Abstract 4860: Pan-HDAC inhibitor treatment sensitizes breast cancer cells to autophagy inhibitor 2-phenylethynesulfonamide: Molecular mechanism

Abstract

Abstract The stress phenotype of transformed cells is associated with increased levels of heat shock proteins (hsp) and autophagy to avoid cell death. Autophagy is a conserved catabolic pathway in which cytoplasmic macromolecules and organelles are sequestered in autophagosomes, which fuse with lysosomes for protein degradation and amino acid recycling. The molecular cascade initiating autophagosome formation is triggered by a lipid kinase (PI3KC3) signaling complex (Vps34, Vps15, ATG6 and ATG14) that mediates phagophore nucleation. We have previously reported that treatment with panobinostat (PS), a pan-histone deacetylase (HDAC) inhibitor induced hyperacetylation of hsp70 and hsp90. In the present study we demonstrate that treatment of MB231 cells with PS increased the association of hyperacetylated hsp70 with the PI3KC3 VPS34, ATG6 (Beclin-1) and KAP1 (an E3 ligase for sumoylation). PS treatment also induced the levels of ATG7, ATG5 and LC3-II (a phosphatidylethanolamine conjugated ATG8) in MB-231 cells. Treatment with 2-phenylethynesulfonamide (PES), a small molecule inhibitor of hsp70 function resulted in the abrogation of PS-induced interaction of hsp70 with hsp40, VPS34 and (the lysosome-associated membrane protein A (LAMP-2A), a protein involved in chaperone mediated autophagy. Consequently, as compared to treatment with each agent alone, co-treatment with PS and PES resulted in significantly more cell death in MB-231 and SUM159PT cells (p < 0.05). Co-treatment with PS also significantly enhanced lethality of breast cancer MB-231 and MCF-7 cells due to autophagy inhibitor 3-methyl-adenosine (3-MA, inhibitor of Vps34), or hydroxychloroquine (inhibitor of lysosome acidification). In orthotopically implanted MB-231 xenografts in the mammary fat pads of NOD/SCID mice, tumor size increase to 100, 200, 500, 1000 and 2000 cu mm demonstrated tumor size-dependent increase in the intracellular levels of hsp90 and hsp70, hyperacetylated hsp70, ATG7, ATG5, Beclin1 as well as LC3-II. These in vivo findings suggest that as breast tumors grow they exhibit the stress-phenotype involving proteotoxic stress and autophagy. Our findings demonstrate the combination therapy with PS and autophagy inhibitor, which eliminates breast cancer cells with stress phenotype including autophagy, may be especially effective against established and enlarging breast cancers. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4860.