Identification and functional characterization of new potentially defective alleles of human CYP2C19

Abstract

CYP2C19 is a clinically important enzyme responsible for the metabolism of a number of therapeutic drugs, such as mephenytoin, omeprazole, diazepam, proguanil, propranolol and certain antidepressants. Genetic polymorphisms in this enzyme result in poor metabolizers of these drugs. There are racial differences in the incidence of the poor metabolizer trait, which represents 13-23% of Asians but only 3-5% of Caucasians. In this study, single nucleotide polymorphisms (SNPs) in CYP2C19 were identified by direct sequencing of genomic DNA from 92 individuals from three different racial groups of varied ethnic background, including Caucasians, Asians and blacks. Several new alleles were identified containing the coding changes Arg114 His (CYP2C19*9), Pro227 Leu (CYP2C19*10), Arg150 His (CYP2C19*11), stop491 Cys (CYP2C19*12), Arg410 Cys (CYP2C19*13), Leu17 Pro (CYP2C19*14) and Ile19 Leu (CYP2C19*15). When expressed in a bacterial cDNA expression system, CYP2C19*9 exhibited a modest decrease in the V(max) for 4'-hydroxylation of -mephenytoin, and no alteration in its affinity for reductase. CYP2C19*10 exhibited a dramatically higher K(m) and lower V(max) for mephenytoin. CYP2C19*12was unstable and expressed poorly in a bacterial cDNA expression system. Clinical studies will be required to confirm whether this allele is defective in vivo. CYP2C19*9, CYP2C19*10 and CYP2C19*12 all occurred in African-Americans, or individuals of African descent, and represent new potentially defective alleles of CYP2C19 which are predicted to alter risk of these populations to clinically important drugs.

Structural Characterization of Human Cytochrome P450 2C19

“…P450 2C19*1A differs from P450 2C19*1B due to a nonsynonymous single nucleotide polymorphism (refSNP rs3758581) that changes codon 331 for valine in the CYP2C19*1B gene to a codon for isoleucine, which is present in NCBI reference sequence for the CYP2C19*1A gene (NG_008384.1). The corresponding proteins do not appear to differ in their catalytic properties (28,29). A three-fragment ligation reaction was used to insert two fragments, generated by SacI/SphI and SphI/ PvuII digestion of pCW2C19*1B plasmid (27), into the pCW2C5LVdH expression vector (30) digested with SacI and PvuII.…”

Evaluation of Predictive Cyp2C19 Genotyping Assays Relative to Measured Phenotype in a South African Cohort

“…The allele was first described in Black Africans by Blaisdell et al [20], where the defining SNV, 55A>C (rs17882687), was found to be in linkage disequilibrium with 80161A>G (rs3758581).…”

“…To achieve this, additional alleles such as the decreased function CYP2C19*9 and *15 alleles were accounted for in this study. The former allele was of particular relevance, as it is known to be African-specific [20,21] and indeed contributed to the improved stratification of CYP2C19 phenotypes.…”

“…However once the AS was adjusted from 0.5 to 1.0 ( Although examining a greater number of alleles and modifying the AS system according to OME did improve the predictive ability of the genotyping assays, this system remains imperfect. The wide range of metabolic ratios within each of the predicted phenotypic groups, the overlap of 20 these groups, and the lack of good correlation ( Figure 1 & 2) reveals discrepancies in the predicted-measured phenotypes of CYP2C19. These discrepancies could be explained, in part,…”

“…The above-mentioned alleles have been prioritised based on observations in cohorts of predominantly European and Asian descent. This approach may not be sufficiently comprehensive in the context of African populations, and many African-specific alleles, such as CYP2C19*9 [20,21], may be important in this context.…”