Abstract 809: BrafV637E mutation as an early change in hepatocarcinogenesis induced by neonatal treatment with diethylnitrosamine in B6C3F1 mice

Abstract

Abstract Neonatal treatment of B6C3F1 mice with diethylnitrosamine (DEN) is often employed as an experimental model for hepatic tumors. In this model, microscopic preneoplastic foci emerge in the early stage and subsequently progress into adenomas, from which hepatocellular carcinoma (HCC) is finally developed. We detected the BrafV637E mutation, that is correspondent to the human BrafV600E mutation, in 4/4 DEN-induced hepatic tumors at 13 months after DEN treatment by whole exome analysis, and further in 16/17 (94.1%) late tumors by the PCR-direct sequencing method. By microdissection, the BrafV637E mutation was also detected in 16/18 (88.8%) and 22/28 (78.5%) microscopic lesions (preneoplastic foci and microadenomas) respectively at 5 and 8-9 months after DEN treatment. Meanwhile, although the BrafV637E mutation was detected in 5/7 (71.4%) hepatic tumors induced by neonatal DEN treatment followed by repeated CCl4 administration, it was not found in 24 hepatic tumors induced by CCl4 alone and 6 spontaneous hepatic tumors in B6C3F1 mice, indicating that the BrafV637E mutation is specific to the DEN-initiated hepatic tumors, presumably being derived from the genotoxic action of DEN. The DEN-induced tumors showed hyperphosphorylation of MEK and ERK1/2, indicating that the BrafV637E mutation activated the downstream MAPK pathway. Furthermore, the DEN-induced tumors overexpressed the pro-survival/pro-proliferative cytokines/chemokines such as complement C5/C5a, soluble intercellular adhesion molecule (sICAM-1), IL-1 receptor antagonist (IL-1ra) and chemokine CXCL9 as well as the oncogene-induced senescence (OIS) markers, p15Ink4b and p19Arf mRNAs, suggesting that the BrafV637E mutation might influence the gene expression that is involved either in cellular growth/survival or OIS. Transgenic mice expressing BrafV637E specifically in the liver under the control of albumin promoter showed enlarged hepatic mass as large as 4-5 times of normal liver, and whole liver was consisted of small basophilic hepatocytes resembling the cells in the early preneoplastic lesions induced by neonatal treatment with DEN. These results indicate that the BrafV637E mutation plays a pivotal role in the DEN-induced hepatocarcinogenesis. Citation Format: Masahiro Yamamoto, Hiroki Tanaka, Xin Bing, Yuji Nishikawa, Kohsuke Yamazaki, Keiko Shimizu, Katsuhiro Ogawa. BrafV637E mutation as an early change in hepatocarcinogenesis induced by neonatal treatment with diethylnitrosamine in B6C3F1 mice. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 809. doi:10.1158/1538-7445.AM2015-809