgA.
• preferred doses include 0.1ml, 0.2ml, 0.6ml and 2ml.
• preferred doses include 0.1ml, 0.2ml, 0,3ml, 0.6 and 2ml.
• EP 2 selective receptor agonist examples include local administration by injection to a particular site or delivery by a catheter to a site. Additional examples can be found in U.S. provisional patent application number 60/335,156, filed November 30, 2001.
• the compounds of this invention can be made by processes which include processes known in the chemical arts, particularly in light of the description contained herein. Certain processes for the manufacture of the compounds of this invention are provided as further features of the invention and are illustrated by the following reaction schemes. Other processes are described in the experimental section. Some substituents (e.g., carboxyl) may best be prepared through conversion of another functional group (e.g., carboxyl substituents may be prepared through conversion of, e.g., hydroxyl or carboxaldehyde) at a point later in the synthetic sequence.
• substituents e.g., carboxyl
• carboxyl substituents may best be prepared through conversion of another functional group (e.g., carboxyl substituents may be prepared through conversion of, e.g., hydroxyl or carboxaldehyde) at a point later in the synthetic sequence.
• Compounds of Formula I wherein B is nitrogen may be prepared using methods described in SCHEMES 1-5. These methods include (a) sequential alkylation of a sulfonamide or amide with two appropriate alkylating agents, generally alkyl halides or alkyl sulfonates; (b) alkylation of a sulfonamide or amide with an alkyl halide or alkyl sulfonate; or (c) reductive amination of an aldehyde followed by reaction with an acylating agent such as an acyl chloride, a chloroformate, an isocyanate or a chlorocarbonyl amide; or a sulfonylating agent such as a sulfonyl chloride.
• an acylating agent such as an acyl chloride, a chloroformate, an isocyanate or a chlorocarbonyl amide
• a sulfonylating agent such as a sul
• one of the alkylating agents When performing sequential alkylation, one of the alkylating agents will contain a Q-Z portion, where the Z portion is suitably protected if necessary, and the other alkylating agent will contain a K-M portion, where any functional groups requiring protection are suitably protected.
• the order of the alkylation i.e., whether the alkylating agent containing the Q-Z portion is added first or second, will depend upon the reactivity of the electrophilic side chain.
• the Q-Z portion When performing a reductive amination, the Q-Z portion may be attached to either the amine reagent or the aldehyde reagent depending upon the ease of preparation of the reagent and the 2004/078169
• the reductive amination is followed by acylation or sulfonylation with an appropriate acylating agent or sulfonyl chloride and, if desired the product is hydrolysed.
• the starting materials including amines, aldehydes and alkylating agents, are prepared using methods well known to those skilled in the art. Certain preferred methods for their preparation are described herein.
• compounds of Formula I wherein B is N are prepared ofthe methods set forth in SCHEMES 1 and 2 below.
• the sequences involve sequential alkylation of an appropriate sulfonamide of Formula 1 or amide of Formula 1 with two appropriate alkyl halides or alkyl sulfonates.
• SCHEMES 1 and 2 differ only in the order of addition of the two alkylating agents.
• the alkylation order is typically chosen depending on the reactivity of the electrophilic side-chain. It is generally preferable to react the less reactive electrophilic side chain first. This reduces the amount of dialkylation which occurs in that first alkylation step, thereby resulting in a greater yield of monoalkylated material to be carried on to the next alkylation.
• one of the alkylating agents contains a carboxylic acid or a carboxylic acid isostere, suitably protected with an appropriate protecting group, if necessary.
• the carboxylic acid precursor of Formula 3 is a carboxylic acid ester where R is a suitable carboxylic acid protecting group.
• the protecting group is either a straight chain lower alkyl, preferably methyl or ethyl, or a fert-butyl or phenyl group.
• Other acid isosteres can be employed by appropriately modifying SCHEMES 1 and 2 of methods well known to those skilled in the art (e.g., see SCHEME 6 which sets forth the preparation of a tetrazole).
• Typical alkylating agents are primary, secondary, benzylic or allylic halides and sulfonates and are preferably alkyl bromides or alkyl iodides.
• the Formula 1 sulfonamide or amide is converted to its anion with a strong base such as sodium hydride, lithium diisopropylamide, lithium bis(trimethylsilyl)amide, potassium bis(trimethylsily!amide, potassium tert-butoxide, etc. in an aprotic solvent such as dimethylformamide, tetrahydrofuran or N,N- dimethylformamide/benzene at a temperature of about -78°C to about 100°C.
• a strong base such as sodium hydride, lithium diisopropylamide, lithium bis(trimethylsilyl)amide, potassium bis(trimethylsily!amide, potassium tert-butoxide, etc.
• an aprotic solvent such as dimethylformamide, tetrahydrofuran or N,N- dimethylformamide/benzene at a temperature of about -78°C to about 100°C.
• the resulting anion is alkylated with an appropriate alkyl halide of Formula 2 or 3 or an appropriate alkyl sulfonate of Formula 2 or 3, wherein X' is the halide or sulfonate portion of the alkylating agent, at a temperature of about 0°C to about 100°C to yield the corresponding mono-alkylated compound of Formula 4 or 5.
• varying amounts of a side-product resulting from dialkylation of the amide or sulfonamide are obtained and can be removed using chromatographic techniques, preferably by flash chromatography (W.C. Still, M. Kahn, A. Mitra, J. Org. Chem. 43, 2923, 1978).
• the compound of Formula 4 or 5 is converted to an anion using a suitable base such as sodium hydride, lithium ! bis(trimethylsilyl)amide, lithium diisopropylamide, potassium bis(trimethylsilyl)amide, potassium tert-butoxide, or potassium carbonate in an aprotic solvent such as N,N- dimethylformamide, tetrahydrofuran, N,N- dimethylformamide/benzene, or acetone at a temperature of about -
• a suitable base such as sodium hydride, lithium ! bis(trimethylsilyl)amide, lithium diisopropylamide, potassium bis(trimethylsilyl)amide, potassium tert-butoxide, or potassium carbonate
• an aprotic solvent such as N,N- dimethylformamide, tetrahydrofuran, N,N- dimethylformamide/benzene, or acetone at a temperature of about -
• the hydrolysis may be carried out by using methods well known to those skilled in the art, for example, methods described in "Protecting Groups in Organic Synthesis," Second Edition, T.W. Greene and P.G.M. Wuts, John Wiley and Sons, Inc., 1991.
• Compounds of Formula I wherein B is N are also prepared from amines as set forth in SCHEMES 3-4.
• the appropriate amine starting materials of Formulas 9 and 10 are commercially obtained or can be prepared using methods well known to those skilled in the art (see “The Chemistry of Amino, Nitroso and Nitro Compounds and their Derivatives,” Ed. S. Patai, J. Wiley, New York, 1982).
• the amine starting materials are prepared from the corresponding nitriles of Formulas 7 or 8.
• nitriles are available from commercial sources or can be prepared using methods well known to those skilled in the art (see Rappaport, "The Chemistry of the Cyano Group,” Interscience, New York, 1970 or Patai and Rappaport, "The Chemistry of Functional Groups,” pt. 2, Wiley, New York, 1983).
• the nitrile of Formula 7 or 8 is reduced with a reducing agent such as borane- tetrahydrofuran complex, borane-methyl sulfide complex or lithium aluminum hydride in an aprotic solvent such as tetrahydrofuran or diethyl ether at a temperature of about -78°C to about 60°C.
• the nitrile is hydrogenated under a hydrogen atmosphere typically at 0 to 50 psi in the presence of Raney nickel or a platinum or palladium catalyst in a protic solvent such as methanol or ethanol at a temperature of about 0°C to about 50°C. It may be desired to add an equivalent of an acid, such as hydrogen chloride, to accomplish the reduction.
• the amine of Formula 9 or 10 thus obtained is converted to the sulfonamide of Formula 11 or 12 by sulfonylation with a sulfonyl chloride or said amine is converted to an amide of Formula 11 or 12 by acylation with an appropriate acyl chloride.
• Both the sulfonylation reactions and the acylation reactions are generally carried out in the presence of a weak base such as triethylamine, pyridine, or 4-methylmorpholine in an aprotic solvent such as methylene chloride or diethyl ether at a temperature of about - 20°C to about 50°C.
• a weak base such as triethylamine, pyridine, or 4-methylmorpholine
• an aprotic solvent such as methylene chloride or diethyl ether
• coupling of amines of Formulas 9 or 10 with carboxylic acids are conveniently carried out in an inert solvent such as dichloromethane or N,N-dimethylformamide by a coupling reagent such as 1-(3- dimethylaminopropyl )-3-ethylcarbodiimide hydrochloride (EDC) or 1 , 3- dicyclohexylcarbodiimide (DCC) in the presence of 1-hydroxybenzotriazole hydrate (HOBT) to generate compounds of Formulas 11 or 12.
• a suitable base such as triethylamine
• the coupling can be effected with a coupling reagent such as benzotriazol-l-yloxy-tris(dimethylamino)- phosphonium hexafluorophosphate (BOP) in an inert solvent such as methanol.
• a coupling reagent such as benzotriazol-l-yloxy-tris(dimethylamino)- phosphonium hexafluorophosphate (BOP) in an inert solvent such as methanol.
• BOP benzotriazol-l-yloxy-tris(dimethylamino)- phosphonium hexafluorophosphate
• inert solvent such as methanol
• the amines of Formulas 9 and 10 are also prepared via reduction of an appropriate amide of Formulas 15 and 16. This reduction is achieved using reagents such as a borane-tetrahydrofuran complex, a borane-methyl sulfide complex, or diisobutyaluminum hydride in an aprotic solvent such as tetrahydrofuran or diethyl ether at a temperature of about -78°C to about 60°C.
