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WO2009152431A1 - Antagonistes des récepteurs p2y<sb>1</sb> - Google Patents

Antagonistes des récepteurs p2y<sb>1</sb> Download PDF

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WO2009152431A1
WO2009152431A1 PCT/US2009/047204 US2009047204W WO2009152431A1 WO 2009152431 A1 WO2009152431 A1 WO 2009152431A1 US 2009047204 W US2009047204 W US 2009047204W WO 2009152431 A1 WO2009152431 A1 WO 2009152431A1
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compound
salt
alkyl
substituted
conjugate
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Kenneth A. Jacobson
Sonia De Castro
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US Department of Health and Human Services
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
    • C07F9/00—Compounds containing elements of Groups 5 or 15 of the Periodic Table
    • C07F9/02—Phosphorus compounds
    • C07F9/547—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom
    • C07F9/6561—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom containing systems of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring or ring system, with or without other non-condensed hetero rings
    • C07F9/65616—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom containing systems of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring or ring system, with or without other non-condensed hetero rings containing the ring system having three or more than three double bonds between ring members or between ring members and non-ring members, e.g. purine or analogs
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
    • C07H19/00—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof
    • C07H19/02—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof sharing nitrogen
    • C07H19/04—Heterocyclic radicals containing only nitrogen atoms as ring hetero atom
    • C07H19/16—Purine radicals
    • C07H19/20—Purine radicals with the saccharide radical esterified by phosphoric or polyphosphoric acids

Definitions

  • the family of P2Y receptors consists of eight subtypes of G protein-coupled receptors (GPCRs) that respond to extracellular purine and pyrimidine nucleotides.
  • GPCRs G protein-coupled receptors
  • Native ligands of the these receptors include ATP, ADP, UTP, UDP, and UDP-glucose.
  • Two subfamilies have been delineated based on second messengers and sequence homology, i.e., a cluster of P2Y h P2Y 2 , P2Y 4 , P2Y 6 , and P2Y n receptors, which activate PLC ⁇ via G q , and a smaller cluster of P2Yi 2 , P2Y 13 , and P2Yi 4 , which inhibit adenylate cyclase via Gj.
  • Platelets possess three P2 receptors for adenine nucleotides: P2Yi and P2Yi 2 , which interact with ADP, and P2Xi, which interacts with ATP.
  • P2Yi receptors are involved in activation of platelets and other hematopoietic cells, as well as in other functions and events such as vascular function, apoptotic events, inflammation, glial cell differentiation and function, and development of vision. Accordingly, there is a desire to find antagonists of the P2Yi receptor for preventing or treating a number of diseases or conditions, particularly platelet aggregation or thrombosis.
  • the invention provides P2Y] receptor antagonists, pharmaceutical compositions comprising the same, and the use of such antagonists in the prevention or treatment of a disease or unfavorable condition that can be ameliorated by antagonizing a P2Yi receptor.
  • the invention also provides conjugates comprising a P2Yi antagonist that is covalently linked to a conjugant such as a dendrimer.
  • the conjugate in an embodiment, includes a dendrimer to which are covalently linked a P2Yi antagonist and one or more ligands other than a P2Yi antagonist.
  • Such conjugates have synergistic effect on the prevention or treatment of diseases or unfavorable conditions.
  • FIG. 1 depicts a reaction scheme (Scheme 1) for preparing compounds 10 to 15 in accordance with an embodiment of the invention.
  • Reagents (i) CuI, (Ph 3 P) 2 PdCl 25 TEA, DMF, rt (room temperature); (ii) TFA, CH 2 Cl 2 , it; (iii) ethylenediamine, MeOH, rt (iv) KOH, H 2 O, rt , (v) TFA, CH 2 Cl 2 , rt.
  • FIG. 2 depicts a reaction scheme (Scheme 2) for preparing biotinylated compounds 16 and 17 or conjugates in accordance with an embodiment of the invention.
  • Reagents (i) biotin, DIEA, HATU, DMF, rt; (ii) biotinamidohexanoic acid 3-sulfo-iV- hydroxysuccinimide ester sodium salt, bicarbonate buffer, DMF; rt (iii) TFA, CH 2 Cl 2 .
  • HATU is the peptide coupling agent 2-(lH-7-Azabenzotriazol-l-yl)— 1,1,3, 3-tetramethyl uronium hexafluorophosphate Methanaminium.
  • Figure 3 depicts a reaction scheme (Scheme 3) to prepare a fluorosulfonyl derivative 18 in accordance with an embodiment of the invention.
  • Reagents (i) DIEA, HATU, DMF, rt.
  • Figure 4 depicts a reaction scheme to (Scheme 4) to prepare compounds 39-42 in accordance with an embodiment of the invention and illustrates the use of Click chemistry to couple P2Yi antagonists to small azido-bearing molecules, wherein AcNH(CH 2 ) 2 - is a representative group of an amine ended dendrimer.
  • Figure 5 depicts a reaction scheme (Scheme 5) illustrating the use of Click chemistry to couple P2Yj antagonists to azido-bearing PAMAM dendrimers in accordance with an embodiment of the invention.
  • the dendrimer may alternatively contain a prosthetic group, for example, coupled to one or more of the free amino groups on the periphery of the polymers.
  • Figure 6 depicts a reaction scheme (Scheme 6) illustrating the use of amide formation and Click chemistry to couple both A 2A adenosine agonists and P2Yi antagonists to the same azido-bearing PAMAM dendrimer in a random fashion, but according to defined ratios, in accordance with an embodiment of the invention.
