Association of high T allele frequency of CYP2B6 G516T polymorphism among ethnic south Indian HIV-infected patients with elevated plasma efavirenz and nevirapine

Sir, Plasma concentrations of the non-nucleoside reverse transcriptase inhibitors efavirenz and nevirapine are known to exhibit a high degree of inter-patient variability.1 A comprehensive analysis of the existing literature on the pharmacogenetic determinants of antiretroviral drug exposure, toxicity and activity has been undertaken.1 Plasma concentrations of efavirenz and nevirapine are known to be influenced by a G to T change at codon 516 of CYP2B6.1 The frequency of the T allele varies widely across populations.2 We report the prevalence of the 516 G>T polymorphism of the CYP2B6 gene and its influence on steady-state plasma efavirenz and nevirapine concentrations in HIV-infected patients in south India. The study participants were HIV-1-infected individuals receiving antiretroviral treatment with either efavirenz (600 mg once daily)- or nevirapine (200 mg twice daily)-based highly active antiretroviral therapy (‘HAART’) regimens for a minimum period of 15 days from the Government Hospital of Thoracic Medicine, Chennai, south India. These patients were not receiving any concomitant medications known to alter the blood levels of efavirenz or nevirapine. The study was approved by the Institutional Ethics Committee, and all patients gave written informed consent. A single timepoint blood collection at 12 and 2 h post-dose for efavirenz and nevirapine, respectively, was done. Genomic DNA was extracted from whole blood, and the CYP2B6 amplicon was directly sequenced using a 3100 Avant Genetic Analyzer.3 Plasma concentrations of efavirenz and nevirapine were quantified using HPLC.4,5 Analysis of data was performed using SPSS, version 14. Shapiro's test was done to confirm normal distribution of the data. Groupwise comparison of plasma concentrations was done using Tukey's multiple comparison test. A P value of <0.05 was considered to be statistically significant. Deviation of genotype frequencies from Hardy–Weinberg equilibrium expectations was evaluated using the χ2 test. Allelic frequency was estimated by dividing total copies of individual alleles by all alleles in the population. A total of 135 HIV-1-infected persons were studied; their mean (range) age and body weight were 35 years (20–59) and 49 kg (26–91), respectively, and 90 were males. Patient adherence to treatment was satisfactory as assessed by regularity of hospital visits to collect drugs and also by the treating physician. The proportions of GG, GT and TT genotypes were 47 (35%), 58 (43%) and 30 (22%), respectively. The G and T allele frequencies in this population were 0.56 and 0.44, respectively; the polymorphism was in Hardy–Weinberg equilibrium. Among the 135 patients, 69 and 66 were receiving treatment with efavirenz- and nevirapine-containing regimens, respectively. The mean 12 h plasma efavirenz and 2 h plasma nevirapine concentrations for the GG, GT and TT genotypes are shown in Table 1. The TT genotypes had significantly higher plasma efavirenz and nevirapine concentrations than the GG and GT genotypes (P<0.001). Drug levels did not significantly differ between GG and GT genotypes. Plasma concentrations of efavirenz (EFV) and nevirapine (NVP) for the different genotypes aP<0.05 versus GG and GT. Plasma concentrations of efavirenz (EFV) and nevirapine (NVP) for the different genotypes aP<0.05 versus GG and GT. The study data showed that CYP2B6 G516T polymorphism significantly influences blood levels of efavirenz and nevirapine in the south Indian population: a finding similar to that reported by others.1 High inter-individual variability in plasma efavirenz and nevirapine levels is largely due to genetic variations in the CYP2B6 gene.1 We obtained a T allele frequency of 0.44, which is higher than that reported in Koreans (0.14), Japanese (0.16), Caucasians (0.25), White Americans (0.25) and African-Americans (0.28), but similar to that of West Africans (0.42), Chinese (0.43) and Hispanics (0.43).2 This implies that differences in pharmacogenetic and pharmacokinetic characteristics appear to be associated with ethnicity. Validated pharmacogenetic and pharmacokinetic analyses should be used to help predict likely patterns of antiretroviral treatment response among different populations worldwide. Plasma exposure to efavirenz may be higher among ethnic south Indians than among other populations. Hence, clinical trials using lower doses of these drugs are required. In fact, the feasibility of genotype-based efavirenz dose reduction in CYP2B6 G516T allelic carriers was studied and the dose reduction was found to relieve patients of efavirenz-associated CNS symptoms.6 Such studies are important, because they would have cost-saving implications, particularly for developing countries. CYP2B6 G516T genotyping at baseline may allow clinicians to optimize antiretroviral therapy in patients who initiate an efavirenz- or nevirapine-based regimen. The clinical implications of altered plasma efavirenz and nevirapine levels, as observed in this study, are not clear. Although an association between CYP2B6 G516T polymorphism and increased adverse events in the CNS during the first week of efavirenz therapy has been previously reported, this association was not apparent during subsequent weeks, despite persistently elevated efavirenz concentrations.1 This is in agreement with our observation that, in spite of TT genotypes having mean plasma efavirenz concentrations at 12 h as high as 10.59 µg/mL (peak levels could be still higher), none of the patients had persistent CNS effects or complaints of efavirenz toxicity. However, ours was a cross-sectional assessment without long-term follow-up. Prospective studies are required to definitively relate blood levels of efavirenz and nevirapine to the occurrence of adverse reactions as well as to determine whether dose modifications are effective and safe. This work was funded in part by a grant from the United States Agency for International Development provided through the World Health Organization—Model DOTS Project. None to declare. We are grateful to Dr P. R. Narayanan, Director, and Dr V. Kumaraswami, Scientist ‘F’, at the Tuberculosis Research Centre, Chennai, India, for their encouragement and support. We also thank Mr P. Kumar for technical assistance related to plasma estimations, antiretroviral treatment (ART) medical officers for patient recruitment and clinical and laboratory staff for organizing blood collections at the Government Hospital of Thoracic Medicine, Tambaram, as well as all of the patients who took part in the study.