• reagents such as a borane-tetrahydrofuran complex, a borane-methyl sulfide complex, or diisobutyaluminum hydride in an aprotic solvent such as tetrahydrofuran or diethyl ether at a temperature of about -78°C to about 60°C.
• the amines of Formulas 9 and 10 are also obtained from the corresponding nitro precursors by reduction of the nitro group using reducing reagents such as zinc/HCI, hydrogenation in the presence of Raney nickel, palladium, or platinum catalysts, and other reagents as described by P.N. Rylander in "Hydrogenation Methods,” Academic Press, New York, 1985. SCHEME 3
• the compounds of Formula I wherein B is N are prepared by reductive amination of an aldehyde containing the appropriate suitably protected acidic functionality with an amine. This sequence is set forth in SCHEME 5.
• the amine may contain the appropriate suitably protected acidic functionality.
• the reductive amination is typically carried out at a pH of between 6 and 8, using a reducing agent such as sodium cyanoborohydride or sodium triacetoxyborohydride.
• a reducing agent such as sodium cyanoborohydride or sodium triacetoxyborohydride.
• the reaction is normally performed in a protic solvent such as methanol or ethanol at temperatures of about -78°C to about 40°C. (e.g., see A. Abdel-Magid, C. Maryanoff, K. Carson, Tetrahedron Lett. 39, 31, 5595-5598, 1990.)
• the reductive amination reaction may also be carried out using titanium isopropoxide and sodium cyanoborohydride (RJ. Mattson et al, J. Org. Chem.
• amine of Formulas 42 and 42A is transformed to the desired amide or sulfonamide by coupling with an acid chloride, sulfonyl chloride, or carboxylic acid as set forth in SCHEMES 3 and 4.
• the amine intermediate of Formulas 42 or 42A may be converted to a urethane by treatment with a chloroformate or to a tetrasubstituted urea by treatment with a chlorocarbonyl amide.
• a sulfonamide or amide of Formula 4 is alkylated with the appropriate alkyl halide or sulfonate (wherein X' is halide or sulfonate), preferably a primary, secondary, benzylic, or allylic alkyl bromide, iodide, or sulfonate, which contains a nitrile to provide a nitrile of Formula 59.
• This alkylation is achieved by treatment of the sulfonamide or amide of Formula 59 with a base such as sodium hydride, lithium bis(trimethylsilyl)amide, potassium bis(trimethy!silyl)amide, potassium tert-butoxide, or potassium carbonate in an aprotic solvent such as dimethylformamide, dimethylformamide/benzene, or acetone followed by reaction of the resulting anion with a suitable alkylating agent. Alkylation occurs at a temperature of about -78°C to about 100°C.
• a base such as sodium hydride, lithium bis(trimethylsilyl)amide, potassium bis(trimethy!silyl)amide, potassium tert-butoxide, or potassium carbonate
• an aprotic solvent such as dimethylformamide, dimethylformamide/benzene, or acetone
• a preferred method for converting the resulting nitrile of Formula 59 to the tetrazole of Formula 60 is treatment of the alkylated nitrile with dibutyltin oxide and trimethylsilylazide, in refluxing toluene (S J. Wittenberger and B.G. Donner, J. Org. Chem. 1993, 58, 4139-4141, 1993).
• S J. Wittenberger and B.G. Donner, J. Org. Chem. 1993, 58, 4139-4141, 1993 For a review of alternative preparations of tetrazoles see R.N. Butler, Tetrazoles, In Comprehensive Heterocyclic Chemistry; Potts, K.T. Ed.; Pergamon Press: Oxford, 1984, Vol. 5, pp 791-838.
• esters of Formula 46 are prepared using the procedures described above in SCHEMES 1 and 2.
• Heck coupling of this intermediate to an arylhalide preferably an aryl bromide or aryl iodide
• an aryl triflate or a ring system which contains a vinyl bromide, iodide, or triflate is accomplished with a palladium catalyst, such as palladium acetate or tetrakis(triphenylphosphine)palladium(0) in the presence of a trialkylamine, such as triethylamine.
• an additive such as a triarylphosphine or triarylarsene may be added to the reaction.
• the reaction is typically performed in an aprotic solvent such as dimethylformamide or acetonitrile at a temperature of about 0°C to about 150°C (see R.F. Heck in Comp. Org. Syn., Vol. 4, Ch. 4.3, p. 833 or Daves and Hallberg, Chem. Rev. 1989, 89, 1433).
• the compound of Formula 47 can be hydrolyzed to the corresponding acid.
• the compound of Formula 47 can be hydrogenated and, if desired, further hydrolyzed to the corresponding acid of Formula 49.
• Hydrogenation is preferably achieved under a hydrogen atmosphere typically at 0 to 50 psi in the presence of a palladium or platinum catalyst in an alcoholic solvent such as ethanol or methanol at a temperature of about 0°C to about 50°C.
• a palladium or platinum catalyst in an alcoholic solvent such as ethanol or methanol at a temperature of about 0°C to about 50°C.
• hydrogenation will generate a fully saturated ring system.
• certain compounds of Formula I wherein B is N are prepared as described in SCHEME 8.
• Compounds of Formula 51 are prepared as described in SCHEMES 1 and 2 by alkylation of compounds of Formula 5 with an electrophile of Formula 2 which contains the appropriate functionality on the M ring. At least one of the substituents on the M ring must be suitable for subsequent conversion to an aldehyde.
• electrophiles of Formula 2 containing a protected alcohol on the M ring may be alkylated and then deprotected and oxidized to the aldehyde, using reagents well known to those skilled in the art, to generate compounds of Formula 51.
• An alternative method is to alkylate with an electrophile of Formula 2 where M contains a vinyl group.
• oxidative cleavage of the double bond provides the desired aldehyde of Formula 51.
• the oxidative cleavage is accomplished by transforming the double bond to the 1 ,2-diol with catalytic osmium tetroxide and N-methylmorpholine followed by oxidative cleavage to the aldehyde using sodium periodate.
• oxidative cleavage via ozonolysis followed by reduction using reagents such as methyl sulfide, triphenylphosphine, zinc/acetic acid, or thiourea, generates the desired aldehyde of Formula 51.
• LMetal is any organometallic reagent such as an organolithium or a Grignard reagent in an aprotic solvent such as diethyl ether or tetrahydrofuran at a temperature of about -78°C to about 80°C, followed by hydrolysis of the ester as described above, provides the desired compound of Formula 50.
• organometallic reagent such as an organolithium or a Grignard reagent in an aprotic solvent such as diethyl ether or tetrahydrofuran at a temperature of about -78°C to about 80°C
• certain compounds of Formula I wherein B is N are prepared as described in SCHEME 9.
• the appropriate sulfonamide or amide of Formula 5 is alkylated using the conditions described in SCHEMES 1 and 2.
• the alkylating agent is an electrophile which contains an aromatic bromide or iodide or a ring system which contains a vinyl bromide or iodide (Ar 1 ) to provide compounds of Formula 53.
• Suzuki-type coupling of the compound of Formula 53 thus obtained with an aryl boronic acid (Ar 2 ) provides Formula 53a compounds.
• the coupling reaction is achieved using about two equivalents of a base, such as sodium carbonate, potassium carbonate, sodium hydroxide, thallium hydroxide or potassium phosphate, in the presence of a palladium catalyst, such as tetrakis(triphenylphosphine)palladium(0), palladium acetate, palladium chloride, tris(dibenzylideneacetone)dipalladium(0) or [1,4- bis(diphenylphosphine)butane]palladium(0).
• a base such as sodium carbonate, potassium carbonate, sodium hydroxide, thallium hydroxide or potassium phosphate
• a palladium catalyst such as tetrakis(triphenylphosphine)palladium(0), palladium acetate, palladium chloride, tris(dibenzylideneacetone)dipalladium(0) or [1,4- bis(diphenylphosphine)butane]palladium(0).
• the reaction may be run in an aqueous alcoholic solvent such as methanol or ethanol; or in other aqueous solvents such as aqueous tetrahydrofuran, aqueous acetone, aqueous glycol dimethyl ether, or aqueous benzene at temperatures ranging from about 0°C to about 120°C.
• aqueous alcoholic solvent such as methanol or ethanol
• other aqueous solvents such as aqueous tetrahydrofuran, aqueous acetone, aqueous glycol dimethyl ether, or aqueous benzene
• Ar 1 is a partially saturated ring
• reduction of the ring to provide a saturated ring system may, if desired, be performed at this point.
• This transformation is achieved by hydrogenating the partially saturated ring in the presence of a catalyst such as palladium or platinum in an alcoholic solvent (ethanol or methanol) and/or ethyl acetate.
• Ester hydrolysis of compounds of Formulas 53 or 53a provides the corresponding acid.
• the resulting acids may contain functional groups on either of the ring systems (Ar 1 or Ar 2 ) which can be modified using methods well known to those skilled in the art. Examples of such modifications are shown in SCHEME 10.
• certain compounds of Formula I wherein B is N are prepared as described in SCHEME 11.
• the starting alcohols of Formula 58 are prepared of methods well known to persons skilled in the art, for example, by using methods described in SCHEMES 1 and 2. It will be recognized by a person of ordinary skill in the art that protecting groups may be required in the synthesis of certain of these alcohols.