  • the dendrimer may alternatively contain a prosthetic group, for example, coupled to one or more of the free amino groups on the periphery of the polymers or coupled to one or more of the azido groups.
  • the combined conjugate inhibits platelet aggregation by two parallel and synergistic mechanisms.
  • R 1 is a Ci-C 6 alkyl
  • R 2 is a substituted C 2 -C 20 alkynyl or a substituted Ci-C 2O alkyl C 2 -C 20 alkynyl, or a substituted or unsubstituted C7-C12 cycloalkyl C 2 -C 2O alkynyl, substituted or unsubstituted C 6 - Ci 4 aryl C 2 -C 20 alkynyl, or substituted or unsubstituted heterocyclyl alkyl C 2 -C 2O alkynyl, wherein the each of the substituted R 2 groups includes one or more substituents selected from the group consisting of halo, hydroxy, alkoxy, amino, alkylamino, dialkylamino, mercapto, thioalkoxy, carboxy, alkoxycarbonyl, alkylcarbonylaminoalkyl, arylalkylaminocarbonyl wherein aryl is optionally substituted, aminocarbonyl, alkyny
  • R 1 is preferably a Ci-C 3 alkyl, particularly methyl or ethyl.
  • R 3 is preferably (a), wherein the cyclopentyl ring is conformationally constrained in the Northern (N) conformation due to the methanocarba modification:
  • aryl refers to aromatic moieties such as phenyl, naphthyl, anthracenyl, and biphenyl.
  • heterocyclyl refers to 3-7 membered rings which can be saturated or unsaturated, comprising carbon and one or more heteroatoms such as O, N, and S, and optionally hydrogen; optionally in combination with one or more aromatic rings.
  • heterocyclyl groups include pyridyl, piperidinyl, piperazinyl, pyrazinyl, pyrolyl, pyranyl, tetrahydropyranyl, tetrahydrothiopyranyl, pyrrolidinyl, furanyl, tetrahydrofuranyl, thiophenyl, tetrahydrothiophenyl, purinyl, pyrimidinyl, thiazolyl, thiazolidinyl, thiazolinyl, oxazolyl, triazolyl, tetrazolyl, tetrazinyl, benzoxazolyl, morpholinyl, thiophorpholinyl, quinolinyl, and isoquinolinyl.
  • the alkyl, alkoxy, and alkylamino groups can be linear or branched.
  • a substituent e.g., halo, amino, alkyl, hydroxyl, alkoxy, and others
  • the aromatic ring hydrogen is replaced with the substituent and this can take place in any of the available hydrogens, e.g., 2, 3, 4, 5, and/or 6-position wherein the 1-position is the point of attachment of the aryl group in the compound of the present invention.
  • halo refers to fluorine, chlorine, bromine, and iodine.
  • R 2 is a substituted C 1 -C 20 alkyl C 2 -C 20 alkynyl or a substituted or unsubstituted heterocyclyl alkyl C 2 -C 20 alkynyl, wherein each of the substituted R 2 groups includes one or more substituents selected from the group consisting of halo, hydroxyl, amino, alkylamino, dialkylamino, mercapto, carboxy, alkoxycarbonyl, alkylcarbonylaminoalkyl, arylalkylaminocarbonyl which is optionally substituted at the aryl, aminocarbonyl, alkylaminocarbonyl, dialkylaminocarbonyl, aminoalkylaminocarbonyl, amino (alkylamino) n alkylaminocarbonyl wherein n is 1 to 6, and aminoalkylcarbonylaminoalkylaminocarbonyl.
  • R 2 is a substituted Cj-C 2O alkyl C 2 -C 2O alkynyl that includes a carboxy substituent on the Ci-C 20 alkyl group, for example, a Ci-C 6 alkyl C 2 -C 20 alkynyl group that includes a carboxy substituent, preferably a Ci-C 6 alkyl ethynyl that includes a carboxy substituent.
  • R 2 is C ⁇ C-(CH 2 ) m -COOH, wherein m is 1 to 6, more particularly m is 2 to 4.
  • R 2 is a substituted Ci-C 20 alkyl C 2 -C 20 alkynyl that includes an alkoxycarbonyl substituent, for example, R 2 is a substituted Ci-C 20 alkyl ethynyl that includes an alkoxycarbonyl substituent.
  • R 2 is C ⁇ C-(CH 2 ) P - COOCH 3 , wherein p is 1 to 6, more particularly p is 2, 3, or 4.
  • R 2 is a substituted Ci-C 20 alkyl C 2 -C 20 alkynyl that includes an aminoalkylaminocarbonyl substituent, for example, R 2 is a substituted Ci-C 20 alkyl ethynyl that includes an aminoalkylaminocarbonyl substituent.
  • R 2 is C ⁇ C-(CH 2 ) q -CONH(CH 2 ) r -NH 2 , wherein q and r are independently 1 to 6, more particularly q is 2, 3, or 4. In any of these embodiments, r is 2 to 6, i.e., 2, 3, 4, 5, or 6.
  • Such compounds can be prepared by any suitable method, for example, as shown in Figure 1.
  • a functionalized chain at the 2-position of the adenine ring was formed by reaction of the protected 2-iodo derivative such as compound 18 with (PPh 3 ) 2 PdCl 2 , CuI, and the appropriate acetylene.
