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A New CYP3A5 Variant, CYP3A5*11, Is Shown to Be Defective in Nifedipine Metabolism in a Recombinant cDNA Expression System

Human Metabolism Section (S.-J.L., J.A.G.), Laboratory of Pharmacology and Chemistry, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, North Carolina; and Department of Clinical Chemistry (I.P.v.d.H., R.H.N.v.S.), Erasmus MC, Rotterdam, The Netherlands

A new CYP3A5 variant, CYP3A5*11, was found in a white European subject by DNA sequencing. The CYP3A5*11 allele contains a single nucleotide polymorphism (SNP) (g.3775A>G) in exon 2, which results in a Tyr53Cys substitution, and a g.6986A>G splice change, the latter SNP previously reported in the defective CYP3A5*3 allele. However, the CYP3A5*3 is not a null allele because this variant is associated with leaky splicing, resulting in small amounts of functional protein still being produced. Therefore, we constructed a cDNA coding for the newly identified CYP3A5.11 protein by site-directed mutagenesis, expressed it in Escherichia coli, and partially purified it. Whereas bacteria transformed with wild-type CYP3A5*1 cDNA expressed predominantly cytochrome P450 (P450), those transfected with CYP3A5*11 expressed a significant amount of denatured cytochrome P420 in addition to P450, suggesting the protein to be unstable. CYP3A5.11 exhibited a 38% decrease in the V_max for nifedipine metabolism, a 2.7-fold increase in the K_m, and a 4.4-fold decrease in the CL_int of nifedipine compared with CYP3A5.1. A polymerase chain reaction-restriction fragment length polymorphism genotyping procedure was developed and used to genotype DNA of 500 white individuals for CYP3A5*11. No additional examples of this allele were identified. In summary, individuals carrying the rare CYP3A5*11 allele are predicted to have lower metabolism of CYP3A5 substrates than individuals expressing CYP3A5*3.