Functional Characterization of Low-Prevalence Missense Polymorphisms in the UDP-Glucuronosyltransferase 1A9 Gene

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The American Society for Pharmacology and Experimental Therapeutics

RESUMO

The UDP-glucuronosyltransferase (UGT) 1A9 has been shown to play an important role in the detoxification of several carcinogens and clearance of anticancer and pain medications. The goal of the present study was to identify novel polymorphisms in UGT1A9 and characterize their effect on glucuronidation activity. The UGT1A9 gene was analyzed by direct sequencing of buccal cell genomic DNA from 90 healthy subjects. In addition to a previously identified single nucleotide polymorphism (SNP) at codon 33 resulting in an amino acid substitution (Met>Thr), two low-prevalence (<0.02) novel missense SNPs at codons 167 (Val>Ala) and 183 (Cys>Gly) were identified and are present in both white and African-American subjects. Glucuronidation activity assays using HEK293 cell lines overexpressing wild-type or variant UGT1A9 demonstrated that the UGT1A933Thr and UGT1A9183Gly variants exhibited differential glucuronidation activities compared with wild-type UGT1A9, but this was substrate-dependent. The UGT1A9167Ala variant exhibited levels of activity similar to those of wild-type UGT1A9 for all substrates tested. Whereas the wild-type and UGT1A933Thr and UGT1A9167Ala variants formed homodimers as determined by Western blot analysis of native polyacrylamide gels, the UGT1A9183Gly variant was incapable of homodimerization. These results suggest that several low-prevalence missense polymorphisms exist for UGT1A9 and that two of these (M33T and C183G) are functional. These results also suggest that although Cys183 is necessary for UGT1A9 homodimerization, the lack of capacity for UGT1A9 homodimerization is not sufficient to eliminate UGT1A9 activity.

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