Reactive intermediates and bioactivation pathways characterization of avitinib by LC-MS/MS: In vitro metabolic investigation

Avitinib (AC0010) is a third generation inhibitor of the EGFR (epidermal growth factor receptor) that was permitted parallel phase I clinical trials in the US and in 2014. It is estimated to enter in market within two years. In the current study, eight in vitro metabolites were detected and their chemical structures were postulated. The main in vitro phase-I metabolic reaction was N-oxidation in piperazine moiety. The generation of reactive metabolites in avitinib metabolism was investigated using rat liver microsomes while adding capturing agents, viz potassium cyanide for reactive iminium intermediates, GSH for iminoquinones and methoxylamine for aldehyde forming stable adducts which are identifiable by LC-MS/MS. Ten reactive intermediates (four iminoquinones, three iminium and three aldehydes) were characterized. The three capturing agents used resulted in proposing four different bioactivation pathways. Upon literature examination, no former articles were found for avitinib metabolism including the produced reactive metabolites.