Chemical aspects of acylfulvene bioactivation to a cytotoxic reactive intermediate.

Abstract

Understanding molecular mechanisms of cytotoxicity is vital to the development of more effective chemotherapies. Acylfulvenes (AFs) are a class of semisynthetic analogues of the natural product illudin S. Minor structural changes between the parent compound and AFs have resulted in a more favorable selectivity profile in preclinical chemotherapy assays. AF cytotoxicity involves alkylation of biological targets, including DNA and cellular proteins. While AFs are capable of direct alkylation, reductive bioactivation to an electrophilic intermediate is correlated with enhanced cytotoxicity. Alkenal/one oxidoreductase (AOR) is a cytosolic enzyme implicated in activating AF in cells that are sensitive to the drug. This study aims to elucidate chemical aspects of acylfulvene activation mechanisms. We compared enzymatic versus chemical activation pathways for AF involving NADPH-dependent AOR or sodium borohydride, respectively. These two processes result in isomeric reactive intermediates. Despite structural differences, these isomers appear to have similar biological activity and give rise to similar patterns of DNA modification. Cell-based studies, utilizing human embryonic kidney cells transiently transfected with an AOR-overexpressing vector, were conducted to test the hypothesis that a chemically activated AF does not require further bioactivation to be cytotoxic. The reactivity of this activated compound was further assessed by measuring its half-life in the presence of acid. On the basis of this study, we anticipate that the chemically activated form of AF will serve as a useful tool for anticipate that the chemically activated form of AF will serve as a useful tool for evaluating protein and nucleic acid interactions, and to gain a further understanding of their contributions to cytotoxicity, independent of bioactivation.