Abstract 596: Endolysosomal trafficking defects leads to caspase independed cell death in colon cancer

Currently approved conventional chemotherapeutics for treating colorectal cancer (CRC) exert their effect through activating apoptotic pathways in cancer cells. However, CRC cells often become resistant due to mutations in apoptotic pathways rendering chemotherapeutics ineffective. Drug resistant CRC typically advances to metastatic stage, resulting in poor prognosis and shorter survival time. We have recently discovered structurally constrained 4-pyridynyl BAPT compounds that cause a form of non-apoptotic cell death in CRC cells characterized by the simultaneous induction of dysregulated macropinocytosis (i.e. self-drinking) and macroautophagy (i.e. self-eating). We define this distinctive type of caspase-independent cell death as ‘methuophagy9 (from the Greek ‘methuo9, to drink to intoxication, and ‘phagy9, self-eating). In particular, our initial lead, BAPT-27, does not induce cell death via activation of the classical apoptotic pathways, but causes methuophagy in submicromolar range. BAPT-27 induces substantial accumulation of fluid-filled, heterogeneous vacuoles and autophagosomes, resulting in metabolic failure, loss of membrane integrity, and detachment of cells from the substratum. The cell death is accompanied by extensive autophagy in the absence of cell shrinkage, chromatin condensation, alterations in mitochondrial membrane potential, or damage to the nuclear membrane. Other hit analogues i.e. BAPT-36, BAPT-38, BAPT-54 also potently trigger this form of non-apoptotic cell death in CRC cells. We observed that methuophagic cell death is accompanied by a significant increase in lysosomal activity, numbers, and biogenesis as confirmed by ultrastructural imaging. Our results suggest that BAPT analogues produce endolysosomal trafficking defects that prevent recycling of lysosomes and cause lysosome-to-nucleus signaling defect, a mechanism that disturb cellular clearance and energy metabolism. Furthermore, they disrupt vesicular trafficking at the lysosomal nexus, manifested by defective processing of procathepsins, impairment of autophagic flux, V-ATPase levels, and accumulation of autophagosomes and acrgo protein p-62. However, specific changes in core pharmacophore could result in loss of methuophagy induction and drive the cell death towards apoptosis. For example, BAPT-42 was found to have limited effects on lysosomal pathways and does not kill the cells through methuophagy, but induces apoptosis. Identifying the biology of methuophagy-induced cell death and lysosomal clearance pathways will allow us to better design this new class of compounds that kill CRC cells via non-apoptotic mechanisms for treatment of patients with apoptotic-resistant and aggressive CRCs. Citation Format: Noor Hussein, Mariah Pasternak, Shikha Kumari, Zaynah Awthe, William Gunning, Paul C. Trippier, Amit Tiwari. Endolysosomal trafficking defects leads to caspase independed cell death in colon cancer [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 596.