  • the deprotection of phosphate groups in derivatives 19 - 21 in strong acid provides the ester derivatives 4 - 6.
  • Each of the ester derivatives was hydrolyzed to yield the corresponding carboxylic acid 7 - 9.
  • ester group was directly aminolyzed using 1 ,2-diaminoethane, 1,3- diaminopropane or 1,4-diaminobutane to provide, after deprotection with TFA, amine congeners 10 - 15.
  • R 2 is a substituted Ci-C 20 alkyl C 2 -C 20 alkynyl that includes an arylalkylaminocarbonyl which is optionally substituted at the aryl, for example, a substituted Ci-C 2O alkyl ethynyl that includes an arylalkylaminocarbonyl which is substituted at the aryl.
  • R 2 is C ⁇ C-(CH 2 ) v -CONH(CH 2 ) w -(4-SO 2 F)Ph, wherein v and w are independently 1 to 6, preferably 2, 3, or 4, and "Ph" stands for phenyl.
  • Figure 3 illustrates a method of preparing compound 18 in accordance with an embodiment of the invention, which involves the condensation of phosphate-unprotected carboxylic acid 7 with the corresponding amine (containing the preformed arylsulfonyl fluoride) in DMF in presence of HATU and DIEA (Scheme 3). The presence of the free phosphate groups did not interfere in this coupling reaction.
  • R 2 is a substituted or unsubstituted heterocyclyl alkyl C 2 -C 20 alkynyl, for example, an unsubstituted heterocyclyl alkyl C 2 -C 20 alkynyl, particularly, an unsubstituted heterocyclyl alkyl ethynyl.
  • R 2 is C ⁇ C-(CH 2 ) x -Hetero-(CH 2 ) y -NHCO-alkyl, wherein "Hetero” stands for a heterocyclic moiety and x and y are independently 1 to 6, for example, R 2 is C ⁇ C-(CH 2 ) X - triazole-(CH 2 ) y -NHCO-alkyl, or R 2 is C ⁇ C-(CH 2 ) x -triazole-(CH 2 ) y -NHCO-CH 3 , wherein x is 2, 3, 4, or 5 and y is 2 or 3.
  • the triazole ring can be produced by the Click chemistry, i.e., by reacting an azido compound with an alkynyl end of the compound of the invention.
  • Examples of pharmaceutically acceptable salts for use in the present inventive pharmaceutical composition include those derived from mineral acids, such as hydrochloric, hydrobromic, phosphoric, metaphosphoric, nitric and sulphuric acids, and organic acids, such as tartaric, acetic, citric, malic, lactic, fumaric, benzoic, glycolic, gluconic, succinic, maleic and arylsulfonic, for example, benzenesulfonic and p-toluenesulfonic, acids.
  • the invention also provides a conjugate comprising at least one conjugant and a a P2Yi receptor antagonist compound or salt as described above, wherein the conjugant is covalently linked to the compound or salt.
  • the at least one conjugant is selected from the group consisting of a label, dye, reporter group, and a dendrimer.
  • the conjugate can include, in addition to a P2Yi receptor antagonist compound or salt, a dye molecule and/or a reporter molecule.
  • the label, dye, reporter group, or dendrimer is covalently linked to R 2 of the compound or salt.
  • R 2 is a substituted Ci-C 2O alkyl C 2 -C 20 alkynyl that includes an aminoalkylaminocarbonyl
  • the amino end is linked to a label, dye, reporter group and/or a dendrimer.
  • R 2 is a substituted C]-C 20 alkyl ethynyl that includes an aminoalkylaminocarbonyl
  • the amino end is linked to a label, dye, reporter group and/or a dendrimer.
  • the conjugate comprises a P2Yi receptor antagonist wherein R 2 is a substituted Ci-C 2O alkyl ethynyl that includes an aminoalkylaminocarbonyl whose amino end is linked to a dye, for example, R 2 is linked to a dye, such as in C ⁇ C-(CH 2 ) q -CONH(CH 2 ) r -NH-dye, wherein q and r are independently 1 to 6.
  • Figure 2 illustrates a method of preparing conjugates 16 and 17 wherein the dye biotin is covalently linked to the P2Yi receptor antagonist.
  • Ligands intended for irreversible receptor binding and avidin complexation were synthesized by attaching an appropriate prosthetic group to the functionalized chains at the C2 position.
  • Biotin conjugates of varying length 16 and 17 were synthesized from the protected intermediate amine 23 (Scheme 2), which was the precursor of amine congener 11.
  • the biotin conjugate 16 was prepared by a HATU coupling with biotin, and the extended chain analogue 17 was prepared from the corresponding biotin- ⁇ -aminocaproyl active ester. Finally, the phosphate groups were deprotected by reaction with TFA.
  • R 2 of the P2Yi receptor antagonist is a substituted C]-C 20 alkyl C 2 -C 20 alkynyl that includes an aminoalkylcarbonylaminoalkylaminocarbonyl whose amino end is linked to a label, dye, reporter group, and/or a dendrimer
  • R 2 is a substituted Ci-C 20 alkyl ethynyl that includes an aminoalkylcarbonylaminoalkylaminocarbonyl whose amino end is linked to a label, dye, reporter group, and/or a dendrimer
  • R 2 is a substituted Ci-C 20 alkyl ethynyl that includes an aminoalkylcarbonylaminoalkylaminocarbonyl group whose amino end is linked to a dye.
