Abstract 1337: Novel model of pancreatic cancer induced by smoking compound NNK

Abstract

Abstract Background & Aims: Cigarette smoking is a major risk factor for pancreatic cancer. However, the molecular signaling mechanisms through which smoking promotes pancreatic cancer remain unknown. Experimental models of smoking-induced pancreatic cancer have not been developed. Here we studied the effect of cigarette smoking compound: nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) on cancer progression in Elastase-Kras (EL-Kras) mice, which have mutated/activated Kras. We further determined the effect of NNK and cigarette smoking extract (CSE) on cell death pathways both in vivo and in vitro in human pancreatic ductal cells. Methods: Wild type and EL-Kras transgenic mice were injected with NNK (100mg/Kg) once a week for 4 weeks. Mice were sacrificed one week later. Analysis of pancreatic lesions, adenocarcinoma, as well as measurement of fibrosis, inflammation, and proliferation were performed by immunohistochemistry. Apoptosis in tissue was assessed by TUNEL assay. Immortalized human ductal cells HPDE6 were cultured for up to 1 week in the presence or absence of NNK (1nM-1µM) or CSE (4ng/ml-4µg/ml). Apoptosis was assessed by measuring DNA fragmentation. Autophagy in cells and in mice pancreatic tissue was assessed by measuring autophagic marker LC3-II with western blot and with immunofluorescence. Results: NNK injections significantly increased (8 times) the number of pancreatic neoplastic lesions in EL-Kras mice. Importantly, NNK induced progression of the pancreatic lesions to the adenocarcinoma stage. Staining of pancreatic tumors with amylase and cytokeratines 7 and 9 showed that cancer cells present the phenotype of both acinar and ductal cells indicating that cells are at the stage of progression to the ductal adenocarcinoma phenotype. Stimulation of pancreatic adenocarcinoma was associated with a decrease in autophagy and apoptosis of exocrine pancreatic cells. NNK inhibited autophagy and apoptosis in pancreas from wild type mice but the effect was much less pronounced. Importantly, NNK injection stimulated fibrosis and stellate cells recruitment and activation. Furthermore, NNK stimulated inflammation. Similarly, exposure to NNK or CSE inhibited apoptosis and autophagy of human ductal cells. NNK and CSE increased Akt phosphorylation both in vivo and in vitro. Pharmacological and molecular inhibition of Akt kinase reversed the smoking inhibitory effect on apoptosis and autophagy, indicating a central role of Akt in the progression of pancreatic cancer by smoking compounds. Further, Akt inhibited autophagy through regulation of AMPK phosphorylation. Conclusion: NNK-induced cancer progression in EL-Kras mice represents the first mouse model of pancreatic cancer induced by smoking and can be utilized to understand the mechanism of progression of the disease. Akt kinase is a key mediator of smoking-induced pancreatic cancer progression. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1337. doi:1538-7445.AM2012-1337