Preclinical Development Combination of Pan-Histone Deacetylase Inhibitor and Autophagy Inhibitor Exerts Superior Efficacy against Triple-Negative Human Breast Cancer Cells

Histone deacetylase (HDAC) inhibitors (HDI) induce endoplasmic reticulum (ER) stress and apoptosis, while promoting autophagy, which promotes cancer cell survival when apoptosis is compromised. Here, we determined the in vitro and in vivo activity of the combination of the pan-HDI panobinostat and the autophagy inhibitor chloroquine against humanestrogen/progesterone receptor andHER2 (triple)-negative breast cancer (TNBC) cells. Treatment of MB-231 and SUM159PT cells with panobinostat disrupted the hsp90/histone deacetylase 6/HSF1/p97 complex, resulting in the upregulation of hsp. This was accompanied by the induction of enhanced autophagic flux as evidenced by increased expression of LC3B-II and the degradation of the autophagic substrate p62. Treatment with panobinostat also induced the accumulation and colocalization ofp62withLC3B-II in cytosolic foci as evidencedby immunofluorescent confocalmicroscopy. Inhibition of panobinostat-induced autophagic flux by chloroquine markedly induced the accumulation of polyubiquitylated proteins and p62, caused synergistic cell death of MB-231 and SUM159PT cells, and inhibited mammosphere formation inMB-231 cells, comparedwith treatmentwith each agent alone. Finally, inmousemammary fat pad xenografts of MB-231 cells, a tumor size–dependent induction of heat shock response, ER stress and autophagywere observed. Cotreatmentwith panobinostat and chloroquine resulted in reduced tumor burden and increased the survival ofMB-231 breast cancer xenografts. Collectively, ourfindings show that cotreatment with an autophagy inhibitor and pan-HDI, for example, chloroquine and panobinostat results in accumulation of toxic polyubiquitylated proteins, exerts superior inhibitory effects on TNBC cell growth, and increases the survival of TNBC xenografts. Mol Cancer Ther; 11(4); 973–83. 2012 AACR.