  • the conjugate includes a covalently linked dye, e.g., where R 2 is C ⁇ C-(CH 2 ) s -CONH(CH 2 ) t -NHCO(CH 2 ) u -NH-dye, wherein s, t, and u are independently 1 to 6.
  • the invention provides a compound or salt, wherein the compound is of the formula:
  • R is selected from the group consisting of:
  • n 2 to 5.
  • the dendrimer described above can further include one or more covalently bonded ligands that are agonists or antagonists of a receptor of the GPCR superfamily, other than the P2Yi receptor antagonists described above.
  • the GPCR family receptors include the four subtypes of adenosine receptors and eight subtypes of purinergic receptors, e.g., nucleotide P2Y receptors, and the muscarinic receptors.
  • the ligand is a functionalized congener of an agonist or antagonist of an adenosine receptor selected from the group consisting of Ai, A 2A , A 2B , and A 3 adenosine receptors, the P2Yi 4 receptors, and the Mi muscarinic receptors.
  • Other purinergic receptors include P2Y 2 , P2Y 4 , P2Y 6 , and P2Yn receptors.
  • the P2Y]-mediated response is associated with the initial shape change and rapid aggregation, and the P2Yi 2 receptor is associated with amplification of the aggregation.
  • P2Yj 2 receptor antagonists used extensively as antithrombotic agents, and new antagonists are under development. Shalito, L, et al., Am J. Ther. 2009, Feb 28. PMID: 19262362.
  • the conjugate comprises a dendrimer that further includes a covalently bonded adenosine receptor agonist, e.g., an A 2A receptor agonist.
  • adenosine receptor agonist can be included, for example, one or more selected from the group consisting of 2-[4-(2- aminoethylaminocarbonylethyl)phenylethylamino]-5'-N-ethylcarboxamidoadenosine (APEC), 2-[4-(2-carboxylethyl)phenylethylamino]-5'-N-ethylcarboxamidoadenosine (CGS21680), adenosine 5'-N-ethylcarboxamide (NECA), (2-[2-(4-chlorophenyl)-ethoxy]- adenosine) (MRE 9004), N 6 -[2-(3,5-
  • the conjugate includes a dendrimer which is covalently linked to a functionalized congener of an A 2A adenosine receptor agonist, wherein the functionalized congener has a purine nucleoside moiety and a functional group at the 2-position of the purine nucleoside moiety, wherein the functional group has the formula (II):
  • Examples of functionalized congener of an A 2A receptor agonist include one of the following formulas (Ha)-(IIe):
  • R' is Ci-C 4 alkyl
  • the conjugate includes a dendrimer which is covalently linked to a functionalized congener of an P2Yi 2 receptor antagonist, for example, a thienopyridine such as clopidrogel, ticlopidine, or prasugrel, or a direct and reversible P2Yj 2 receptor antagonist such as cangrelor or A2D6140.
  • a functionalized congener of an P2Yi 2 receptor antagonist for example, a thienopyridine such as clopidrogel, ticlopidine, or prasugrel
  • a direct and reversible P2Yj 2 receptor antagonist such as cangrelor or A2D6140.
  • the compounds and conjugates of the invention can be co-administered with a P2Yi 2 receptor antagonist, for example, a thienopyridine such as clopidrogel, ticlopidine, or prasugrel, or a direct and reversible P2Yi 2 receptor antagonist such as cangrelor or A2D6140.
  • a P2Yi 2 receptor antagonist for example, a thienopyridine such as clopidrogel, ticlopidine, or prasugrel, or a direct and reversible P2Yi 2 receptor antagonist such as cangrelor or A2D6140.
  • Dendrimers are known materials and are classified as polymers. Sajita, J., et al., Crit. Rev. Ther. Drug Carrier Syst, 2007, 24(3): 257-306; Yang, H., et al., J. Biomater Sd Poly Ed., 2006, 17(1-2): 3-19. Dendrimers are made from branched monomers through the iterative organic synthesis by adding one layer (i.e., generation) at each step to provide a perfectly symmetrical structure. The solution conformation of higher generation dendrimers may closely mimic the size and shape of a protein.
  • dendrimers possess favorable characteristics: structural integrity, control of component functional groups-and their physical properties-by chemical synthesis, feasibility to conjugate multiple functional units at the peripheries and interiors, and a low enzymatic degradation rate.
  • Dendrimers possess numerous chain end groups, which can serve as multivalent binding sites for interaction with biological receptors and cell surfaces in the construction of targeted drug delivery systems.
  • Dendritic architecture is characterized by unique properties for drug delivery such as structural uniformity, high purity, efficient membrane transport, high drug payload, targeting potential, and good colloidal, biological, and shelf stability.
  • the dendrimer can be of any suitable generation, e.g., from 2 to 10 or 2 to 6, or more, including fractional generations, particularly 2 to 8, e.g., 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, or 7.5.
  • the dendrimer can be anionic or cationic.
  • the half generations are carboxyl terminated and full generations are amine terminated.
  • the conjugate of the invention can include any suitable dendrimer, particularly a poly(amidoamine) (PAMAM) dendrimer.
  • PAMAM poly(amidoamine)
  • dendrimers include amine terminated dendrimers, polyester dendrimers, citric acid dendrimers, arginine dendrimers, and carbohydrate dendrimers.
