CYTOCHROME P450 3A-MEDIATED PHARMACOKINETIC VARIATIONS FOR ANTI-INFECTIVE AGENTS
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Two highly homologous enzymes, cytochrome P450 enzyme 3A4 (CYP3A4) and CYP3A5 play predominant roles in drug elimination by metabolizing more than half of the marketed drugs. This thesis focuses on pharmacokinetics of anti-infective drugs that are CYP3A4/5 substrates, inducers, or inhibitors in order to provide a mechanistic understanding of CYP3A4/5-related pharmacokinetic variations. In a dose-escalation study in healthy volunteers for daily doses of the anti-tuberculosis drug rifapentine, we developed and validated Ultra Performance Liquid Chromatrography-Mass Spectrometry (UPLC-MS)-based quantitation assays for measurement of plasma concentrations of rifapentine and the comparator drug, rifampin, and their desacetyl metabolites. We identified a less-than-proportional increase in rifapentine plasma peak concentration (Cmax) and area under the plasma concentration-time curve (AUC) as the rifapentine dose increased from 5 mg/kg to 20 mg/kg. Co-administration of rifapentine decreased AUC of a CYP3A4/5 substrate drug, midazolam, in a dose-independent manner and the reduction was bigger than that caused by the co-administration of rifampin. Ritonavir is a CYP3A4/5 inhibitor used in fixed combination with several anti-HIV drugs to prolong their half-lives by inhibiting CYP3A4/5-mediated metabolism of those drugs. We conducted a drug-drug interaction study to evaluate the effect of ritonavir-boosted anti-HIV drug lopinavir (LPV/r) on the anti-malarial drug quinine, which has a narrow therapeutic window. We developed a High Performance Liquid Chromatrography (HPLC) interfaced with fluorescence detector assay for simultaneous measurement of plasma concentrations of quinine and its major pharmacologically active metabolite, 3-hydroxyquinine. We found that LPV/r significantly decreased the exposure of quinine and 3-hydroxyquinine, in both total and protein-bound free forms. The finding highlights the complex nature of the influence exerted by LPV/r on CYP3A4/5 and other drug-metabolizing enzymes involved in quinine disposition including UDP-glucuronosyltransferase, and P-glycoprotein. Through studying metabolism of the anti-HIV drug maraviroc, we found CYP3A5 was the principal enzyme responsible for the formation of an abundant metabolite, namely, M1. The M1 formation capacity of CYP3A5 was 23 times higher than that of CYP3A4 in vitro. Further, human liver microsomes isolated from donors with two loss-of-function CYP3A5*3 alleles exhibited a 79% decrease in M1 formation as compared to those with two wild-type CYP3A5*1 alleles. These results indicate that maraviroc can be used as a chemical phenotyping probe to examine CYP3A5 activity in vitro. The excellent safety profile of maraviroc allowed us to further examine the utility of maraviroc to assess CYP3A5 activity in clinical settings with potential for use as a phenotyping probe. Following an oral dose of 300 mg maraviroc, people with two CYP3A5*1 alleles had 41% lower maraviroc AUC and 66% higher apparent clearance compared with those who do not have CYP3A5*1 allele. Subjects who do not have CYP3A5*1 allele had > 2 times the AUC ratio of maraviroc to its M1 metabolite compared with those who carry at least one CYP3A5*1 allele. Strong correlations between plasma concentration ratios and AUC ratios of maraviroc to M1 were observed from 4 to 10 h following the dose with R2 > 0.87. These results suggest that maraviroc has the potential to be used in clinical studies, possibly with single-point blood collections after the dose, to determine CYP3A5 activity. In conclusion, we characterized the changes in pharmacokinetics of CYP3A4/5 substrate drugs caused by anti-infective agents that are CYP3A4/5 inducers or inhibitors; and discovered a novel use of maraviroc to assess activity of the highly variable enzyme CYP3A5 in both in vitro and in clinical settings. The results from this thesis inform several anti-infective dosing regimens and provide a phenotyping tool for identification of CYP3A5 substrates, inducers, or inhibitors.