• Intermediate 58 is coupled with a variety of aryl alcohols (M is as defined above) using Mitsonobu coupling conditions (for a review see O. Mitsonobu, Synthesis, 1 , 1981 ).
• the coupling is achieved by addition of a coupling agent such as triphenylphosphine and diethyl azodicarboxylate or diisopropyl azodicarboxylate in an inert solvent such as methylene chloride or tetrahydrofuran at a temperature of about 0°C to about 80°C. If desired, subsequent hydrolysis yields the corresponding acid.
• a coupling agent such as triphenylphosphine and diethyl azodicarboxylate or diisopropyl azodicarboxylate
• an inert solvent such as methylene chloride or tetrahydrofuran
• certain compounds of Formula I wherein B is N are prepared as described in SCHEME 12.
• a compound of Formula 102 is added to a compound of Formula 105 (wherein X is as defined above for the compound of Formula I) in the presence of a Lewis acid such as titanium tetrachloride or a mineral acid such as hydrochloric acid.
• a Lewis acid such as titanium tetrachloride or a mineral acid such as hydrochloric acid.
• the ester of Formula 106 can be converted to the corresponding acid by hydrolysis or deprotection.
• chloromethyl compounds of Formula 104 can be treated with a Lewis acid such as titanium tetrachloride and an appropriately substituted vinyl silane in an aprotic solvent such as methylene chloride at a temperature of about -50°C to about 50°C to give compounds of Formula 108.
• a Lewis acid such as titanium tetrachloride and an appropriately substituted vinyl silane in an aprotic solvent such as methylene chloride at a temperature of about -50°C to about 50°C
• the compounds of Formula 108 may subsequently be hydrolyzed or deprotected as described above to yield the corresponding acid.
• reduction of the double bond can be accomplished using conditions described in SCHEME 7.
• Formula I compounds wherein B is C(H) and Q, G, M and K are as described above in the Summary of the Invention can be prepared of SCHEME 16.
• Formula 113 beta-ketoesters are alkylated sequentially with Formula 114 compounds to form Formula 115 compounds followed by alkylation with Formula 116 compounds to give Formula 117 compounds (J. Med. Chem. 26, 1993, p335-41).
• Alkylations can be carried out in a suitable solvent such as DMF, THF, ether, or benzene using an appropriate base such as sodium hydride, LDA, or potassium carbonate at a temperature of about -78°C to about 80°C.
• the resulting Formula 117 disubstituted keto esters are hydrolyzed and decarboxylated to give the corresponding Formula 118 compound by using an aqueous base such as sodium hydroxide to hydrolyze the ester, followed by an acidic quench such as with aqueous hydrochloric acid to effect decarboxylation.
• an aqueous base such as sodium hydroxide to hydrolyze the ester
• an acidic quench such as with aqueous hydrochloric acid to effect decarboxylation.
• Formula I compounds wherein B is C(H) and G, Q, M and K are as described above in the Summary of the Invention may be prepared of SCHEME 17.
• Sequential alkylation of a malonate derivative of Formula 119 provides the Formula 121 dialkylated compound.
• Deprotection of the ester group by treatment with a strong acid such as TFA or HCI in ethanol at a temperature of about -20°C to about 50°C leads to the Formula 122 decarboxylated product.
• Formula 123 compounds are suitable substrates for addition of various organometallic species, e.g., Grignard reagents and organo- cadmium reagents which, after hydrolysis of the terminal ester, provide the keto-acid compounds of Formula 118.
• organometallic species e.g., Grignard reagents and organo- cadmium reagents which, after hydrolysis of the terminal ester, provide the keto-acid compounds of Formula 118.
• Formula 118 compounds can be prepared using methods described previously in Schemes 7-11 where one or both of the side chains are further functionalized after attachment to the alkanoyl fragment.
• Certain amides or sulfonamides described by Formulas 21 , 22, and 23 wherein W and Z are as described above in the Summary of the Invention and X and M are aromatic or saturated ring systems are prepared as set forth in SCHEME 18.
• Alkynyl amides or sulfonamides of Formulas 25, 26 and 27 are prepared by coupling an alkynyl sulfonamide or alkynyl amide of Formula 24 to an aromatic or vinyl halide, preferably an aromatic or vinyl bromide or iodide wherein W and Z are as defined above and where X and M represent an aromatic ring or a partially saturated ring system.
• the coupling is typically accomplished in the presence of copper iodide, a palladium catalyst, such as palladium chloride, bis(triphenylphosphine)palladium dichloride, or te.rakis(triphenylphosphine)palladium(0), and an amine such as triethylamine, diisopropylamine, or butylamine in an aprotic solvent such as acetonitrile at a temperature of about 0°C to about 100°C.
• a palladium catalyst such as palladium chloride, bis(triphenylphosphine)palladium dichloride, or te.rakis(triphenylphosphine)palladium(0)
• an amine such as triethylamine, diisopropylamine, or butylamine
• an aprotic solvent such as acetonitrile
• the alkynes of Formulas 25, 26 and 27 are converted to the corresponding alkanes of Formulas 21 , 22 or 23 via hydrogenation in the presence of a palladium or platinum catalyst in a solvent such as methanol, ethanol, and/or ethyl acetate at a temperature of about 0°C to about 50°C.
• a palladium or platinum catalyst in a solvent such as methanol, ethanol, and/or ethyl acetate at a temperature of about 0°C to about 50°C.
• M represents a partially saturated ring system
• hydrogenation will convert M to a fully saturated ring system.
• the alkynes are converted to cis-alkenes using the Lindlar catalyst (Pd-CaC0 3 -PbO) or other suitable catalyst. Alkylation and deprotection as described in SCHEMES 1 and 2 affords the corresponding compounds of Formula I.
• Compounds of Formula 33 are prepared from a suitable amine of Formula 32 (e.g., methoxyarylalkylamine).
• Amines of Formula 32 are commercially available or are prepared by methods well known to those skilled in the art (for example, see SCHEME 4).
• Amines of Formula 32 are converted to sulfonamides or amides of Formula 31 using methods, for example, described in SCHEME 3 and 4.
• the resulting aromatic methyl ether of Formula 31 is deprotected with reagents such as boron tribromide, pyridinium hydrochloride, hydrogen bromide/acetic acid, or other reagents as described in Protecting Groups in Organic Synthesis, Second Edition, T.W. Greene and P.G.M.
• N- bromosuccinimide N- bromosuccinimide
• AIBN 2,2'- azobisisobutyronitrile
• a peroxide preferably benzoyl peroxide.
• the reaction can be initiated with light.
• the reaction is performed in an inert solvent such as carbon tetrachloride or chloroform at a temperature of about 50°C to about 100°C.
• SCHEME 21 sets forth the synthesis of alkylating agents useful for preparing compounds of Formula I where M represents a biaryl or aryl cyclic group.
• Suzuki-type coupling of an aryl iodide or bromide or a ring system containing a vinyl bromide or iodide (Ar 2 ) with a methylaryl boronic acid (Ar 1 ) using the conditions described in SCHEME 9 provides compounds of Formula 34. Where a vinyl bromide or iodide is used, compounds of Formula 34 can be reduced to generate a fully saturated ring.
• R H, alkyl
• Alcohols are obtained from commercial sources or can be prepared using methods well known to those skilled in the art.
• SCHEME 22 sets forth the reduction of a carboxylic acid or ester to the corresponding alcohol using reagents such as, but not limited to, sodium borohydride, lithium aluminum hydride, borane-tetrahydrofuran complex, borane-methyl sulfide complex, etc.
• the corresponding alkyl chlorides are typically prepared from the alcohols with reagents such as hydrogen chloride, thionyl chloride, phosphorous pentachloride, phosphorous oxychloride, or triphenylphosphine/carbon tetrachloride.
• reagents such as hydrogen chloride, thionyl chloride, phosphorous pentachloride, phosphorous oxychloride, or triphenylphosphine/carbon tetrachloride.
• the alcohol is commonly treated with reagents such as hydrogen bromide, phosphorous tribromide, triphenylphosphine/bromine, or carbonyldiimidazole/allyl bromide (Kamijo, T., Harada, H., lizuka, K. Chem. Pharm. Bull. 1983, 38, 4189).
• alkyl chlorides can be converted to the more reactive alkyl bromides or alkyl iodides by reaction with an inorganic salt such as sodium bromide, lithium bromide, sodium iodide, or potassium iodide in solvents such as acetone or methyl ethyl ketone.
• Alkyl sulfonates can also be used as electrophiles or can be converted to alkyl halides.
• Alkyl sulfonates are prepared from the corresponding alcohol using a mild base such as triethylamine or pyridine and a sulfonyl chloride in an inert solvent such as methylene chloride or diethyl ether. If desired, conversion to the halide is accomplished by treating the alkyl sulfonate with an inorganic halide (sodium iodide, sodium bromide, potassium iodide, potassium bromide, lithium chloride, lithium bromide, etc) or a tetrabutylammonium halide.
• a mild base such as triethylamine or pyridine
• a sulfonyl chloride in an inert solvent such as methylene chloride or diethyl ether.
• conversion to the halide is accomplished by treating the alkyl sulfonate with an inorganic halide (sodium iodide, sodium bromide, potassium iod
• Cinnamic acids or esters are commonly available from commercial sources and can by converted to alkylating agents of Formulas 37 or 38 as follows (see SCHEME 23).
• the cinnamic acid or ester derivatives are reduced by hydrogenation in the presence of palladium or platinum catalysts typically in protic solvents (e.g., methanol or ethanol), tetrahydrofuran, or ethyl acetate.
