Atazanavir metabolism according to CYP3A5 status: an in vitro-in vivo assessment

Abstract

The current study was a follow-up to an in vivo study in which atazanavir oral clearance was shown to be dependent on genetically determined CYP3A5 expression status, but only in non-African Americans. The aim of this study was to identify atazanavir metabolites generated by CYP3A5 and to evaluate this metabolite pattern in the African-American versus non-African-American CYP3A5 expressors from the previous study. First, the in vitro metabolism of atazanavir was evaluated using human liver microsomes (HLM) and CYP3A4 and CYP3A5 isoforms. Second, the metabolite pattern generated by CYP3A5 was evaluated in human plasma samples from the previous study. Atazanavir metabolites were analyzed using liquid chromatography-tandem mass spectrometry methods. Metabolite areas under the time-concentration curves (AUCs) were normalized to atazanavir AUC to generate an AUC ratio. Sixteen metabolites were observed in human liver microsomal incubations representing five "phase I" biotransformation pathways. Mono-oxidation products (M1 and M2) were formed by CYP3A5 at a faster rate than CYP3A4 by 32- and 2.6-fold, respectively. This finding was replicated in HLM from a genetically determined CYP3A5 expressor versus nonexpressor. In the in vivo samples, the M1 and M2 AUC ratios were approximately 2-fold higher in CYP3A5 expressors versus nonexpressors (P < 0.05), and the difference was similar in African Americans and non-African Americans. Thus, CYP3A5 produced a unique metabolite "signature" for atazanavir in vitro and in vivo, independent of race. Therefore, other pharmacological factors are likely to explain the apparent lack of effect of genetically determined CYP3A5 expressor status on atazanavir oral clearance in African Americans from the previous study.

Figures

Fig. 1.

Proposed atazanavir metabolite pathways (major and minor) commonly observed via human liver microsomal, CYP3A4, or CYP3A5 incubates and via human in vivo samples.

Fig. 2.

Major atazanavir metabolites observed in human liver microsomes in vitro incubations (0–30 min): ■, M1; ▴, M2; ▾, M5 and M6; ○, M7; ●, M9. Data are presented as means ± S.D. n = 6.

Fig. 3.

Major atazanavir metabolites observed via human CYP3A4 and CYP3A5 in vitro incubations (0–30 min): □, M1 (CYP3A4); ■, M1 (CYP3A5); ▵, M2 (CYP3A4), ▴, M2 (CYP3A5). Data are presented as means ± S.E.M. n = 4.

Fig. 4.

Metabolite M1/atazanavir AUC ratios in the three groups: group 1 (G1), non-African-American CYP3A5 nonexpressors (G1-NAA-NE); group 2 (G2), non-African-American CYP3A5 expressors (G2-NAA-E); and group 3 (G3), African-American CYP3A5 expressors (G3-AA-E). Data are presented as means ± S.E.M. ***, P < 0.001.

Fig. 5.

Metabolite M2/atazanavir AUC ratios in the three groups: group 1 (G1), non-African-American CYP3A5 nonexpressors (G1-NAA-NE); group 2 (G2), non-African-American CYP3A5 expressors (G2-NAA-E); and group 3 (G3), African-American CYP3A5 expressors (G3-AA-E). Data are presented as means ± S.E.M. *, P < 0.05; ***, P < 0.001.