The overexpression of anti-apoptotic proteins belonging to the Bcl2 family (Bcl-2, Bcl-xL, Bcl-w, Mcl-1, Bcl2A1) is one of the resistance mechanisms used by cancer cells to evade regulatory apoptosis. The expression and importance of each anti-apoptotic protein varies with cancer type and tumor development stage, and may change during chemotherapy. Therefore, inhibiting one or more of these oncogenic proteins has the potential to restore apoptosis in different types and stages of cancer, prompting the search for potent inhibitors of variable Bcl2 family specificity.

A 26-residue peptide BimBH3 binds indiscriminately to multiple oncogenic Bcl2 proteins but, being a linear peptide, lacks the necessary cell penetration and proteolytic stability to induce apoptosis of cancer cells in vivo. Here, we have reengineered the promiscuous Bim peptide to bind specifically to proteins Bcl2A1 and Mcl-1, whose overexpression is linked to the progression some cancers, particularly, melanoma. Optimization initially required N- and C-terminal truncation, insertion of one and then two sidechain to sidechain helix-inducing crosslinks, as well as an N-terminal acrylamide electrophile for covalent binding to Cys55 in Bcl2A1. Subtle variation of the nature and positioning of the helix-inducing constraints subsequently produced 14-residue bicyclic peptides with no electrophile that were potent dual inhibitors of Bcl2A1 and Mcl-1, with high selectivity over Bcl-w, Bcl-xL and Bcl-2 proteins. The bicyclic helix induced apoptosis of SkMel-28 melanoma cells, which was enhanced by the anticancer drug etoposide.