Abstract 5220: Development of first-in-class small molecule inhibitors of FLIP which activate caspase-8, the nodal regulator of apoptosis, necroptosis and pyroptosis

Abstract

Abstract Caspase-8 is the initiator caspase for the extrinsic apoptotic pathway and now recognized as the molecular “switch” that controls 3 major forms of programmed cell death: apoptosis, necroptosis and pyroptosis. As such, methods to selectively activate caspase-8 in appropriate disease contexts represent an exciting new therapeutic paradigm. In cancer, evasion of cell death is a fundamental cause of resistance to therapy, prompting the development of therapeutics that reactivate cell death pathways, with the first of these (targeting the intrinsic apoptotic pathway) recently clinically approved. The apoptosis modulator FLIP is a non-redundant inhibitor of caspase-8 activation in the extrinsic apoptotic pathway and is the only human pseudo-caspase. FLIP is frequently overexpressed in a number of cancers and leukemias and, through its modulation of caspase-8, has been shown to be a major mediator of resistance to standard-of-care chemotherapies, targeted therapies and radiotherapy. Moreover, by regulating caspase-8 activity, FLIP is a key determinant of cell death induced by immune effector cells. Thus, targeting FLIP's interaction with caspase-8 represents a unique therapeutic approach for enhancing standard-of-care anti-cancer therapies and promoting anti-tumor immunity. We have identified a series of potent (nM), selective small molecule inhibitors which, by disrupting the binding of FLIP to effector proteins, are able to activate caspase-8-dependent apoptosis in multiple cancer cell lines. These protein-protein interaction inhibitors, which are relatively low MW and have favorable drug-like properties, have potent in vitro activity as single agents in several clinically-relevant settings, including KRAS mutant non-small cell lung cancer (NSCLC). Moreover, these FLIP inhibitors have been shown to potentiate the effects of standard-of-care chemotherapeutics and targeted anti-cancer agents in pre-clinical models. Recently, FLIP has emerged as a key mediator of resistance to the 3rd generation EGFR inhibitor Osimertinib in EGFR mutant NSCLC. We found that Osimertinib-resistant EGFR mutant NSCLC cells express elevated levels of FLIP and are more sensitive to FLIP inhibitors. Most importantly, compared to treatment with Osimertinib alone, co-treatment with Osimertinib and FLIP inhibitors prevented tumor regrowth in vivo after treatment cessation. Similar effects have been observed in vitro with the MEK inhibitor Selumetinib in KRAS mutant NSCLC models. In summary, we have identified novel, drug-like activators of caspase-8 with a unique mechanism-of-action and successfully completed proof-of-concept in vivo efficacy studies which have strong translational relevance to the clinical situation in EGFR mutant NSCLC, where despite excellent response rates to Osimertinib, patient relapse is inevitable. Moreover, these compounds have potential for broad application in treatment of several cancers both as single agents and in combination with standard-of-care therapy and other clinically-relevant targeted agents. Citation Format: Daniel B. Longley, Catherine Higgins, Jennifer Fox, Jamie Z. Roberts, Ray Boffey, Sophie Williams, Trevor Perrior, Martin J. Page, Tim Harrison. Development of first-in-class small molecule inhibitors of FLIP which activate caspase-8, the nodal regulator of apoptosis, necroptosis and pyroptosis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5220.