Impact of Efavirenz on Heme Protein Induction and Biomarkers: Insights From In Vitro Experiments and a Clinical Study

Coproporphyrin I (CPI) is increasingly recognized as a reliable endogenous biomarker for hepatic OATP1B transporters. However, confounding mechanisms such as increased heme biosynthesis driven by induction of CYP enzymes, which are hemoproteins, may affect CPI levels independently of OATP1B1 inhibition. The aim of this study was to investigate whether activation of nuclear receptors and subsequent CYP induction by efavirenz affects CPI levels in vitro and in vivo. We first evaluated the potential of efavirenz to interact with OATP1B1 in vitro. We then assessed efavirenz-induced changes in vitro, analyzing the mRNA expression of CYP2B6 and both mRNA and protein expression of ALAS1 (5-aminolevulinate synthase 1), the rate-limiting enzyme in heme biosynthesis. Additionally, we quantified CPI levels in the cell culture supernatant. Finally, we determined plasma CPI levels in healthy volunteers pre- and post-induction with efavirenz. In transport studies, efavirenz did not alter OATP1B1-mediated uptake of CPI, whereas rosuvastatin reduced their intracellular accumulation. Treatment with efavirenz significantly increased CYP2B6 mRNA expression, confirming CYP induction in vitro, but had no effect on ALAS1 expression at either mRNA or protein level. Correspondingly, CPI levels in the cell supernatant remained unchanged after efavirenz treatment compared to solvent control. In vivo, plasma CPI levels were also unaffected by efavirenz. Our in vitro findings demonstrate that while efavirenz induces CYP expression, it does not impact ALAS1 expression or alter CPI levels, supported by our in vivo findings where efavirenz did not change plasma CPI, suggesting that CYP enzyme induction does not relevantly affect coproporphyrin levels.