Novel human CYP2A6 alleles confound gene deletion analysis.

Cytochrome P450 (CYP) 2A6 metabolizes a number of drugs and a variety of procarcinogens. CYP2A6 also catalyzes nicotine C-oxidation leading to cotinine formation, a major metabolic pathway of nicotine in humans. There are genetic polymorphisms in the human CYP2A6 gene and a relationship between the CYP2A6 genotype and smoking habits as well as the incidence of lung cancer has been indicated. CYP2A64 alleles are the whole deleted type and are completely deficient in the enzymatic activity. An unequal crossover junction is located in the 3'-flanking region in the CYP2A64A allele, whereas the junction is located in either intron 8 or exon 9 in the CYP2A64D allele. In the present study, a novel genotyping method to distinguish between two different whole deleted alleles of CYP2A64A and CYP2A64D was established. In the process, two novel alleles, CYP2A61F and CYP2A61G, were found. The CYP2A61F has a single nucleotide polymorphism (SNP) of C5717T in exon 8, and the CYP2A61G has two SNPs, C5717T in exon 8 and A5825G in intron 8. The SNP of C5717T corresponds to C1224T on the cDNA sequence and is a synonymous mutation. Since the CYP2A61F produces a recognition site of the restriction enzymes that is the same as CYP2A64D, the presence of the CYP2A61F allele could cause a mistyping as the CYP2A64D allele. According to an improved genotyping method, the allele frequencies of CYP2A64A, CYP2A64D, CYP2A61F, and CYP2A61G in 165 Caucasians were 3.0%, 0%, 1.8%, and 1.2%, respectively. The allele frequencies of CYP2A64A, CYP2A64D, CYP2A61F, and CYP2A61G in 94 African-Americans were 0%, 0.5%, 0%, and 13.3%, respectively. This is the first report of a method that can distinguish between CYP2A64A, CYP2A64D, and CYP2A61F which could otherwise cause a mistyping as CYP2A6*4D.