In vivo metabolism of epothilone B in tumor-bearing nude mice: identification of three new epothilone B metabolites by capillary high-pressure liquid chromatography/mass spectrometry/tandem mass spectrometry.

Abstract

Epothilone B is a 16-membered macrolide produced by the myxobacterium Sorangium cellulosum and is currently under clinical investigation. Experimentally, epothilone B demonstrates potent antiproliferative activity at the nanomolar level in vitro, and potent regression-producing antitumor activity in vivo, similar to paclitaxel (Taxol). In order to foster the design of improved derivatives, the potential biotransformation products of epothilone B formed in liver of tumor-bearing mice after intravenous administration of 10 mg/kg were characterized in an early stage of compound development. Solely on the basis of capillary high-performance liquid chromatography, combined either with electrospray tandem mass spectrometry (in precursor and product ion scan mode) and single analyzing time-of-flight mass spectrometry (H/D exchange and accurate mass measurement), three main metabolites could be detected. The three metabolites, formed by the liver, have in common that the epoxide ring was hydrolyzed and that the macrocyclic lactone ring was opened to the acid. In two cases it is assumed that open-chain intermediates re-cycled either to a lactone or, after conjugation with taurine, to the respective lactam. The proposed structures were additionally supported by the determination of the number of the exchangeable hydrogen atoms and by confirmation of the proposed elemental composition by exact mass measurement.