288 HDAC-1 Inhibition Prevents Smoking Induced PanIN Lesions and EMT in Kras Mouse Model of Pancreatic Cancer

Abstract

Background: Histone deacetylases (HDAC) inhibitors have been shown to suppress tumor growth in many cancers. HDAC class 1 regulates pancreatic cancer growth and resistance to cell death through a mechanism that involves NF-kB. We have developed a mouse model of pancreatic cancer induced by cigarette smoke. We hypothesized that HDAC-1 mediates progression of the disease induced by smoking and that inhibition of HDAC-1 will prevent both cancer cell survival and EMT/metastasis in vitro and in vivo. Methods: Pdx-Cre;LSLKras mice were exposed to filtered air or to cigarette smoke in smoke chambers for 6 weeks. A set of mice were injected with HDAC1 inhibitor SAHA (50mg/Kg) 5 times per week. Pancreatic intraepithelial neoplasia (PanIN) lesions, fibrosis, and inflammation weremeasured by immunohistochemistry (IHC). Pancreas levels of proliferation, apoptosis, and EMT markers were measured by IHC andWestern. Pharmacological and molecular inhibitors of HDAC1 were applied in MIA PaCa-2 and AsPC-1 cell lines. Inhibition of Zeb1, a regulator of EMT, was achieved by using Zeb1 siRNA. EMT markers and transcription factors as well as proteins acetylation level were measured by Western. Cell survival and apoptosis in cells were measured by MTT assay and DNA fragmentation ELISA assay, respectively. Results: Mice exposed to cigarette smoke showed increased level of PanIN lesions, fibrosis, and inflammation. The three pro-cancer effects were prevented in mice exposed to HDAC-1 inhibitor SAHA. SAHA decreased the levels of PanIN lesions and fibrosis, not only to a level lower than in mice exposed to cigarette smoke, but also lower than in control mice exposed to filtered air. SAHA decreased the protein level of vimentin, zeb1, sox2 and CD133 and increased E-cadherin level in pancreatic tissue indicting that it prevented EMT and stemness. Pharmacological and molecular inhibitions of HDAC-1 also significantly and dose-dependently decreased proliferation and at a lesser extent stimulated apoptosis in pancreatic cancer cells. Zeb1 siRNA decreased the level of vimentin and increased E-cadherin in pancreatic cancer cells. Conclusion: We found that smoking stimulates an increase in PanIN lesions through a mechanism that involves HDAC-1 in Kras transgenic mice. HDAC-1 not only affected proliferation of the cancer cells, but also EMT, which is involved in metastasis and cancer stemness. These data suggest an important role for HDAC-1 in pancreatic carcinogenesis.