  • the conjugate of the invention can contain any suitable degree of loading of the agonist or antagonist or both, e.g., a degree of loading greater than about 0.1%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or more; or 100% or less, about 95% or less, about 85% or less, about 75% or less, about 65% or less, about 55% or less, about 45% or less, about 35% or less, about 25% or less, about 15% or less, or about 5% or less, for example, about 1% to about 99%, about 10% to about 90%, about 20% to about 80%, about 30% to about 70%, or about 40% to about 60%, of the theoretical capacity of the dendrimer.
  • a degree of loading greater than about 0.1%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or more; or 100% or less, about 95% or less, about 85% or less, about 75% or less, about 65% or less
  • the conjugate of the invention can optionally include a covalently bonded marker, e.g., a reporter group, a dye or a fluorescent marker, e.g., biotin, Rhodamine Green, Cy5, Evo30, Gnothis blue 1, Gnothis blue 2, Gnothis blue 3, and Atto655; see, e.g., Giller, G., et al., Nucleic Acids Research, 2003, 31, 2630-2635.
  • fluorescent probes are BODIPY and FITC.
  • Cy3B is an example of a cyanine dye.
  • the conjugates of the invention can optionally include one more surface modifying moieties to modify one or more of the surface properties of the conjugate or a group that protects the surface functional groups such as amine or carboxylic functional groups.
  • the surface modifying moiety can be an amine protecting group.
  • An example of an amine protecting group is Ci-C 6 alkyl carbonyl, preferably CpC 3 alkyl carbonyl, wherein the alkyl group is unsubstituted or substituted with a substituent selected from the group consisting Of Cj-C 6 alkoxy, hydroxy, halo, nitro, cyano, amino, amido, and mercapto.
  • the surface modifying moiety comprises a hydrophilic group, e.g., a polyethylene glycol moiety.
  • the polyethylene glycol moiety can be linked to the dendrimer through any suitable bond, e.g., amide, hydrazide, ether, urethane, urea, thiourea, ester, carbonate, carbamate, hydrazone, carbazone, secondary amine, tertiary amine, and quaternary amine.
  • the amine or carboxyl ends of a dendrimer could also be covalently linked to amino acids, peptides, nucleic acids, glycosides, and one or more small molecules.
  • the invention further provides a pharmaceutical composition
  • a pharmaceutical composition comprising a compound of formula I described above or salt thereof , or a conjugate thereof, and a pharmaceutically acceptable carrier.
  • the invention further provides a method of preventing or treating a disease or condition that is preventable or treatable by antagonizing a P2Yj receptor of an animal comprising administering the animal an effective amount of a compound or salt or conjugate described above.
  • diseases or condition preventable or treatable by antagonizing a P2Y] receptor of an animal are arthrosclerosis, platelet aggregation, and thrombosis.
  • the pharmaceutically acceptable carrier can be any of those conventionally used and is limited only by chemico-physical considerations, such as solubility and lack of reactivity with the compound, and by the route of administration.
  • the compounds, salts, or conjugates of the present invention can be formulated as inclusion complexes, such as cyclodextrin inclusion complexes, or liposomes.
  • inclusion complexes such as cyclodextrin inclusion complexes, or liposomes.
  • pharmaceutically acceptable carriers described herein for example, vehicles, adjuvants, excipients, or diluents, are well known to those who are skilled in the art and are readily available to the public. It is preferred that the pharmaceutically acceptable carrier be one which is chemically inert to the active compounds and one which has no detrimental side effects or toxicity under the conditions of use.
  • compositions of the present invention are merely exemplary and are in no way limiting.
  • Formulations suitable for oral administration can consist of (a) liquid solutions, such as an effective amount of the compound, salt, or conjugate dissolved in diluents, such as water, saline, or orange juice; (b) capsules, sachets, tablets, lozenges, and troches, each containing a predetermined amount of the active ingredient, as solids or granules; (c) powders; (d) suspensions in an appropriate liquid; and (e) suitable emulsions.
  • Liquid formulations may include diluents, such as water and alcohols, for example, ethanol, benzyl alcohol, and the polyethylene alcohols, either with or without the addition of a pharmaceutically acceptable surfactant, suspending agent, or emulsifying agent.
  • diluents such as water and alcohols, for example, ethanol, benzyl alcohol, and the polyethylene alcohols, either with or without the addition of a pharmaceutically acceptable surfactant, suspending agent, or emulsifying agent.
  • Capsule forms can be of the ordinary hard- or soft-shelled gelatin type containing, for example, surfactants, lubricants, and inert fillers, such as lactose, sucrose, calcium phosphate, and cornstarch.
  • Tablet forms can include one or more of lactose, sucrose, mannitol, corn starch, potato starch, alginic acid, microcrystalline cellulose, acacia, gelatin, guar gum, colloidal silicon dioxide, croscarmellose sodium, talc, magnesium stearate, calcium stearate, zinc stearate, stearic acid, and other excipients, colorants, diluents, buffering agents, disintegrating agents, moistening agents, preservatives, flavoring agents, and pharmacologically compatible carriers.
  • Lozenge forms can comprise the active ingredient in a flavor, usually sucrose and acacia or tragacanth, as well as pastilles comprising the active ingredient in an inert base, such as gelatin and glycerin, or sucrose and acacia, emulsions, gels, and the like containing, in addition to the active ingredient, such carriers as are known in the art.
