HCV Antiviral Drugs Have the Potential to Adversely Perturb the Fetal-Maternal Communication Axis through Inhibition of CYP3A7 DHEA-S Oxidation

Abstract

The hepatitis C virus (HCV) poses a great risk to pregnant people and their developing fetus, yet no HCV antiviral treatment guidelines have been established. While there has been a substantial increase in the development of HCV antivirals, the effect they have on the developing fetus remains poorly defined. Many of these drugs are metabolized through the cytochrome P450 CYP3A pathway, which is mediated by cytochrome P450 3A7 (CYP3A7) in the fetus and developing infant. In this study, we sought to investigate the effect HCV antivirals have on CYP3A7 metabolism, as this CYP enzyme plays a vital role in proper fetal and neonatal development. Of the 13 HCV antivirals we investigated, 8 (∼62%) inhibited CYP3A7 metabolic activity by 50% or more at a concentration of 20 µM. Furthermore, paritaprevir, asunaprevir, simeprevir, danoprevir, and glecaprevir all had observed half-maximal inhibitory concentrations between the range of 10 and 20 µM, which is physiologically relevant in comparison with the K_m of dehydroepiandrosterone-sulfate (DHEA-S) oxidation (reported to be between 5 and 20 µM). We also discovered that paritaprevir is a time-dependent inhibitor of CYP3A7, which shifts the IC₅₀ ∼twofold from 11 µM to 5 µM. Upon further characterization, paritaprevir inactivates DHEA-S metabolism by CYP3A7, with K_I and K_inact values of 4.66 µM and 0.00954 minute^-1, respectively. Depending on treatment plan and off-label drug use, HCV treatment could adversely affect the fetal-maternal communication axis by blocking fetal CYP3A7 metabolism of important endogenous hormones. SIGNIFICANCE STATEMENT: The prevalence of HCV in pregnant people is estimated at between 1% and 8% of the global population, yet little to no information exists about the risk antiviral treatment poses to the developing fetus. There is a potential risk of drugs adversely affecting mother-fetal communication by inhibiting fetal hepatic CYP3A7, an integral enzyme for estriol production. We discovered that five HCV antivirals inhibited DHEA-S metabolism by CYP3A7, and paritaprevir inactivated the enzyme. Our studies demonstrate the potential threat these drugs pose to proper fetal development.

Fig. 1.

CYP3A7 inhibition by HCV antivirals as assessed by the DBF high-throughput screening fluorescence assay. All drugs were tested in triplicate, with the black circles representing individual test points and error bars representing the standard deviation. Calculations were based upon the solvent control (representing 100% activity).

Fig. 2.

Dose-response curves of CYP3A7 activity depletion by (A) danoprevir, (B) glecaprevir, (C) asunaprevir, (D) simeprevir, (E) paritaprevir, and (F) ritonavir. Black circles represent the average of triplicate runs (n = 3) and error bars represent the standard deviation. Curves were fit to the [Inhibitor] versus response (three parameters) equation in GraphPad Prism (version 10.0.2) to determine IC₅₀s and goodness-of-fits for each drug (R²).

Fig. 3.

Dose–response curve shifts of (A) paritaprevir and (B) ritonavir following 30-minute NADPH preincubation. Black circles and squares represent the average of triplicate experiments (n = 3) in which there was and was not a 30-minute preincubation with NADPH, respectively. Error bars represent the standard deviation of each concentration tested. Points were fitted to the [Inhibitor] versus response (three parameters) curve in GraphPad Prism (version 10.0.2).

Fig. 4.

Time- and concentration-dependent inhibition of CYP3A7 DHEA-S 16α-hydroxylation by paritaprevir. (A) Pseudo-first-order kinetic plots at five paritaprevir concentrations based on the percent CYP3A7 activity remaining versus the preincubation time in the presence of NADPH. All points represent the average of triplicate incubations, and error bars represent the standard deviation. (B) Inactivation rate constants (k_obs) at the five paritaprevir concentrations assessed (R²: 0.864).

Fig. 5.

Docking of paritaprevir within the CYP3A7 active site. (A) Most energetically favorable pose of paritaprevir (delta G = –12.7 kcal/mol) docked in the CYP3A7 active site (paritaprevir show in cyan, heme prosthetic group shown in red, and the CYP3A7 protein backbone shown in tan). (B) Close-up of paritaprevir in the CYP3A7 active site showing the relative distance of the C9 of paritaprevir to the heme iron and interacting amino acid residues (phenylalanines shown in green, tyrosine in yellow, and tryptophan in magenta; other colors same as panel A). (C) Cut-away view of paritaprevir in the CYP3A7 active site showing the solvent accessible surface of the protein and distance from C9 to the heme iron (red; other colors same as panels A and B). (D) Space filling model of paritaprevir in the CYP3A7 active site (color scheme same as previous).