Polymorphisms in the human gene for cytochrome p450 polypeptide 2c8 and their use in diagnostic and therapeutic applications

Abstract

The present invention relates to a polymorphic CYP2C8-polynucleotide. Moreover, the invention relates to genes or vectors comprising the polynucleotides of the invention and to a host cell genetically engineered with the polynucleotide or gene of the invention. Further, the invention relates to methods for producing molecular variant polypeptides or fragments thereof, methods for producing cells capable of expressing a molecular variant polypeptide and to a polypeptide or fragment thereof encoded by the polynucleotide or the gene of the invention or which is obtainable by the method or from the cells produced by the method of the invention. Furthermore, the invention relates to an antibody which binds specifically the polypeptide of the invention. Moreover, the invention relates to a transgenic non-human animal. The invention also relates to a solid support comprising one or a plurality of the above mentioned polynucleotides, genes, vectors, polypeptides, antibodies or host cells. Furthermore, methods of identifying a polymorphism, identifying and obtaining a prodrug or drug or an inhibitor are also encompassed by the present invention. In addition, the invention relates to methods for producing of a pharmaceutical composition and to methods of diagnosing a disease. Further, the invention relates to a method of detection of the polynucleotide of the invention. Furthermore, comprised by the present invention are a diagnostic and a pharmaceutical composition. Even more, the invention relates to uses of the polynucleotides, genes, vectors, polypeptides or antibodies of the invention. Finally, the invention relates to a diagnostic kit.