• protic solvents e.g., methanol or ethanol
• Reduction and conversion to the alkyl halide or sulfonate as described in SCHEME 22 provides the compounds of Formula 38.
• the cinnamic acids or esters are converted directly to the alcohols of Formula 39 by treat those cinnamic acids or esters with reagents such as lithium aluminum hydride in an inert solvent such as tetrahydrofuran and diethyl ether.
• reagents such as lithium aluminum hydride in an inert solvent such as tetrahydrofuran and diethyl ether.
• the cinnamic acid or ester can be reduced to an allylic alcohol of Formula 40 using reagents such as lithium aluminum hydride/aluminum chloride, diisobutylaluminum hydride or lithium borohydride. Conversion to the allylic halide or sulfonate as described in SCHEME 22 provides the compounds of Formula 37.
• SCHEME 24 Conversion to the allylic halide or sulfonate as described in SCHEME 22 provides the compounds of Formula 37.
• alkylating agents of Formula 41 wherein W and M are as described above in the Summary of the Invention is set forth in SCHEME 24.
• Compounds of Formula 42 can be alkylated with a variety of bases. The choice of base is dependent on the nature of W and M. Some preferred bases include, but are not limited to, sodium hydroxide, sodium hydride, lithium diisopropylamide, lithium bis(trimethylsilyl)amide, potassium bis(trimethylsilyl)amide and potassium tert- butoxide. Treatment of the resulting anion with one of a variety of dialkylhalides generates the desired alkylating agents of Formula 41.
• Aldehydes useful for the method described in SCHEME 5 are available from commercial sources or can be prepared from available intermediates using methods well known to those skilled in the art (for a general reference see "The Chemistry of the Carbonyl Group,” Ed. S. Patai, Interscience, New York (1966-70)).
• SCHEME 25 demonstrates an exemplary method used to prepare Formula 43 hydroxy aldehydes where M in SCHEME 5 contains a hydroxy substituted alkyl group.
• Aldehydes useful for the method described in SCHEME 5 may be prepared using the methods described in SCHEMES 26-28.
• an aryl boronic acid which contains an aldehyde can be coupled to an aryl bromide, aryl iodide, or a ring system which contains a vinyl bromide or iodide using the Suzuki protocol described for SCHEME 9 to provide aldehydes of Formula 60.
• SCHEME 27 describes the preparation of aldehydes of Formula 62 which contain a suitably protected acid moiety and can be used for the preparation of compounds of Formula 42A described in SCHEME 5.
• Selective reduction of nitriles (see SCHEMES 3-4 for preparations) of Formula 61 provides aldehydes of Formula 62.
• a preferred method for this reduction involves heating the nitrile with aluminum- nickel (Raney) alloy in the presence of an acid such as formic acid.
• Aldehydes of Formula 64 useful for the preparation of compounds of Formula 42 may be prepared from starting nitriles of Formula 63 by treatment with a variety of reducing agents such as diisobutylaluminum hydride, tin (II) chloride/hydrogen chloride, or lithium triethoxyalanate.
• reducing agents such as diisobutylaluminum hydride, tin (II) chloride/hydrogen chloride, or lithium triethoxyalanate.
• a method for the preparation of proprionaldehydes of Formula 65 is described in SCHEME 28 and follows the procedures described by Kang (J. Org. Chem. 1996, 61 , 2604) and by Jeffery (J. Chem. Soc. Chem. Commun. 1984, 19, 1287).
• An aryl iodide or bromide is coupled to allyl alcohol in the presence of a suitable palladium catalyst, preferably palladium acetate.
• the reaction is performed in a suitable polar, aprotic solvent, preferably dimethylformamide, with addition of a base, such as sodium bicarbonate, and an ammonium salt, such as tetrabutylammonium chloride and provides proprionaldehydes of.
• Formula 65 A method for the preparation of proprionaldehydes of Formula 65.
• Intermediate chloromethyl compounds can be prepared as described in SCHEMES 29 and 30.
• the appropriate Formula 66 or 68 sulfonamide or carboxamide is treated with a formaldehyde equivalent such as paraformaldehyde in an inert organic solvent such as methylene chloride or chloroform with a suitable catalyst such as HCI, zinc chloride or trimethylsilyl chloride at temperatures ranging from about 0°C to about 60°C to give the Formula 67 and 69 chloromethyl derivatives, respectively. 04/078169
• SCHEME 31 sets forth the synthesis of biaryl aldehydes of Formula 60.
• Fluorobenzonitriles of Formula 70 are heated with a nucleophilic group such as a pyrrazole or imidazole in a suitable solvent, preferably DMF to effect displacement of the fluoro group yielding intermediates of Formula 71.
• the desired biaryl aldehydes of Formula 60 are obtained by reduction of the nitrile via hydrogenation with Raney alloy in formic acid, or by reduction with a hydride source such as diisobutylaluminum hydride.
• Example 1 7-((4-Butyl-benzyl.-(pyridine-3-sulfonyl)-amino)-heptanoic acid Step A: Reductive Amination 7-(4-Butyl-benzylamino)-heptanoic acid methyl ester.
• Step A After cooling to 0°C, NaBH (0.342 g, 9.04 mmol) was added and the reaction was stirred for 15 minutes at room temperature. The mixture was quenched with 1 :1 NaHC0 3 :H 2 0 and the MeOH was removed in vacuo. The resulting residue was diluted with CH 2 CI 2 and the organic solution was washed with water and brine, dried over MgS0 4 , filtered and concentrated in vacuo to afford the title compound of Step A (1.4 g).
• Step C Ester Hydrolysis 7-((4-Butyl-benzyl)-(pyridine-3-sulfonyl)-amino)-heptanoic acid.
• Examples 1a-1an were prepared from the appropriate starting materials in a manner analogous to the method of Example 1 , with variations in reaction time, temperature, and reagents as noted.
• Example 1b (3-(((1-Methyl-1 H-indol-3-ylmethyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)- acetic acid 1 H NMR (400 MHz, (CDpl 3 ) ⁇ 8.93 (s, 1 H), 8.66 (s, 1H), 7.96 (d, 1H), 7.39 (d, 1 H), 7.01-7.37 (m, 9H), 6.77 (s, 1 H), 4.56 (s, 2H), 4.41 (s, 2H), 3.66 (s, 3H), 3.52 (s, 2H); MS 448 (M-1).
• Example 1c (3-(((1-Methyl-1 H-indol-3-ylmethyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)- acetic acid 1 H NMR (400 MHz, (CDpl 3 ) ⁇ 8.93 (s, 1 H), 8.66
• Example 1e 3-(((4-Phenethylsulfanyl-benzv ⁇ -(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)-acetic acid Step A: Reaction time of 4 h at room temperature.
• 1 H NMR 400 MHz, CDCI 3 ) ⁇ 8.00 (d, 1H), 7.50 (bs, 1H), 6.90-7.38 (m, 15H), 4.31 (s, 4H), 3.49 (s, 2H), 3.11 (t, 2H), 2.87 (t, 2H); MS 531 (M-1).
• Example 1f (3-(((3-Hvdroxy-4-propoxy-benzyl.-(pyridine-3-sulfonv ⁇ -amino)-methyl)-phenyl)-acetic acid Step A: Reaction time of 3.5 h at room temperature.
• Example ig (3-(((4-Pentyl-benzv ⁇ -(pyridine-3-sulfonv ⁇ -amino)-methyl)-phenyl)-acetic acid Step A: Reaction time of 3.5 h at room temperature.
• 1 H NMR 400 MHz, CDCI 3 ) ⁇ 8.98 (s, 1 H), 8.74 (s, 1H), 8.00 (d, 1H), 7.39 (m, 1 H), 7.14-7.26 (m, 2H), 6.95-7.05 (m, 6H), 4.35 (s, 4H), 3.54 (s, 2H), 2.54 (t, 2H), 1.56 (m, 2H), 1.29 (m, 4H), 0.88 (t, 3H); MS 465 (M-1 ).
• Example 1i (3-(((4-lsopropoxy-benzv ⁇ -(pyridine-3-sulfonv ⁇ -amino)-methylVphenv ⁇ -acetic acid Step A: Reaction time of 3.5 h at room temperature.
• 1 H NMR 400 MHz, CDCI 3 ) ⁇ 8.97 (s, 1 H), 8.74 (s, 1 H), 8.03 (m, 1H), 7.42 (m, 1H), 6.94-7.25 (m, 6H), 6.72 (m, 2H), 4.48 (m, 1H), 4.32 (m, 4H), 3.52 (s, 2H), 1.29 (t, 6H); MS 453 (M-1).
• Example 1i (3-(((4-Chloro-thiophen-2-ylmethyl)-(pyridine-3-sulfonv ⁇ -amino)-methv ⁇ -phenv ⁇ - acetic acid
• Step A Reaction time of 3.5 h at room temperature.
• 1 H NMR 400 MHz, CDCI 3 ) ⁇ 9.01 (s, 1H), 8.79 (s, 1H), 8.07 (d, 1H), 7.45 (m, 1 H), 7.20-7.29 (m, 2H), 7.12 (d, 1H), 7.10 (s, 1H), 7.07 (s, 1 H), 4.46 (s, 2H), 4.42 (s, 2H), 3.60 (s, 2H); MS 435 (M-1).
• Example 1k (3-(((4-Butyl-benzv ⁇ -(4-nitro-benzenesulfonyl)-amino)-methv ⁇ -phenvO-acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 8.23 (m, 2H), 7.85 (m, 2H), 7.15-7.25 (m, 2H), 695-7.02 (m, 6H), 4.32 (m, 4H), 3.53 (s, 2H), 2.52 (m, 2H), 1.51 (m, 2H), 1.30 (m, 2H), 0.89 (t, 3H); MS 495 (M-1).
