Amphiphilic alpha-helical peptides that induces apoptosis through interacton with Bcl-2

Abstract

Bcl-2 is one of the pro-survival factors of Bcl-xl family that inhibits apoptosis signaling by interacting with Bak or Bax protein. If it is over-expressed, pro-survival Bcl-2 proteins can lead to non-apoptotic cancerous cells. Thus, a blockage of interaction of Bcl-2 with counter-part proteins could reduce survival factors and finally induce apoptosis. Then, inhibitor of interaction with Bcl-2 might be pro-apoptotic and could be used as an anti-cancer agent. Analogues of Bak or Bax peptides and their mimetic synthetic substance and even small molecules have been intensively studied as pro-apoptotic cancer therapeutics. Structural studies reveal that Bak or Bax binds as an amphipathic and high alpha-helical peptide and makes extensive hydrophobic contacts with the protein. Therefore, we thought that a simple alpha-helical and amphiphilic peptide could bind to Bcl-2 as an anchoring molecule. Then, we could apply a strategy for selecting a tailor-made peptide against Bcl-2 by carrying out mutations in both hydrophilic and hydrophobic faces of the simple peptide, generating selective peptides. We designed and synthesized a library of amphiphilic peptides mainly composed of Arg and Leu, against an intra-cellular protein target, Bcl-2. For changes of hydrophilic and hydrophobic faces of the peptide, alpha-aminoadipic acid (Aad) and Trp were introduced to increase conformational stability and to induce more interactions with the target protein. A selected mutant in hydrophilic face, 13Aad, showed strong pro-apoptotic propensity (EC50 = 8.4 nM). There are several reasons why this peptide is so apoptotic. Firstly, position and carbon number of 13Aad was optimized to induce the highest alpha-helical propensity. Secondly, cellular concentration of peptides could be dependent on cell permeability that is correlated with alpha-helical propensity. Finally, the strongest binding affinity of the peptide against Bcl-2 is also a strong pro-apoptotic factor. The selected peptide could be used as a cancer therapeutic agent inducing apoptosis in its low nanomolar concentration.