Myocyte-Damaging Effects and Binding Kinetics of Boronic Acid and Epoxyketone Proteasomal-Targeted Drugs

The proteasome inhibitors bortezomib, carfilzomib, and ixazomib, which are used in the treatment of multiple myeloma have greatly improved response rates. Several other proteasome inhibitors, including delanzomib and oprozomib, are in clinical trials. Carfilzomib and oprozomib are epoxyketones that form an irreversible bond with the 20S proteasome, whereas bortezomib, ixazomib, and delanzomib are boronic acids that form slowly reversible adducts. Several of the proteasome inhibitors have been shown to exhibit specific cardiac toxicities. A primary neonatal rat myocyte model was used to study the relative myocyte-damaging effects of five proteasome inhibitors with a view to identifying potential class differences and the effect of inhibitor binding kinetics. Bortezomib was shown to induce the most myocyte damage followed by delanzomib, ixazomib, oprozomib, and carfilzomib. The sensitivity of myocytes to proteasome inhibitors, which contain high levels of chymotrypsin-like proteasomal activity, may be due to inhibition of proteasomal-dependent ongoing sarcomeric protein turnover. All inhibitors inhibited the chymotrypsin-like proteasomal activity of myocyte lysate in the low nanomolar concentration range and exhibited time-dependent inhibition kinetics characteristic of slow-binding inhibitors. Progress curve analysis of the inhibitor concentration dependence of the slow-binding kinetics was used to measure second-order "on" rate constants for binding. The second-order rate constants varied by 90-fold, with ixazomib reacting the fastest, and oprozomib the slowest. As a group, the boronic acid drugs were more damaging to myocytes than the epoxyketone drugs. Overall, inhibitor-induced myocyte damage was positively, but not significantly, correlated with their second-order rate constants.