Abstract 1350: Inhibiting VPS34 suppresses Wnt/beta-catenin signaling and promotes chemotherapeutic efficacy in colon cancer cells

Abstract

Abstract A significant obstacle in the successful management of colon cancer (CRC) patients remains intrinsic or acquired drug resistance. Addressing these obstacles remain priority for reducing CRC-associated deaths. Autophagy is a survival-promoting program and upregulated in cancers to sustain metabolism and survive microenvironmental stress. In preclinical cancer models, inhibition of autophagy can restore chemosensitivity and enhance tumor cell death. However, clinical applicability of the currently available autophagy inhibitors is problematic due to potential toxicity as these inhibitors primarily interfere with lysosomal trafficking program, implicated in essential cellular processes. To overcome this limitation, we examined the effects of inhibiting VPS34, a class III PI-3 Kinase implicated in autophagy initiation, to overcome resistance to therapy by inhibiting autophagy. Notably, #36-077, the inhibitor used in our studies was synthesized based on the recently described novel VPS34 inhibitor, SAR405. Using colon cancer cells and colon tumoroids from APCmin mice, we confirmed marked upregulation of autophagy. Tumoroids exposure to #36-077 inhibited autophagy (LC-3I/II and P-62 expression) and induced apoptosis (Cleaved caspase-3 expression), and thus confirmed survival role for autophagy in CRC. We further examined efficacy of using #36-077 for adjuvant therapy. Combinatorial treatment efficacy of #36-077with 5-Fluorouracil (5-FU) and Erlotinib (anti-EGFR therapy) was tested. SW480 (highly tumorigenic) and HCT116 (Highly Metastatic) cells were used. MTT-assay helped determine cell viability. We found synergy between #36-077 and 5-FU in the killing CRC cells, and the EC50 was 4.9µm, 4.7µm and 2.40µm for 5-FU, #36-077 and 5-FU+#36-077, respectively. Combinatorial use of the #36-077 also boosted the cancer cell killing effects of Erlotinib, and the EC50 was 25.05µm ,5.59µm and 4.09 µm for Erlotinib, #36-077 and Erlotinib+#36-077, respectively. In further studies, we focused on the β-catenin regulation due to its role in promoting Wnt-signaling and autophagy. Indeed, in both, colon cancer cells and tumoroids, combinatorial treatment with 5FU+#36-077 resulted in sharp increases in phospho-β-catenin (Ser33) expression. Of note, phosphorylation of β-catenin at Ser33 by GSK-3β leads to its destabilization and degradation. TOP-FLASH reporter analysis (p<0.001 vs control) further confirmed significant inhibition of Wnt-signaling in cells subjected to combinatorial treatment. In accordance, in cells subjected to combinatorial treatment, Survivin expression (a Wnt-signaling target; p<0.001 vs control) was markedly suppressed while cleaved caspse-3 expression (p<0.05 vs control) was upregulated. Taken together, our data provide strong rationale for the therapeutic targeting of VPS34 in improving therapeutic efficacy of anti-CRC treatments. Citation Format: Balawant Kumar, Rizwan Ahmad, Sandeep Rana, Amarnath Natarajan, Punita Dhawan, Amar B. Singh. Inhibiting VPS34 suppresses Wnt/beta-catenin signaling and promotes chemotherapeutic efficacy in colon cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1350.