• Example 11 (3-(((4-Butyl-benzyl)-(4-cvano-benzenesulfonyl)-amino)-methv ⁇ -phenv ⁇ -acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 8.21 (d, 1H), 7.67-7.84 (m, 3H), 6.89-7.24 (m, 8H), 4.46 (s, 1H), 4.38 (s, 1 H), 4.32 (m, 2H), 3.54 (s, 1H), 3.38 (s, 1H), 2.55 (m, 2H), 1.58 (m, 2H), 1.33 (m, 2H), 1.29 (s, 1 H), 0.89 (t, 3H); MS 475 (M-1 ).
• Example 1s (3-(((4-lmidazol-1-yl-benzyl,-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)-acetic acid
• 1 H NMR 400 MHz, CD 3 OD
• Example 1t (3-(((Pyridine-3-sulfonyl)-(4-pyrimidin-2-yl-benzv ⁇ -amino)-methyl)-phenyl)-acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 9.10 (s, 1 H), 8.80 (m, 3H), 8.14 (d, 1 H), 8.02 (d, 2H), 7.47 (m, 1 H), 7.06-7.25 (m, 6H), 6.83 (s, 1H), 4.40 (s, 2H), 4.33 (s, 2H), 3.41 (s, 2H); MS 473 (M-1).
• Example 1u (3-(((Pyridine-3-sulfonyl)-(4-thiazol-2-yl-benzyl)-amino)-methyl)-phenyl)-acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 9.11 (s, 1 H), 8.85 (s, 1 H), 8.15 (d, 1H), 7.87 (s, 2H), 7.63 (d, 2H), 7.51 (m, 1 H), 7.37 (s, 1H), 7.07-7.27 (m, 6H), 6.83 (s, 1H), 4.37 (s, 2H), 4.33 (s, 2H), 3.41 (s, 2H).
• Step B N,N-diisopropylethylamine was replaced by triethylamine.
• 1 H NMR 400 MHz, CD 3 OD
• Example 1x (3-(((4-Cvclohexyl-benzyl)-(pyridine-3-sulfonv ⁇ -amino)-methv ⁇ -phenoxy)-acetic acid
• Step B N,N-diisopropylethylamine was replaced by triethylamine.
• Example 1aa (3-(((4-tert-Butyl-benzv ⁇ -(pyridine-3-sulfonyl)-amino)-methv ⁇ -phenoxy)-acetic acid
• Step B N,N-diisopropylethylamine was replaced by triethylamine.
• Example 1ab (3-(((3-(3-Chloro-phenv ⁇ -propyl)-(pyridine-3-sulfonyl)-amino)-methv ⁇ -phenoxy)- acetic acid
• Step B N,N-diisopropylethylamine was replaced by triethylamine.
• Step B N,N-diisopropylethylamine was replaced by triethylamine.
• 1 H NMR 400 MHz, CDCI 3 ) ⁇ 7.66 (s, 1 H), 7.08-7.31 (m, 6H), 6.70-6.78 (m, 3H), 4.54 (s, 2H), 4.35 (s, 4H), 3.68 (s, 3H), 1.27 (s, 9H); MS 469.9 (M-1).
• Example 1ad (3-(((4-Cvclohexyl-benzyl)-(pyridine-3-sulfonv ⁇ -amino)-methv ⁇ -phenv ⁇ -acetic acid
• Step B N,N-diisopropylethylamine was replaced by triethylamine.
• 1 H NMR 400 MHz, CDCI 3 ) ⁇ 8.98 (bs, 1 H), 8.75 (bs, 1 H), 7.98 (d, 1 H), 7.39 (bs, 1H), 6.97-7.25 (m, 8H), 4.36 (d, 4H), 3.54 (s, 2H), 2.44 (s, 1H), 1.72-1.82 (m, 4H), 1.24-1.36 (m, 5H); MS 476.9 (M-1).
• Example 1af (3-(((4-Phenoxy-benzyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)-acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 9.00 (bs, 1 H), 8.76 (bs, 1 H), 8.04 (d, 1 H), 7.41 (t, 1H), 7.35 (m, 1 H), 6.86-7.32 (m, 10H), 6.84 (d, 2H), 4.37 (d, 4H), 3.54 (s, 2H); MS 487 (M- 1).
• Example lag (3-(((4-(2-Oxo-pyrrolidin-1-yl)-benzyl)-(pyridine-3-sulfonv ⁇ -amino)-methyl)-phenyl)- acetic acid
• Step B N,N-diisopropylethylamine was replaced by triethylamine.
• Example 1ah (3-((Benzo[1.3ldioxol-5-ylmethyl-(pyridine-3-sulfonyl)-amino)-methyl.-phenv ⁇ -acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 8.98 (s, 1 H), 8.76 (s, 1 H), 8.04 (d, 1 H), 7.41 (m, 1 H), 7.14-7.20 (m, 2H), 7.00 (d, 1H), 6.94 (s, 1H), 6.64 (t, 2H), 6.55 (d, 1H), 4.34 (s, 2H), 4.26 (s, 2H), 3.54 (s, 2H); MS 439 (M-1 ).
• Example 1ai (3-((Benzo[1.3ldioxol-5-ylmethyl-(pyridine-3-sulfonyl)-amino)-methyl.-phenv ⁇ -acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 8.98 (s, 1 H
• Example 1ai (3-(((Pyridine-3-sulfonyl)-(4-pyrimidin-5-yl-benzyl)-aminoVmethv ⁇ -phenyl)-acetic acid 1 H NMR (400 MHz, CD 3 OD) ⁇ 9.17 (s, 1 H), 9.01 (s, 1 H), 8.77 (s, 1 H), 7.57 (m, 4H), 7.45 (d, 2H), 7.05-7.16 (m, 5H), 4.48 (s, 2H), 4.43 (s, 2H), 3.45 (s, 2H).
• Example 1ak (3-(((Pyridine-3-sulfonyl)-(4-pyrimidin-5-yl-benzyl)-aminoVmethv ⁇ -phenyl)-acetic acid 1 H NMR (400 MHz, CD 3 OD) ⁇ 9.17 (s, 1 H), 9.01 (s, 1 H), 8.77 (s, 1 H), 7.57 (m, 4H), 7.45
• Example 1am (3-(((4-Butyl-benzvO-phenylmethanesulfonyl-amino)-methyl.-phenyl)- acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.31-6.96 (m, 13H), 4.13 (s, 2H), 4.05 (s, 2H), 4.03 (s, 2H), 3.62 (s, 2H), 2.60 (t, 2H), 1.58 (m, 2H), 1.33 (m, 2H), 0.91 (t, 3H); MS 464 (M-1).
• Step A Triethylamine was replaced with N,N-diisopropylethylamine.
• 1 H NMR 400 MHz, CDCI 3 ) ⁇ 9.18 (d, 1 H), 8.82 (d, 1 H), 8.05 (d, 1 H), 7.73-7.20 (m, 8H), 6.60 (d, 1H), 4.35 (s, 2H), 3.22 (t, 2H), 2.70 (t, 2H), 1.85-1.70 (m, 2H).
• Example 2 (3-(((2-(3-Chloro-phenoxy)-ethyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)-acetic acid
• Step A Alkylation (3-(((2-(3-Chloro-phenoxy)-ethyl)-(pyridine-3-sulfonyl)-amino)-methyl)-phenyl)-acetic acid methyl ester.
• Examples 2a-2c Examples 2a-2c were prepared from the appropriate starting materials in a manner analogous to the method of Example 2.
• 1 H NMR 400 MHz, CDCI 3 ) ⁇ 9.08 (bs, 1 H), 8.81 (bs, 1 H), 8.11 (d, 1 H), 7.48 (bs, 1 H), 7.12-7.28 (m, 4H), 6.98 (s, 2H), 6.19 (d, 1H), 5.86 (m, 1H), 4.38 (s, 2H), 3.93 (d, 2H), 3.58 (s, 2H).
• Example 2b (3-(((2-(3.5-Dichloro-phenoxy)-ethyl)-(pyridine-3-sulfonyl)-amino)-methv ⁇ -phenv ⁇ - acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 8.96 (bs, 1 H), 8.70 (bs, 1 H), 8.04 (d, 1 H), 7.41 (m, 1 H), 7.24-7.09 (m, 4H), 6.86 (s, 1 H), 6.47 (s, 2H), 4.44 (s, 2H), 3.86 (m, 2H), 3.49 (s, 2H), 3.31 (m, 2H).
• Example 3 5-(3-((2-Benzylsulfanyl-ethv ⁇ -(pyridine-3-sulfonyl)-amino)-propyl)-thiophene-2- carboxylic acid
• Step A Reductive Amination 5-(3-(2-Benzylsulfanyl-ethylamino)-propyl)-thiophene-2-carboxylic acid tert-butyl ester. Step A was performed in a manner analogous to the method of Step A of Example 1.
• Step B Amide Formation 5-(3-((2-Benzylsulfanyl-ethvO-(pyridine-3-sulfonyl)-amino)-propyl)-thiophene-2- carboxylic acid tert-butyl ester.
• Step B was performed in a manner analogous to the method of Step B of Example 1 , except triethylamine was used in place of N,N- diisopropylethylamine.
