Evaluation of the Effectiveness of l-Ergothioneine in Conjunction with Reactive Intermediates Derived from Model Pharmaceuticals

Abstract

During our investigation into a compound’s disposition within our discovery portfolio, we identified an l-ergothioneine to generate reactive intermediates. These drugs included acetaminophen, diclofenac, carbamazepine, clozapine, nefazodone, raloxifene, tamoxifen, ticlopidine, troglitazone, and ethacrynic acid. The drugs were incubated with human liver microsomes supplemented with NADPH and ET, followed by analysis with liquid chromatography–mass spectrometry (LC-MS). This process led to the detection of ET conjugates in six of the ten compounds, which exhibited structural differences: for instance, acetaminophen, raloxifene, and troglitazone presented with +ET-2H, while diclofenac, nefazodone, troglitazone, and tamoxifen showed +O + ET-2H. Additionally, nefazodone yielded +ET + O-HCl. The paper discusses structure–activity relationships (SAR) and underlying mechanisms. The proposed structures indicate that ET effectively incorporates reactive intermediates featuring highly conjugated moieties, such as quinones and quinone-imines, yet is less effective with epoxides, α-β-unsaturated ketones, and nitrenium ions. To further investigate ET’s detoxification capabilities, we analyzed metabolic products from acetaminophen, diclofenac, nefazodone, and raloxifene using rat, monkey, and human hepatocytes without GSH and ET supplementation. Interestingly, we detected conjugates of ET and GSH corresponding to +ET/GSH-2H and +O + ET/GSH-2H. Notably, our findings suggest that, in addition to scavenging reactive oxygen species, ET can also shield cells from reactive xenobiotic intermediates, similar to GSH. This research presents the first evidence of ET’s role as a trapping agent for reactive drug intermediates.