  • the compounds, salts, or conjugates of the present invention alone or in combination with other suitable components, can be made into aerosol formulations to be administered via inhalation. These aerosol formulations can be placed into pressurized acceptable propellants, such as dichlorodifluoromethane, propane, nitrogen, and the like. They also may be formulated as pharmaceuticals for non-pressured preparations, such as in a nebulizer or an atomizer.
  • Formulations suitable for parenteral administration include aqueous and nonaqueous, isotonic sterile injection solutions, which can contain anti-oxidants, buffers, bacteriostats, and solutes that render the formulation isotonic with the blood of the intended recipient, and aqueous and non-aqueous sterile suspensions that can include suspending agents, solubilizers, thickening agents, stabilizers, and preservatives.
  • the compound, salt, or conjugate can be administered in a physiologically acceptable diluent in a pharmaceutical carrier, such as a sterile liquid or mixture of liquids, including water, saline, aqueous dextrose and related sugar solutions, an alcohol, such as ethanol, isopropanol, or hexadecyl alcohol, glycols, such as propylene glycol or polyethylene glycol, glycerol ketals, such as 2,2- dimethyl-l,3-dioxolane-4-methanol, ethers, such as poly(ethyleneglycol) 400, an oil, a fatty acid, a fatty acid ester or glyceride, or an acetylated fatty acid glyceride with or without the addition of a pharmaceutically acceptable surfactant, such as a soap or a detergent, suspending agent, such as pectin, carbomers, methylcellulose, hydroxypropylmethylcellulose, or carboxymethylcellulose, or emuls
  • Oils which can be used in parenteral formulations include petroleum, animal, vegetable, or synthetic oils. Specific examples of oils include peanut, soybean, sesame, cottonseed, corn, olive, petrolatum, and mineral. Suitable fatty acids for use in parenteral formulations include oleic acid, stearic acid, and isostearic acid. Ethyl oleate and isopropyl myristate are examples of suitable fatty acid esters.
  • Suitable soaps for use in parenteral formulations include fatty alkali metal, ammonium, and triethanolamine salts
  • suitable detergents include (a) cationic detergents such as, for example, dimethyl dialkyl ammonium halides, and alkyl pyridinium halides, (b) anionic detergents such as, for example, alkyl, aryl, and olefin sulfonates, alkyl, olefin, ether, and monoglyceride sulfates, and sulfosuccinates, (c) nonionic detergents such as, for example, fatty amine oxides, fatty acid alkanolamides, and polyoxyethylene-polypropylene copolymers, (d) amphoteric detergents such as, for example, alkyl-beta-aminopropionates, and 2-alkyl-imidazoline quaternary ammonium salts, and (3) mixtures thereof.
  • the parenteral formulations will typically contain from about 0.5 to about 25% by weight of the compound, salt, or conjugate in solution. Suitable preservatives and buffers can be used in such formulations. In order to minimize or eliminate irritation at the site of injection, such compositions may contain one or more nonionic surfactants having a hydrophile-lipophile balance (HLB) of from about 12 to about 17. The quantity of surfactant in such formulations ranges from about 5 to about 15% by weight. Suitable surfactants include polyethylene sorbitan fatty acid esters, such as sorbitan monooleate and the high molecular weight adducts of ethylene oxide with a hydrophobic base, formed by the condensation of propylene oxide with propylene glycol.
  • HLB hydrophile-lipophile balance
  • parenteral formulations can be presented in unit-dose or multi-dose sealed containers, such as ampoules and vials, and can be stored in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example, water, for injections, immediately prior to use.
  • sterile liquid carrier for example, water
  • Extemporaneous injection solutions and suspensions can be prepared from sterile powders, granules, and tablets of the kind previously described.
  • the compounds, salts, or conjugates of the present invention may be made into injectable formulations.
  • the requirements for effective pharmaceutical carriers for injectable compositions are well known to those of ordinary skill in the art. See Pharmaceutics and Pharmacy Practice, J. B. Lippincott Co., Philadelphia, Pa., Banker and Chalmers, eds., pages 238-250 (1982), and ASHP Handbook on Injectable Drugs, Toissel, 4th ed., pages 622-630 (1986).
  • the compounds, salts, or conjugates of the present invention may be made into suppositories by mixing with a variety of bases, such as emulsifying bases or water-soluble bases.
  • bases such as emulsifying bases or water-soluble bases.
  • Formulations suitable for vaginal administration may be presented as pessaries, tampons, creams, gels, pastes, foams, or spray formulas containing, in addition to the active ingredient, such carriers as are known in the art to be appropriate.
  • Examples of conditions or diseases preventable or treatable by an A 2A adenosine receptor agonists include sleep disorders, respiratory disorders, reperfusion injury, platelet aggregation, thrombosis, hypertension, kidney ischemia, rheumatoid arthritis, inflammation, wound healing, sepsis, and sepsis syndrome, particularly inflammation, platelet aggregation, and thrombosis.
  • Examples of A], A 2A , A 2B , and/or A 3 adenosine receptor agonists can be found in the PCT/US09/38026; WO 2006/031505 Al and WO 01/51490 Al, WO 2008/006369; WO 2006/128159; WO 2006/113204; US Patents 7,199,127; 7,087,589; 6,586,413; 6,376,521; 6,316,423; 6,211,165; 5,620,676; 5,284,834; 5,280,015; 5,840,728; 5,688,774; and 5,773,423; and US 2007/0232626 Al and US 2007/0265223 Al; the disclosures of which are incorporated by reference in their entireties.
