Abstract LB-241: Photoreactive stapled peptides for target discovery in cancer
Authors: Braun Craig R. , Mintseris Julian , Gavathiotis Evripidis , Bird Gregory H. , Gygi Steven P. , Walensky Loren D. 
Published: June 2012

Protein interactions mediate the pathologic cellular activities that lead to the development, maintenance, and recurrence of cancer. Whether fleeting or stable, homeostatic or aberrant, protein partnerships and their sites of contact form the basis for discovery of biological pathways, disease mechanisms, and opportunities for therapeutic intervention. Like the teeth of a key that perfectly match a lock, complementary protein shape is critical to the execution of biological interactions. These contact points are embedded within a complex protein structure that provides the infrastructure to maintain the essential bioactive fold. When the biological activity of cancer-causing proteins is assigned to discrete subdomains, their mode of action, menu of protein targets, and explicit sites of target engagement are frequently unknown. Ideally, these evolutionarily honed substructures could be used to capture and thereby catalogue their protein targets; however, out of context from the whole protein, bioactive subdomains often unfold, resulting in loss of biological shape, potency, and specificity.

To address the shortcomings of peptides and reclaim their remarkable capacity to selectively bind and modulate protein targets, we previously generated hydrocarbon-stapled alpha-helices that recapitulate the native bioactive structure, exhibit marked protease resistance, and penetrate intact cells. To date, we have deployed stapled peptides to advance a new therapeutic strategy for targeting protein interactions in vivo, uncover an unanticipated function for a death protein in metabolism, structurally define the elusive activation site on an essential executioner protein of the cell death pathway, and identify a natural alpha-helical peptide that can function as an exclusive inhibitor of a formidable anti-apoptotic protein linked to cancer. Here, we combine peptide alpha-helix stabilization with photoaffinity labeling, proteomic analysis, and structure calculation to generate a new strategy for both expanding our grasp of the alpha-helical interactome and localizing alpha-helix/protein interaction sites. We demonstrate the capacity of photoreactive helices modeled after the BH3 death domains of BCL-2 family proteins to (1) covalently trap both static and dynamic protein interactors and (2) identify explicit sites of engagement, providing a critical link between interactome discovery and targeted drug design. We believe that the development of photoreactive stapled peptides will extend the potential for discovery of novel and unforeseen protein interactions and how they impact cancer pathogenesis. Importantly, the stapled peptide constructs used to capture protein targets can be applied to interrogate them in a cellular context and provide the templates for developing next-generation cancer therapeutics.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-241. doi:10.1158/1538-7445.AM2011-LB-241