Optimization and Chemoproteomic Profiling of a Selective, Covalent Bfl-1-Targeting Cellular Tool

We
describe herein the discovery and optimization of a potent and
irreversible cellular probe for selective labeling of Bfl-1, a member of the Bcl-2 family. This chemical series
demonstrates robust selectivity for Bfl-1 over other related antiapoptotic
proteins and exhibits favorable cellular potency as well as promising in vivo pharmacokinetics. Notably, compound 25 achieves a kinact/KI value of 9300 M–1s–1 and elicits caspase activation at submicromolar concentrations in
cellular assays. To comprehensively profile proteome-wide selectivity,
we performed chemoproteomic analyses on compound 25 alongside
our previously reported Bfl-1 inhibitors. This enabled critical insights
into potential off-target interactions and facilitated direct comparison
of off-target profiles among distinct chemotypes targeting Bfl-1.