Haplotype-specific PCR for NAT2 diplotyping

Abstract

N-acetyltransferase 2 (NAT2) is an enzyme that acetylates many kinds of drugs, including the antituberculosis drug isoniazid. The NAT2 gene is highly diverse across populations. An individual can be classified as having a slow acetylator (SA), an intermediate acetylator (IA), or a rapid acetylator (RA) phenotype based on its two haplotypes (diplotype) of NAT2. SA individuals are at a higher risk for isoniazid-induced hepatitis, while the RA phenotype contributes to failure in tuberculosis treatment. Being able to predict individual NAT2 phenotypes is important for dose adjustment of isoniazid. NAT2 haplotypes are commonly determined via an indirect method of statistical haplotype inference from SNP genotyping. Here, we report a direct NAT2 haplotyping method using haplotype-specific PCR (HS-PCR) for the 6 most commonly found NAT2 haplotypes: NAT2*4, NAT2*5B, NAT2*6A, NAT2*7B, NAT2*12A, and NAT2*13A. Validation of this HS-PCR method via comparison with a sequencing method in 650 Thai DNA samples (107 RA, 279 IA, and 264 SA samples) showed a concordance rate for diplotype calls of 99.23% (645/650 samples). The discordant results in 5 samples were due to 3 rare NAT2 haplotypes: NAT*5C (n = 3), NAT2*7C (n = 1), and NAT2*11A (n = 1). This novel HS-PCR method allows direct NAT2 diplotyping, enabling the implementation of NAT2 acetylator phenotypes in clinical pharmacogenetic testing.