1 . A polynucleotide comprising a polynucleotide selected from the group consisting of:
 (a) a polynucleotide having the nucleic acid sequence of SEQ ID NO: 54, 57, 60, 63, 66, 69, 72, 75, 78, 81, 84, 87, 90, 93, 96, 99, 102, 105, 108, 111, 114, 117, 120, 123, 126, 129, 132, 135, 138, 141, 144, 147, 150, 153, 156, 159, 162, 165, 168, 171, 174, 177, 180, 183, 183, 189, 192, 195, 198, 201, 210, 213, 216, 219, 222, 225, 228, 231, 234, 237, 240, 243, 246, 249, 252, 255, 258, 261, 264, 267, 270, 273, 276, 279, 282, 285, 288, 291, 306, 309, 318, 321, 324, 327, 330, 333, 342, 345, 348, 351, 354, 357, 360, 363, 366, 369, 384, 387, 390, 393, 396 or 399;   (b) a polynucleotide encoding a polypeptide having the amino acid sequence of SEQ ID NO: 6, 8, 10, 12, 18, 377, 379 or 381;   (c) a polynucleotide capable of hybridizing to a CYP2C8 gene, wherein said polynucleotide is having at a position corresponding to position 411, 560, 713, 817, 824, 831, 879, 886, 1058, 1627, 1668, 1767, 1887, 1905 or 1952 (GenBank accession No: AF136830.1), at a position corresponding to position 171 or 258 (GenBank accession No: AF136832.1), at a position corresponding to position 122, 150, 182, 334, 339 or 378 (GenBank accession No: AF136833.1), at a position corresponding to position 162, 163, 243 (GenBank accession No: AF136834.2) or at position 583 (GenBank accession No: NM — 000770.1), at a position corresponding to position 13 or 180 (GenBank accession No: AF136835.1), at a position corresponding to position 116, 132, 172 or 189 (GenBank accession No: AF136836.1), at a position corresponding to position 42 or 101 (GenBank accession No: AF136837.1), at a position corresponding to position 309 (GenBank accession No: AF136838.1), at a position corresponding to position 1135 (GenBank accession No: NM — 000770.1), at a position corresponding to position 232 (GenBank accession No: AF136840.1), at a position corresponding to position 206 (GenBank accession No: AF136842.1), at a position corresponding to position 30, 87, 167, 197, 212, 221, 255 or 271 (GenBank accession No: AF136843.1), at a position corresponding to position 118 (GenBank accession No: AF136844.1), at a position corresponding to position 44 (GenBank accession No: AF136845.1) of the cytochrome 2C8 gene (GenBank accession No: GI: 13787189) a nucleotide substitution, at a position corresponding to position 306 to 307, 1271 to 1273 or 1397 to 1398 of the CYP2C8 gene (GenBank accession No: AF136830.1), at a position corresponding to position 329 of the CYP2C8 gene (GenBank accession No: AF136833.1), at a position corresponding to position 87 of the CYP2C8 gene (GenBank accession No: AF136834.2) a deletion of one or more nucleotides or at a position corresponding to position 1785/1786 of the CYP2C8 gene (GenBank accession No: AF136830.1) or at a position corresponding to position 180/181 of the CYP2C8 gene (GenBank accession No: AF36833.1) an insertion of one or more nucleotides;   (d) a polynucleotide capable of hybridizing to a CYP2C8 gene, wherein said polynucleotide is having at a position corresponding to position 411, 817, 824, 831, 879, 1058, 1767 or 1887 of the CYP2C8 gene (GenBank accession No: AF136830.1) an A, at a position corresponding to position 560 or 1668 of the CYP2C8 gene (GenBank accession No: AF136830.1) a G, at a position corresponding to position 713 or 886 of the CYP2C8 gene (GenBank accession No: AF36830.1) a T, at a position corresponding to position 1627, 1905 or 1952 of the CYP2C8 gene (GenBank accession No: AF136830.1) a C, at a position corresponding to position 258 of the CYP2C8 gene (GenBank accession No: AF136832.1) a T, at a position corresponding to position 171 of the CYP2C8 gene (GenBank accession No: AF136832.1) a C, at a position corresponding to position 122, 150 or 334 of the CYP2C8 gene (GenBank accession No: AF136833.1) an A, at a position corresponding to position 182 or 378 of the CYP2C8 gene (GenBank accession No: AF136833.1) a C, at a position corresponding to position 162, 163, 243 [identical to position corresponding to position 583 of the CYP2C8 gene (GenBank accession No: NM — 000770.1) of the CYP2C8 gene (GenBank accession No: AF136834.2) an A, at a position corresponding to position 180 of the CYP2C8 gene (GenBank accession No: AF136835.1) an A, at a position corresponding to position 13 of the CYP2C8 gene (GenBank accession No: AF136835.1) a G, at a position corresponding to position 116 or 132 of the CYP2C8 gene (GenBank accession No: AF136836.1) a G, at a position corresponding to position 172 of the CYP2C8 gene (GenBank accession No: AF136836.1) a G, at a position corresponding to position 189 of the CYP2C8 gene (GenBank accession No: AF136836.1) a C, at a position corresponding to position 42 or 101 of the CYP2C8 gene (GenBank accession No: AF136837.1) a G, at a position corresponding to position 1135 of the CYP2C8 gene (GenBank accession No: GI: 13787189) an A, at a position corresponding to position 309 of the CYP2C8 gene (GenBank accession No: AF136838.1) a T, at a position corresponding to position 232 (GenBank accession No: 136840.1) a T, at a position corresponding to position 30 or 212 of the CYP2C8 gene (GenBank accession No: AF136843.1) a T, at a position corresponding to position 87 of the CYP2C8 gene (GenBank accession No: AF136843.1) a G, at a position corresponding to position 167 or 197 of the CYP2C8 gene (GenBank accession No: AF136843.1) an A, at a position corresponding to position 221, 255 or 271 of the CYP2C8 gene (GenBank accession No: AF136843.1) a C, at a position corresponding to position 118 of the CYP2C8 gene (GenBank accession No AF136844.1) an A, at a position corresponding to position 44 of the CYP2C8 gene (GenBank accession No: AF136845.1) a T;   (e) a polynucleotide encoding a molecular CYP2C8 variant polypeptide or fragment thereof, wherein said polypeptide comprises an ammo acid substitution at a position corresponding to any one of position 159, 181, 209, 244, 263, 274, 343 or 365 of the CYP2C8 polypeptide (GI: 13787189); and   (f) a polynucleotide encoding a molecular CYP2C8 variant polypeptide or fragment thereof, wherein said polypeptide comprises an amino acid substitution of T to P at position corresponding to position 159 (frameshift), V to I at a position corresponding to position 181, N to S at a position corresponding to position 209, I to V at a position corresponding to position 244, F to L at a position corresponding to position 263, E to Stop at a position corresponding to position 274, G to S at a position corresponding to position 365 or S to I at a position corresponding to position 343 of the CYP2C8 polypeptide (GenBank accession No: GI: 13787189).   
     
     
         2 . (canceled) 
     
     
         3 . (canceled) 
     
     
         4 . (canceled) 
     
     
         5 . (canceled) 
     
     
         6 . (canceled) 
     
     
         7 . (canceled) 
     
     
         8 . (canceled) 
     
     
         9 . (canceled) 
     
     
         10 . (canceled) 
     
     
         11 . A polypeptide or fragment thereof encoded by the polynucleotide of  claim 1 . 
     
     
         12 . An antibody which binds specifically to the polypeptide of  claim 11 . 
     
     
         13 . (canceled) 
     
     
         14 . (canceled) 
     
     
         15 . A transgenic non-human animal comprising the polynucleotide of  claim 1 . 
     
     
         16 . (canceled) 
     
     
         17 . A solid support comprising the polynucleotide of  claim 1  the polypeptide of  claim 11 , or the antibody of  claim 12  in immobilized form. 
     
