Different Reactive Metabolites of Nevirapine Require Distinct Glutathione S-Transferase Isoforms for Bioinactivation

Abstract

Nevirapine (NVP) is a non-nucleoside reverse transcriptase-inhibitor, which is associated with severe idiosyncratic skin rash and hepatotoxicity. These adverse drug reactions are believed to be mediated by the formation of epoxides and/or quinone methide formed by oxidative metabolism by P450s and 12-sulfoxyl-NVP formed by sequential 12-hydroxylation and O-sulfonation. Although different GSH-conjugates and corresponding mercapturic acids have been demonstrated previously in vitro and in vivo, the role of the glutathione S-transferases in the inactivation of the different reactive metabolites has not been studied so far. In the present study the activity of 10 recombinant human glutathione S-transferases (GSTs) in the detoxification of the different reactive metabolites of NVP was studied. The results show that GSTP1-1 is a highly active catalyst of GSH-conjugation of the oxidative metabolites of NVP, even at high GSH-concentration. Experiments with trideuterated NVP suggest involvement of a reactive epoxide rather than quinone methide in the formation of the GSH-conjugate formed after oxidative bioactivation. GSH-conjugation of 12-sulfoxyl-NVP forming NVP-12-GSH was only catalyzed by GSTM1-1, GSTA1-1, and GSTA3-3. Although the exact expression levels of these enzymes in the skin is unknown, the relatively low activity of this catalysis makes it unlikely that GSTs can provide significant protection against this metabolite. However, since NVP-12-GSH is specifically formed via the 12-sulfoxyl-NVP, its corresponding urinary mercapturic acid can be considered as a biomarker for recent internal exposure to this protein-reactive sulfate. However, it has to be taken into account that 12-sulfoxyl-NVP is not completely trapped by GSH and that rates of bioinactivation will differ between patients due to variability in expression of GSTM1, GSTA1, and GSTA3.

Nevirapine Biotransformation Insights: An Integrated In Vitro Approach Unveils the Biocompetence and Glutathiolomic Profile of a Human Hepatocyte-Like Cell 3D Model

“…GSTA1/2 share both the expression and regulation mechanisms and substrates, and account for 3% of total hepatocyte cytoplasmic proteins [43]. 12-Sulfoxy-NVP and the arene oxide precursor of 3-OH-NVP are the only metabolites known to form glutathione conjugates [44]. From those, only the formation of 12-sulfoxy-NVP-derived glutathione adducts have been reported to be GST-dependent, mediated by GSTA1-1, GSTM1-1 and GSTA3-1 [44].…”

“…12-Sulfoxy-NVP and the arene oxide precursor of 3-OH-NVP are the only metabolites known to form glutathione conjugates [44]. From those, only the formation of 12-sulfoxy-NVP-derived glutathione adducts have been reported to be GST-dependent, mediated by GSTA1-1, GSTM1-1 and GSTA3-1 [44]. In this study, NVP induced GSTA1-A2 in 3D-HLCs but not in 2D-HLCs, which may be a cellular adaptive mechanism for protection against oxidative stress [39].…”

Nevirapine induced mitochondrial dysfunction in HepG2 cells

“…This bioactivation occurs as part of the detoxification process which is divided into three phases. Phase I enzymes such as cytochrome P450 proteins mediate the initial detoxification process 24 generating hydroxyl-metabolites such as 2-OH-NVP, 3-OH-NVP and 12-OH-NVP 25 . 12-OH-NVP can be O-sulfonated to the protein reactive 12-Sulfoxyl-NVP 25 .…”

“…12-OH-NVP can be O-sulfonated to the protein reactive 12-Sulfoxyl-NVP 25 . However, other pathways involving the generation of NVP-epoxides have been proposed 26 , 27 with phase II enzymes such as glutathione S-transferase mediating the subsequent glutathionylation of the metabolite 24 . Phase III enzymes mediate the subsequent excretion of the metabolites from the cells.…”

Metabolic Epoxidation Is a Critical Step for the Development of Benzbromarone-Induced Hepatotoxicity

“…It is widely accepted that cytochrome P450 (P450)-mediated metabolic activation is most frequently responsible for drug-induced idiosyncratic hepatotoxicity and drug withdrawal from the market (Leung et al, 2012;Dekker et al, 2016;Xuan et al, 2016). Our early metabolic studies illustrated that CYP3A metabolized BBR to epoxide intermediate(s) that reacted with sulfur nucleophiles of protein to form protein covalent binding both in vitro and in vivo (Wang et al, 2016a,b).…”