  • animals include mammals, e.g., the order Rodentia, such as mice, and the order Logomorpha, such as rabbits. It is preferred that the mammals are from the order Carnivora, including Felines (cats) and Canines (dogs). It is more preferred that the mammals are from the order Artiodactyla, including Bovines (cows) and Swines (pigs) or of the order Perssodactyla, including Equines (horses). It is most preferred that the mammals are of the order Primates, Ceboids, or Simioids (monkeys) or of the order Anthropoids (humans and apes). An especially preferred mammal is the human.
  • the invention further provides a diagnostic method for determining a treatment of a patient for a possible antagonist of the P2Y] receptor, comprising:
  • determining a treatment regimen comprising administering an antagonist of the receptor whose expression was higher in the patient than that of the normal population.
  • the present invention also provides a method for blocking P2Yj receptors in a cell comprising contacting the cell with an effective amount of one or more of the inventive compounds, salts, or conjugates thereof.
  • the invention further provides a method for blocking P2Yi receptors in a cell and activating an A 2A adenosine receptor in the cell, comprising contacting the cell with an effective amount of one or more of the inventive compounds, salts, or conjugates thereof.
  • the contacting can be in vitro or in vivo. When the contacting is done in vitro, the contacting can be done by any suitable method, many of which are known in the art.
  • the cell can be provided in a culture medium and the inventive compound introduced into the culture medium per se, or as a solution of the compound in an appropriate solvent.
  • the invention further provides a method of preparing a dendrimer conjugate comprising a P2Yi receptor antagonist and a dendrimer that is covalently linked to the P2Yi receptor antagonist through a triazole moiety, comprising: (a) providing a dendrimer having an azido terminal groups;
  • the dendrimer conjugate further comprises an A 2A adenosine receptor agonist covalently linked to the dendrimer, which is prepared by the above method, which further comprises (b) reacting the dendrimer of (a), in any order, with a P2Yi receptor antagonist having a terminal acetylenic group and an A 2A adenosine receptor agonist, or a mixture thereof, in the presence of a copper (I) catalyst.
  • a suitable copper (I) catalyst is a combination of CuSO 4 and sodium ascorbate.
  • Figure 5 illustrates a reaction scheme where AcNHCH 2 CH 2 N 3 serves as a model for a dendrimer having an azido terminal group.
  • the model compound is reacted with a P2Yi antagonist that has been modified at the 2-position to include a terminal alkynyl group.
  • the terminal alkynyl group reacts with the azido terminal group on the model compound through Click chemistry to produce a model compound covalently linked to the antagonist through a triazole ring.
  • Click chemistry see US 2009/0069561A1 and Wu et al., Chem. Commun., 2005, 5775-5777 (2005).
  • Figure 6 illustrates a reaction scheme where an A 2 A adenosine receptor agonist 52 is covalently linked to a dendrimer, 43, which is followed by covalent linking of protected P2Yi receptor antagonists 54 - 57 modified to have alkynyl terminal groups at the 2-position.
  • the antagonist is covalently linked through the use of the Click chemistry.
  • the nucleotide derivatives were purified by HPLC before biological testing and the purity of each was shown to be >95%.
  • the mobile phases consisted of System A: linear gradient solvent system of CH 3 CN/triethyl ammonium acetate from 5/95 to 60/40 in 20 min, flow rate 1.0 mL/min;
  • System B linear gradient solvent system of CH 3 CN/tetrabutyl ammonium phosphate from 20/80 to 60/40 in 20 min, flow rate 1.0 mL/min.
  • HRMS high resolution mass spectroscopic
  • the crude product was purified by HPLC with a Luna 5 1 RP-C18(2) semipreparative column (250 x 10.0 mm; Phenomenex, Torrance, CA) and using the following conditions: flow rate of 2 mL/min; 10 mM triethylammonium acetate (TEAA)- CH 3 CN from 100:0 to 60:40 in 30 min (and isolated in the triethylammonium salt form).
  • TEAA triethylammonium acetate
  • the crude was purified by HPLC with a Luna 5 1 RP-C 18(2) semipreparative column (250 x 10.0 mm; Phenomenex, Torrance, CA) and using the following conditions: flow rate of 2 mL/min; 10 mM triethylammonium acetate (TEAA)- CH 3 CN from 100:0 to 60:40 in 30 min (and isolated in the triethylammonium salt form).
  • TEAA triethylammonium acetate
  • the crude was purified the crude was purified by HPLC with a Luna 5 1 RP-C 18(2) semipreparative column (250 x 10.0 mm; Phenomenex, Torrance, CA) and using the following conditions: flow rate of 2 mL/min; 10 mM triethylammonium acetate (TEAA)-CH 3 CN from 100:0 to 60:40 in 30 min (and isolated in the triethylammonium salt form).
  • TEAA triethylammonium acetate
  • This Example illustrates a method of preparing conjugates comprising a P2Y1 receptor antagonist that is covalently linked to a dye in accordance with an embodiment of the invention.
  • the crude was purified by HPLC with a Luna 5 1 RP-C18(2) semipreparative column (250 x 10.0 mm; Phenomenex, Torrance, CA) and using the following conditions: flow rate of 2 mL/min; 10 mM triethylammonium acetate (TEAA)-CH 3 CN from 100:0 to 60:40 in 30 min to give to give 0.6 mg of 18 (25 %, isolated in the triethylammonium salt form).