     
         18 . (canceled) 
     
     
         19 . An in vitro method for identifying a single nucleotide polymorphism said method comprising the steps of:
 (a) isolating the polynucleotide of  claim 1  from a plurality of subgroups of individuals, wherein one subgroup has no prevalence for a CYP2C8 associated disease and at least one or more further subgroup(s) do have prevalence for a CYP2C8 associated disease; and   (b) identifying a single nucleotide polymorphism by comparing the nucleic acid sequence of said polynucleotide or said gene of said one subgroup having no prevalence for a CYP2C8 associated disease with said at least one or more further subgroup(s) having a prevalence for a CYP2C8 associated disease.   
     
     
         20 . A method for identifying and obtaining a pro-drug or a drug capable of modulating the activity of a molecular variant of a CYP2C8 polypeptide comprising the steps of:
 (a) contacting the polypeptide of  claim 11  in the presence of components capable of providing a detectable signal in response to drug activity with a compound to be screened for pro-drug or drug activity; and   (b) detecting the presence or absence of a signal or increase or decrease of a signal generated from the pro-drug or the drug activity, wherein the absence, presence, increase or decrease of the signal is indicative for a putative pro-drug or drug.   
     
     
         21 . A method for identifying and obtaining an inhibitor of the activity of a molecular variant of a CYP2C8 polypeptide comprising the steps of:
 (a) contacting the protein of  claim 11  in the presence of components capable of providing a detectable signal in response to drug activity with a compound to be screened for inhibiting activity; and   (b) detecting the presence or absence of a signal or increase or decrease of a signal generated from the inhibiting activity, wherein the absence or decrease of the signal is indicative for a putative inhibitor.   
     
     
         22 . (canceled) 
     
     
         23 . A method of identifying and obtaining a pro-drug or drug capable of modulating the activity of a molecular variant of a CYP2C8 polypeptide comprising the steps of:
 (a) contacting the polypeptide of  claim 11  with the first molecule known to be bound by a CYP2C8 polypeptide to form a first complex of said polypeptide and said first molecule;   (b) contacting said first complex with a compound to be screened, and   (c) measuring whether said compound displaces said first molecule from said first complex.   
     
     
         24 . A method of identifying and obtaining an inhibitor capable of modulating the activity of a molecular variant of a CYP2C8 polypeptide or its gene product comprising the steps of:
 (a) contacting the protein of  claim 11  with the first molecule known to be bound by a CYP2C8 polypeptide to form a first complex of said polypeptide and said first molecule;   (b) contacting said first complex with a compound to be screened, and   (c) measuring whether said compound displaces said first molecule from said first complex.   
     
     
         25 . (canceled) 
     
     
         26 . (canceled) 
     
     
         27 . (canceled) 
     
     
         28 . A method for the production of a pharmaceutical composition comprising the steps of the method of  claim 20  and the further step of formulating the compound identified and obtained or a derivative thereof in a pharmaceutical acceptable form. 
     
     
         29 . A method of diagnosing a disorder related to the presence of a molecular variant of a CYP2C8 gene or susceptibility to such a disorder comprising determining the presence of a polynucleotide of  claim 1  in a sample from a subject. 
     
     
         30 . (canceled) 
     
     
         31 . A method of diagnosing a disorder related to the presence of a molecular variant of a CYP2C8 gene or susceptibility to such a disorder comprising determining the presence of a polypeptide of  claim 11  in a sample from a subject. 
     
     
         32 . (canceled) 
     
     
         33 . (canceled) 
     
     
         34 . A method of detection of the polynucleotide of  claim 1  in a sample comprising the steps of
 (a) contacting the solid support comprising the polynucleotide with the sample under conditions allowing interaction of the polynucleotide with the immobilized targets on a solid support and 
 (b) determining the binding of said polynucleotide or said gene to said immobilized targets on a solid support. 
 
     
     
         35 . An in vitro method for diagnosing a disease comprising the steps of the method of  claim 34 , wherein binding of said polynucleotide or gene to said immobilized targets on said solid support is indicative for the presence or the absence of said disease or a prevalence for said disease. 
     
     
         36 . A diagnostic composition comprising the polynucleotide of  claim 1 , the polypeptide of  claim 11  or the antibody of  claim 12 . 
     
     
         37 . A pharmaceutical composition comprising the polynucleotide of  claim 1 , the polypeptide of  claim 11  or the antibody of  claim 12 . 
     
     
         38 . (canceled) 
     
     
         39 . (canceled) 
     
     
         40 . (canceled) 
     
     
         41 . A diagnostic kit for detection of a single nucleotide polymorphism comprising the polynucleotide of  claim 1 , the polypeptide of  claim 11 , the antibody of  claim 12 .