| Summary: | Coding‐region variants of CYP2C9 can dramatically influence the pharmacokinetics and drug response of therapeutic agents, an important clinical consideration for commonly used CYP2C9 narrow therapeutic index substrates, including (S) ‐warfarin and phenytoin. In the Yup'ik Alaska Native people, novel CYP2C9 protein variants M1L, N218I, and P279T are expressed with higher frequencies than the well‐characterized CYP2C9*2 and CYP2C9*3 alleles. This study's objective was to determine the relative expression of these CYP2C9 variants in HepG2 cells and to characterize the function of the purified reconstituted enzymes expressed in E. coli , towards several CYP2C9 drug substrates. While mRNA expression of the CYP2C9 M1L, N218I, and P279T gene variants and the reference (wildtype) protein in HepG2 cells were similar, the protein was undetectable for the M1L variant. This is expected, due to disruption of the start codon in CYP2C9 M1L. His‐tagged CYP2C9 reference protein and N218I and P279T variants expressed well in E. coli and were highly purified after affinity chromatography. Following reconstitution with cytochrome P450 reductase and cytochrome b5, the N218I and P279T variants metabolized (S) ‐warfarin, but not ( R) ‐warfarin, to the principal 7‐ and 6‐hydroxy metabolites formed by the reference enzyme. Kinetic studies of (S) ‐warfarin metabolism demonstrate that the N218I variant has reduced function, with less than 40% of the catalytic efficiency of reference enzyme, while the P279T variant had similar catalytic efficiency to that of reference CYP2C9. Similar, although not identical concentration–rate profiles were obtained for phenytoin, flurbiprofen and naproxen metabolism. The potential of CYP2C9 M1L, N218I, and perhaps even P279T alleles to alter the pharmacokinetics of drugs metabolized by CYP2C9 may put carriers at risk of exacerbated therapeutic effects from drugs that rely predominantly on CYP2C9 for their metabolic clearance. Further clinical implications for the Alaska Native people include phenotypic ...
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