• Step C Ester Hydrolysis 5-(3-((2-Benzylsulfanyl-ethv ⁇ -(pyridine-3-sulfonv ⁇ -amino)-propy ⁇ -thiophene-2- carboxylic acid «TFA.
• a solution of 5-(3-((2-benzylsulfanyl-ethyl)-(pyridine-3-sulfonyl)- amino)-propyl)-thiophene-2-carboxylic acid tert-butyl ester prepared of Example 3, Step B (0.038 g) in 1 mL CH 2 CI 2 was cooled to 0°C and 1 mL TFA was added. The mixture was warmed to room temperature and was stirred for 1 h. The CH 2 CI 2 and TFA were removed by evaporation, azeotroping with added CH 2 CI 2 to yield the title compound (46.3 mg).
• MS 475 (M-1 ).
• Examples 3a-3i were prepared from the appropriate starting materials in a manner analagous to the method of Example 3 with variations thereto noted.
• Example 3b (3-(((Pyridine-3-sulfonyl)-(4-thiazol-2-yl-benzylVamino)-methyl)-phenoxy)-acetic acid»2TFA 1 H NMR (400 MHz, CDCI 3 ) ⁇ 9.40 (bs, 1 H), 8.98 (s, 1 H), 8.84 (s, 1H), 8.28 (m, 1 H), 8.10 (s, 1H), 7.78 (m, 2H), 7.68 (m, 1H), 7.51 (s, 1H), 7.24 (m, 3H), 7.12 (t, 1H), 6.77 (m, 1 H), 6.48 (s, 1 H), 4.53 (s, 2H), 4.45 (s, 2H), 4.34 (s, 2H); MS 494 (M-1).
• Example 3c (3-(((Pyridine-3-sulfonyl)-(4-thiazol-2-yl-benzylVamino)-methyl)-phenoxy)-acetic acid
• Example 3d (3-(((1-Methyl-1H-imidazole-4-sulfonyl.-(4-thiazol-2-yl-benzv ⁇ -amino.-methyl)- phenoxy)-acetic acid*2TFA 1 H NMR (400 MHz, CD 3 OD) ⁇ 7.93 (s, 1 H), 7.85 (d, 1H), 7.76 (d, 2H), 7.70 (s, 1H), 7.60 (d, 1 H), 7.26 (d, 2H), 7.09 (t, 1 H), 6.75 (d, 2H), 6.68 (s, 1 H), 4.51 (s, 2H), 4.41 (s, 2H), 4.35 (s, 2H), 3.76 (s, 3H); MS 498 (M+).
• Step A No triethylamine was used in Step A.
• the TFA salt was converted to the HCI salt by stirring in 2 equivalents 1 N HCI followed by removal of water and drying in vacuo.
• Step A No triethylamine was used in Step A.
• the TFA salt was converted to the HCI salt by stirring in 2 equivalents 1 N HCI followed by removal of water and drying in vacuo.
• Example 3g (3-(((Pyridine-3-sulfonyl)-(4-pyridin-3-yl-benzyl)-amino)-methyl)-phenoxy)-acetic acid»HCI
• TFA salt was converted to the HCI salt by stirring in 2 equivalents 1 N HCI followed by removal of water and drying in vacuo.
• Example 3h
• Example 4 5-(3-((3-(3-Chloro-phenyl)-propyl)-(pyridine-3-sulfonyl)-amino)-propyl)-thiophene-2- carboxylic acid
• Step A Sulfonamide formation 5-(3-((3-(3-Chloro-phenyl)-propyl)-(pyridine-3-sulfonyl)-amino)-propy ⁇ -thiophene-2- carboxylic acid methyl ester.
• Step B Ester Hydrolysis 5-(3-((3-(3-Chloro-phenv ⁇ -propylHpyridine-3-sulfonylVamino)-propyl)-thiophene-2- carboxylic acid.
• Examples 4a-4h were prepared from the appropriate starting in a manner analogous to the method of Example 4.
• Example 4c 5-(3-(Benzenesulfonyl-(3-(3-chloro-phenyl,-propyl)-amino)-propyl)-thiophene-2- carboxylic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 6.70-7.92 (m, 11H), 3.26 (m, 4H), 3.05 (m, 4H), 2.73 (m, 2H), 2.50 (m, 2H), 1.70 (m, 2H); MS 578(M+1 ), 576 (M-1 ).
• Example 4f 5-(3-((3-(3-Chloro-phenyl)-propy ⁇ -(pyridine-3-sulfonyl)-amino)-propyl)-furan-2- carboxylic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 9.00 (m, 1H), 8.70
• Example 5 5-(3-((3-(3-Chloro-phenv ⁇ -propyl)-(pyridine-3-carbonyl)-amino)-propyl)-thiophene-2- carboxylic acid
• Step A Amide formation 5-(3-((3-(3-Chloro-phenyl)-propyl)-(pyridine-3-carbonyl)-amino)-propylHhiophene-2- carboxylic acid methyl ester.
• Step B Ester Hydrolysis Step B was performed in a manner analogous to the method of Step B of Example 4.
• Examples 5a-5b were prepared from the appropriate starting in a manner analogous to the method of Example 5.
• Example 5a was prepared from the appropriate starting in a manner analogous to the method of Example 5.
• Example 6 5-(3-((3-(3-Chloro-phenyl)-propyl)-(pyridin-3-yl-acetyl)-amino)-propyl)-thiophene-2- carboxylic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.
• Step A Amide formation 5-(3-((2-Chloro-benzenesulfonv ⁇ -(3-(3-chloro-phenv ⁇ -propyl)-amino)-propyl)- thiophene-2-carboxylic acid tert-butyl ester.
• Step B Ester Hydrolysis
• Examples 6a-6i Examples 6a-6j were prepared from the appropriate starting material in a manner analogous to the method of Example 6.
• Example 6b 5-(3-((3-(3-Chloro-phenv ⁇ -propy ⁇ -(2.4-dioxo-1 ,2.3,4-tetrahvdro-guinazoline-6- sulfonyl)-amino)-propyl)-thiophene-2-carboxylic acid
• Example 6d 5-(3-((3-(3-Chloro-phenyl.-propyn-(4-(3,5-dioxo-4.5-dihvdro-3H- ⁇ .2.41triazin-2-yl)- benzenesulfonyl)-amino)-propyl)-thiophene-2-carboxylic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.60-7.92 (m, 4H), 6.80 (m, 7H), 3.22 (m, 4H), 2.80 (m, 2H), 2.60 (m, 2H), 1.82 (m, 2H), 1.22 (m, 2H); MS 587.1 (M-1).
• Example 6g 5-(3-((4-Bromo-benzenesulfonylH3-(3-chloro-phenyl.-propyl)-amino)-propyl)- thiophene-2-carboxylic acid 1 H NMR (400 MHz, CDCI 3
• Example 7 5-(3-(1-(3-(3-Chloro-phenv ⁇ -propy
• Step A Isocyanate addition 5-(3-(1-(3-(3-Chloro-phenyl)-propy ⁇ -3-ethyl-ureido)-propy ⁇ -thiophene-2-carboxylic acid tert-butyl ester.
• a stock solution of 5-(3-(3-(3-chloro-phenyl)-propylamino)- propyl)-thiophene-2-carboxylic acid tert-butyl ester (from Preparation 9, 0.10 g, 0.254 mmol) in 10 mL CH 2 CI 2 was prepared and 1 mL (0.010 g, 0.0254 mmol) was added to a 1 dram vial. Triethylamine (0.7 mL, 0.051 mmol) and ethyl isocyanate (0.004 g, 0.051 mmol) were added and the mixture was stirred overnight at room temperature. The solution was diluted with 2 mL CH 2 CI 2 .
• Step B Ester Hydrolysis
• Step B was performed in a manner analogous to the method of Step B of Example 6.
• Examples 7a-7j were prepared from the appropriate starting materials in a manner analogous to the method of Example 7.
• Example 7c 5-(3-(1-(3-(3-Chloro-phenyl)-propyl)-3-(3.4-dichloro-phenv ⁇ -ureido)-propyl)- thiophene-2-carboxylic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 6.80-7.60 (m, 9H), 3.20 (m, 4H), 2.90 (m, 2H), 2.60 (m, 2H), 1.86-2.00 (m, 4H); MS 527.0 (M+1), 525.0 (M-1).
• Example 7e 5-(3-(1-(3-(3-Chloro-phenyl)-propyl)-3-propyl-ureido)-propyl)- thiophene-2-carboxylic acid 1 H NMR (400 MHz, CDCI 3 )
• Example 7f 5-(3-(1-(3-(3-Chloro-phenyl)-propyl ' )-3-(2.3-dichloro-phenyl)-ureido)-propyl)- thiophene-2-carboxylic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.70 (bs, 1H), 7.00-7.30 (m, 7H), 6.80 (bs, 1H), 3.20 (m, 4H), 2.80 (m, 2H), 2.60 (m, 2H), 1.75-2.00 (m, 4H); MS 527 (M+1 ), 525.1 (M-1 ).
• Example 7h 5-(3-(1-(3-(3-Chloro-phenyl)-propy ⁇ -3-(2.6-difluoro-phenyl)-ureido)-propyl.-thiophene-
• Example 7i 5-(3-(1-(3-(3-Chloro-phenv ⁇ -propyl)-3-(4-fluoro-phenyl)-ureido)-propyl)-thiophene-2- carboxylic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.70 (bs, 1 H), 7.00-7.60 (m, 8H), 6.80 (bs, 1H), 3.30 (m, 4H), 2.90 (m, 2H), 2.60 (m, 2H), 1.80-2.00 (m, 4H); MS 475.1 (M+1), 473.1 (M-1).