  • TEAA triethylammonium acetate
  • This Example illustrates the binding affinities of some embodiments of the compounds of the invention.
  • Pharmacology 2-MeSADP was purchased from Sigma (St. Louis, MO). P2Yj receptor binding was measured at the human P2Yi receptor stably expressed in 132 INl human astrocytoma cells as previously described. Harden, T. K., et al., Biochem. J. 1988, 252, 583-593; Boyer, J. L., et al., J. Biol. Chem. 1989, 264, 884-890.
  • the affinities of bisphosphate analogues for the human P2Yj receptor were directly determined by using [ 125 I]MRS2500 ((l'R,2'S,4'S,5'S)-4-(2-iodo-6-methylamino-purin-9-yl)-l-[(phosphato)- methyl]-2-(phosphato)-bicyclo[3.1.0]hexane) in a radioligand binding assay. Binding and functional parameters were estimated using GraphPAD Prism software (GraphPAD, San Diego, CA). The results obtained are set forth in Table 1.
  • the three terminal ester derivatives 4 - 6 displayed micromolar affinity at the human P2Y1 receptor.
  • the shortest analogue 4 having two methylenes in the alkynyl chain, was slightly more potent than the longer analogues.
  • the carboxylic acid derivatives 7 - 9 were more potent than the corresponding ester derivatives. There was a clearly favored analogue among the carboxylic acid derivatives 7 - 9, i.e., compound 7 with a K, of 23 nM.
  • a comparison of the hexynyl derivative 3 with the corresponding carboxylic acid derivative 7 indicates a gain of 1.4 log units in affinity due to the presence of the terminal carboxylate group.
  • Variation of the chain length and position of an intermediate amide group revealed high affinity of a carboxylic congener 7 (Kj 23 nM) and an extended amine congener 14 (Kj 132 nM), both of which contained a 2-(l-pentynoyl) group.

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Abstract

Cette invention concerne des antagonistes des récepteurs P2Y1 de formule (I); R1-R3 pour la formule (I) étant tels que définis dans la description, ainsi que des compositions pharmaceutiques les contenant, et l'utilisation de ces antagonistes pour traiter une maladie ou une affection que l'on peut faire reculer en s'opposant aux effets d'un récepteur P2Y1, par exemple, l'agrégation des plaquettes, l'athérosclérose ou la thrombose. L'invention décrit également des conjugués comprenant un antagoniste P2Y1 lié par covalence à un partenaire de conjugaison tel qu'un dendrimère.
PCT/US2009/047204 2008-06-13 2009-06-12 Antagonistes des récepteurs p2y<sb>1</sb> Ceased WO2009152431A1 (fr)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1316559A1 (fr) * 2000-09-08 2003-06-04 Toa Eiyo Ltd. D riv s d'ad nosine et leurs applications
EP1733742A1 (fr) * 2005-06-17 2006-12-20 Universiteit Utrecht Holding B.V. Dendrimères substitués de façon multivalente par des groupes actifs

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1316559A1 (fr) * 2000-09-08 2003-06-04 Toa Eiyo Ltd. D riv s d'ad nosine et leurs applications
EP1733742A1 (fr) * 2005-06-17 2006-12-20 Universiteit Utrecht Holding B.V. Dendrimères substitués de façon multivalente par des groupes actifs

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Title
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CRISTALLI G ET AL: "Effects of 5'-phosphate derivatives of 2-hexynyl adenosine and 2-phenylethynyl adenosine on responses of human platelets mediated by P2Y receptors", JOURNAL OF MEDICINAL CHEMISTRY, vol. 48, no. 8, 21 April 2005 (2005-04-21), pages 2763 - 2766, XP009123000, ISSN: 0022-2623, DOI: 10.1021/jm0493562 *
DONDONI A: "Triazole: the keystone in glycosylated molecular architectures constructed by a click reaction", CHEMISTRY, AN ASIAN JOURNAL, vol. 2, no. 6, 4 June 2007 (2007-06-04), pages 700 - 708, XP009120242, ISSN: 1861-4728, DOI: 10.1002/asia.200700015 *
GILLER G ET AL: "Incorporation of reporter molecule-labeled nucleotides by DNA polymerases. I. Chemical synthesis of various reporter group-labeled 2'-deoxyribonucleoside-5'-triphosphates", NUCLEIC ACIDS RESEARCH, vol. 31, no. 10, 15 May 2003 (2003-05-15), pages 2630 - 2635, XP009122995, ISSN: 1362-4962, DOI: 10.1093/nar/gkg370 *
KIM H S ET AL: "2-substitution of adenine nucleotide analogues containing a bicyclo[3.1.0]hexane ring system locked in a northern conformation: Enhanced potency as P2Y1 receptor antagonists", JOURNAL OF MEDICINAL CHEMISTRY, vol. 46, no. 23, 14 October 2003 (2003-10-14), pages 4974 - 4987, XP002388426, ISSN: 0022-2623, DOI: 10.1021/jm030127+ *
MATHIEU R ET AL: "Synthesis and biological activity of 2-alkylated deoxyadenosine bisphosphate derivatives as P2Y(1) receptor antagonists", BIOORGANIC & MEDICINAL CHEMISTRY, vol. 12, no. 7, 1 April 2004 (2004-04-01), pages 1769 - 1779, XP009122991, ISSN: 0968-0896, DOI: 10.1016/j.bmc.2003.12.041 *
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