• Step A Amide formation 5-(3-(1-(3-(3-Chloro-phenyl)-propyO-3-ethyl-ureido)-propyl)-thiophene-2-carboxylic acid tert-butyl ester.
• Step B Ester Hydrolysis Step B was performed in a manner analogous to the method of Step B of Example 6.
• Examples 8a-8c Examples 8a-8c were prepared from the appropriate starting material in a manner analogous to the method of Example 8.
• Step A Reductive Amination (3-,(4-Butyl-benzylamino.-methvO-phenyl)-acetic acid methyl ester.
• a solution of 4- butyl-benzylamine (from Preparation 15, 0.918 g, 6 mmol) in MeOH was added to 4N HCI in dioxane (0.75 mL, 3 mmol) followed by addition of (3-formyl-phenyl)-acetic acid methyl ester (from Preparation 13, 0.534 g, 3.0 mmol).
• NaCNBH 3 0.194 mL, 3 mmol
• Step B Amide Formation (3-(((4-Butyl-benzv ⁇ -(pyridine-3-sulfonyl)-aminoVmethyl)-phenyl)-acetic acid methyl ester. Step B was performed in a manner analogous to the method of Step B of Example 1 to provide the title compound.
• Step C Ester Hydrolysis (3-(((4-Butyl-benzv ⁇ -(pyridine-3-sulfonv ⁇ -amino)-methyl)-phenyl)-acetic acid. Step C was performed in a manner analogous to the method of Step C of Example 1 to provide the title compound.
• Examples 9a-9d were prepared from the appropriate starting materials in a manner analogous to the method of Example 9.
• Example 9a (3-((Benzenesulfonyl-(4-butyl-benzviyamino)-methv0-phenvD-acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.83 (d, 2H), 7.46-7.58 (m, 3H), 7.24 (s, 1H), 7.14 (m, 2H), 6.86-6.98 (m, 5H), 4.29 (d, 4H), 3.51 (s, 2H), 2.52 (t, 2H), 1.53 (m, 2H), 1.30 (m, 2H), 0.90 (t, 2H); MS 450 (M-1 ).
• Example 9b (3-(((4-Butyl-benzylV(thiophene-2-s ⁇ lfonyl)-amino)-methyl)-phenv ⁇ -acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.53 (m, 2H), 7.16 (m, 2H), 6.89-7.14 (m, 7H), 4.27 (d, 4H), 3.52 (s, 2H), 2.49 (t, 2H), 1.51 (m, 2H), 1.29 (m, 2H), 0.88 (t, 2H); MS 456 (M- 1).
• Example 9c (3-(((4-Acetylamino-benzenesulfonyl)-(4-butyl-benzv ⁇ -amino)-methyl)-phenyl)-acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.69 (m, 2H), 7.49 (d, 2H), 7.06-7.23 (m, 6H), 6.91 (d, 1 H), 6.68 (s, 1 H), 4.30 (d, 4H), 3.44 (s, 2H), 2.54 (t, 2H), 2.17 (s, 3H), 1.54 (m, 2H), 1.29 (m, 2H), 0.89 (t, 2H); MS 507(M-1).
• Example 9d (3-(((BenzoH ,2.51oxadiazole-4-sulfonylH4-butyl-benzvO-aminoVmethyl)-phenyl)- acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.94 (d, 1 H), 7.88 (d, 2H), 7.36 (t, 1 H), 7.07 (s, 2H), 6.90-6.96 (m, 6H), 53 (d, 4H), 3.46 (s, 2H), 2.46 (t, 2H), 1.47 (m, 2H), 1.26 (m, 2H), 0.88 (t, 2H); MS 4.92 (M-1).
• Example 10 (3-(((BenzoH ,2.51oxadiazole-4-sulfonylH4-butyl-benzvO-aminoVmethyl)-phenyl)- acetic acid 1 H NMR (400 MHz, CDCI 3 ) ⁇ 7.94 (d, 1 H), 7.88 (d, 2
• Step A Reductive Amination (3-((4-Pyrimidin-2-yl-benzylamino,-methyl,-phenoxy)-acetic acid t-butyl ester. Step A was performed in a manner analogous to the method of Step A of Example 1.
• Step B Amide Formation (3-(((1-Methyl-1 H-imidazole-4-sulfonyl)-(4-pyrimidin-2-yl-benzyl.-amino)-methyl)- phenoxy,-acetic acid tert-butyl ester.
• Step B was performed in a manner analogous to the method of Step B of Example 1 using triethylamine in place of N,N- diisopropylethylamine as base.
• Step C Ester Hydrolysis (3-(((1-Methyl-1 H-imidazole-4-sulfonyl)-(4-pyrimidin-2-yl-benzyl)-amino)-methv ⁇ - phenoxyVacetic acid»HCI.
• Step B Aldehyde formation 5-(3-Oxo-propyl)-thiophene-2-carboxylic acid tert-butyl ester.
• Step A (0.50 g, 1.89 mmol) in 5 mL DMF was added allyl alcohol (0.51 mL, 7.57 mmol) followed by NaHC0 3 (0.397 g, 4.72 mmol), tetrabutylammonium chloride (0.525g, 1.89 mmol), and palladium acetate (0.021 g, 0.094 mmol).
• the reaction was placed in an oil bath heated to 65°C and was heated to 90°C for 2 h.
• the mixture was diluted with EtOAc and 25 mL water and the solids were removed by filtration through Celite ® .
• Step A 5-(3-tert-Butoxycarbonylamino-prop-1-vnyl)-thiophene-2-carboxylic acid methyl ester.
• Step B 5-(3-tert-Butoxycarbonylamino-propyl)-thiophene-2-carboxylic acid methyl ester.
• the reaction ' was filtered through Celite ® with the aid of MeOH and the filtrate was concentrated in vacuo to afford the title compound of Step B as a solid (1.93 g).
• Step C 5-(3-Amino-propyl)-thiophene-2-carbo ⁇ ylic acid methyl ester.
• a solution of 5-(3-tert- butoxycarbonylamino-propyl)-thiophene-2-carboxylic acid methyl ester prepared of Preparation 5, Step B (0.118 g, 0.5 mmol) in 50 mL MeOH was cooled to 0°C and was saturated with HCI (g). The reaction was stirred at room temperature for 90 minutes. The solution was concentrated to a solid which was partitioned between EtOAc and saturated NaHCO 3 . The layers were separated, and the combined organic layers were washed with brine, dried over MgSO 4 , filtered and concentrated in vacuo to afford the title compound as an oil (399 mg). MS 200 (M+1 ).
• Preparation 6 5-(3-Amino-propyO-furan-2-carboxylic acid methyl ester hydrochloride salt.
• the compound of Preparation 6 was prepared from the appropriate starting materials in a manner analogous to the method of Preparation 5 with the following exceptions: (1) the hydrogenation performed in Step B was carried out for 5.5 h; and (2) in Step C, the reaction was stirred for 16 h at room temperature and was concentrated in vacuo to provide the title compound as the hydrochloride salt.
• Step A Prop-2-ynyl-carbamic acid benzyl ester.
• benzylchloroformate 13.37 g, 78.2 mmol
• the reaction was stirred for 16 h and the volatiles were removed in vacuo.
• the residue was dissolved in EtOAc and the organic solution was washed with water (2x).
• the organic solution was washed with dilute aqueous HCI followed by saturated NaHCO 3 .
• the organic solution was dried over MgSO , filtered, and concentrated in vacuo to provide the title compound of Step A(4.43 g).
• Step B 5-(3-Benzyloxycarbonylamino-prop-1 -vnyl)-thiophene-2-carboxylic acid tert-butyl ester.
• the title compound of Step B was prepared from the appropriate starting material in a manner analagous to the method used in Step A of Preparation 5.
• Step C 5-(3-Amino-propyl.-thiophene-2-carboxylic acid tert-butyl ester.
• Preparations 9-10 The compounds of Preparations 9 and 10 were prepared from the appropriate starting materials in a manner analogous to the method of Preparation 8.
• the mixture was concentrated in vacuo to remove THF and was diluted with water.
• the aqueous solution was extracted with EtOAc.
• the organic solution was washed with water, dried over MgS0 4 , filtered and concentrated in vacuo.
• the residue was dissolved in CHCI 3 and the organic solution was washed with 1M HCI.
• the aqueous solution was basified to pH 11 with 1M NaOH and the product was extracted into CHCI 3 .
• the organic solution was dried over MgS0 , filtered and concentrated in vacuo to afford the title compound as a yellow oil (0.134 g).
• Preparation 14 (3-((Pyridine-3-sulfonylamino.-methyl.-phenyl,-acetic acid methyl ester.
• PREPARATION 18 (3-Aminomethyl-phenyl)-acetic acid methyl ester hydrochloride.
• the title compound was prepared from (3-cyano-phenyl)-acetic acid methyl ester (from Preparation 13, Step A) using the procedure described for Preparation 15, Method B except the hydrogenation was performed in MeOH.
• the catalyst was removed via filtration and the organic solution was concentrated in vacuo. The resulting solid was stirred in
• Step A (3-Formyl-phenoxyVacetic acid tert-butyl ester.
• 3- hydroxybenzaldehyde (5.00 g, 40.9 mmol) in DMF (40 mL) was added 1M potassium tert-butoxide in tert-butanol (40.9 mL, 40.9 mmol).
• the reaction was stirred for 2 minutes and tert-butyl bromoacetate (6.61 mL, 40.9 mmol) was added.
• the reaction was stirred for 1 hour and was quenched with 200 mL water.
• the product was extracted into EtOAc and the organic solution was washed with water, dried over MgS0 4 , filtered, and concentrated in vacuo.
• Step B (3-(Hvdroxyimino-methyl)-phenoxy)-acetic acid tert-butyl ester.
• NH 2 OH «HCI (0.66 g, 9.54 mmol)
• pyridine 3.5 mL, 43.4 mmol
• Preparations 22-27 were prepared from the appropriate starting materials in a manner analogous to the method of Preparation 21.
• STEP B Benzylic Bromination A mixture of 4-[(1-acetyloxy)-hexyl]-toluene prepared of Preparation 30, Step A (2.082 g, 8.89 mmol), N-bromosuccinimide (1.58 g, 8.89 mmol), and catalytic 2,2'- azobisisobutyronitrile in carbon tetrachloride (30 mL) was heated at reflux for 2 h. The reaction was cooled and was washed with aqueous NaHC0 3 (saturated), dried over MgS0 4 , filtered, and concentrated. The product was purified by flash chromatography (5% EtOAc/hexanes) to afford the title compound of Preparation 30
• Preparation 35 4-Formyl-N-methyl-benzenesulfonamide.
• the title compound can be prepared using the method described by Koetschet in Helv. Chim. Acta., 12, 682, 1929.
• Preparation 36 4-Chloro-thiophene-2-carbaldehvde.
• the title compound can be prepared using the method described by Raggon and coworkers in Org. Prep. Proced. Int.; EN, 27 , 2, 233-6, 1995.
• Preparation 37
• Preparation 38 4-lmidazol-1-yl-benzaldehvde.
• the title compound can be prepared using the method described by Sircar and coworkers in J. Med. Chem. 30, 6, 1023-9, 1987.
• Atmospheric pressure chemical ionization (APCI) mass spectra were obtained on a Fisons Platform II Spectrometer. Chemical ionization mass spectra were obtained on a Hewlett-Packard 5989 instrument (Hewlett-Packard Co., Palo Alto, California) (ammonia ionization, PBMS). Where the intensity of chlorine or bromine-containing ions are described the expected intensity ratio was observed (approximately 3:1 for 35 CI/ 37 CI-containing ions) and 1 :1 for 79 Br/ 81 Br- containing ions) and the intensity of only the lower mass ion is given.
• APCI Atmospheric pressure chemical ionization
• the terms "concentrated” and “coevaporated” refer to removal of solvent at water aspirator pressure on a rotary evaporator with a bath temperature of less than 45°C. Reactions conducted at "0-20°C” or “0-25°C” were conducted with initial cooling of the vessel in an insulated ice bath which was allowed to warm to room temperature over several hours. The abbreviation “min” and “h” stand for “minutes” and “hours” respectively. DTT means dithiothreitol. DMSO means dimethyl sulfoxide. EDTA means ethylenediamine tetraacetic acid.
• Some of the preparation methods useful for the preparation of the compounds described herein may require protection of remote functionality (e.g., primary amine, secondary amine, carboxyl in Formula I precursors).
• the need for such protection will vary depending on the nature of the remote functionality and the conditions of the preparation methods. The need for such protection is readily determined by one skilled in the art. The use of such protection/deprotection methods is also within the skill in the art. For a general description of protecting groups and their use, see T.W. Greene, Protective Groups in Organic Synthesis. John Wiley & Sons, New York, 1991. Biological Assays Determination of cAMP Elevation in 293-S Cell Lines Stably Overexpressing Recombinant Human EP? and EP Receptors.
• cDNAs representing the complete open reading frames of the human EP 2 and EP receptors are generated by reverse transcriptase polymerase chain reaction using oligonucleotide primers based on published sequences (1, 2) and RNA from primary human kidney cells (EP 2 ) or primary human lung cells (EP 4 ) as templates.
• cDNAs are cloned into the multiple cloning site of pcDNA3 (Invitrogen Corporation, 3985B Sorrento Valley Blvd., San Diego, CA 92121) and used to transfect 293-S human embryonic kidney cells via calcium phosphate co-precipitation. G418-resistant colonies are expanded and tested for specific [ 3 H]PGE 2 binding.
• Transfectants demonstrating high levels of specific [ 3 H]PGE 2 binding are further characterized by Scatchard analysis to determine Bmax and Kds for PGE 2 .
• Constituitive expression of both receptors in parental 293-S cells is negligible.
• Cells are maintained in RPMI supplemented with fetal bovine serum (10% final) and G418 (700 ug/ml final).
• cAMP responses in the 293-S/EP 2 and 293-S/EP 4 lines are determined by detaching cells from culture flasks in 1 ml of Ca++ and Mg++ deficient PBS via vigorous pounding, adding serum-free RPMI to a final concentration of 1 X 10 6 cells/ml, and adding 3-isobutyl-1-methylxanthine (IBMX) to a final concentration of 1 mM.
• IBMX 3-isobutyl-1-methylxanthine
• the compound to be tested is then added to cells at 1 :100 dilutions such that final DMSO or ethanol concentrations is 1%.
• the tubes are covered, mixed by inverting two times, and incubated at 37 °C for 12 minutes. Samples are then lysed by incubation at 100 °C for 10 minutes and immediately cooled on ice for 5 minutes. Cellular debris is pelleted by centrifugation at 1000 X g for 5 minutes, and cleared lysates are transferred to fresh tubes.
• cAMP concentrations are determined using a commercially available cAMP radioimmunoassay kit RIA (NEK-033, DuPont/NEN Research Products, 549 Albany St., Boston, MA 02118) after diluting cleared lysates 1:10 in cAMP RIA assay buffer (included in kit). Typically, one treats cells with 6-8 concentrations of the compound to be tested in 1 log increments. EC50 calculations are performed on a calculator using linear regression analysis on the linear portion of the dose response curves.
• the cells are lysed by sonification with a Branson Sonifier (Model #250, Branson Ultrasonics Corporation, Danbury, CT) in 2 fifteen second bursts. Unlysed cells and debris are removed by centrifugation at 100 x g for 10 min. Membranes are then harvested by centrifugation at 45,000 x g for 30 minutes.
• a Branson Sonifier Model #250, Branson Ultrasonics Corporation, Danbury, CT
• Pelleted membranes are resuspended to 3-10 mg protein per ml, protein concentration being determined of the method of Bradford [Bradford, M., Anal. Biochem., 72, 248 (1976)]. Resuspended membranes are then stored frozen at -80 °C until use.
• Binding Assay Frozen membranes prepared as above are thawed and diluted to 1 mg protein per ml in Buffer A above. One volume of membrane preparation is combined with 0.05 volume test compound or buffer and one volume of 3 nM 3 H-prostaglandin E 2 ( #TRK 431 , Amersham, Arlington Heights, IL) in Buffer A. The mixture (205 ⁇ L total volume) is incubated for 1 hour at 25°C. The membranes are then recovered by filtration through type GF/C glass fiber filters ( #1205-401 , Wallac, Gaithersburg, MD ) using a Tomtec harvester ( Model Mach II/96, Tomtec, Orange, CT).
• the membranes with bound 3 H-prostaglandin E 2 are trapped by the filter, while the buffer and unbound 3 H- prostaglandin E 2 pass through the filter into waste. Each sample is then washed 3 times with 3 ml of [50 mM Tris-HCl (pH 7.4), 10 mM MgCI 2 , 1 mM EDTA].
• the filters are then dried by heating in a microwave oven. To determine the amount of 3 H- prostaglandin bound to the membranes, the dried filters are placed into plastic bags with scintillation fluid and counted in a LKB 1205 Betaplate reader (Wallac, Gaithersburg, MD). IC50s are determined from the concentration of test compound required to displace 50% of the specifically bound 3 H-prostaglandin E 2 .
• the full length EP-i receptor is made as disclosed in Funk et al., Journal of Biological Chemistry, 1993, 268, 26767-26772.
• the full length EP 2 receptor is made as disclosed in Regan et al., Molecular Pharmacology, 1994, 46, 213-220.
• the full length EP 3 receptor is made as disclosed in Regan et al., British Journal of Pharmacology, 1994, 112, 377-385.
• the full length EP 4 receptor is made as disclosed in Bastien, Journal of Biological Chemistry, 1994, 269, 11873-11877.
• These full length receptors are used to prepare 293S cells expressing the EP ⁇ EP 2 , EP 3 and EP 4 receptors. 293S cells expressing either the human EP-i, EP 2 , EP 3 or EP 4 prostaglandin
• E 2 receptors are generated according to methods known to those skilled in the art.
• PCR polymerase chain reaction
• primers corresponding to the 5' and 3' ends of the published full length receptor are made according to the well known methods disclosed above and are used in an RT-PCR reaction using the total RNA from human kidney (for EP-i), human lung (for EP 2 ), human lung (for EP 3 ) or human lymphocytes (for EP 4 ) as a source.
• PCR products are cloned by the TA overhang method into pCR2.1 (Invitrogen, Carlsbad, CA) and identity of the cloned receptor is confirmed by DNA sequencing.
• 293S cells (Mayo, Dept. of Biochemistry, Northwestern Univ.) are transfected with the cloned receptor in pcDNA3 by electroporation. Stable cell lines expressing the receptor are established following selection of transfected cells with G418.
• Clonal cell lines expressing the maximal number of receptors are chosen following a whole cell 3 H-PGE 2 binding assay using unlabeled PGE 2 as a competitor.

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