Abstract A05: Context-dependent transformation with activated Ras isoforms in human normal epithelial cells

The classical mammalian ras genes exist three isoforms K-, H- and N-Ras. Ras mutations are genetic events that have been detected in 30 % of all human cancers. Unfortunately, mutated Ras tumors have not responded to chemotherapy, an IgG1 chimeric monoclonal antibody and tyrosine-kinase inhibitors against EGFR. Therefore, there is need for development of new therapeutic strategies to mutated Ras tumors. Although the function of three Ras isoforms is considered to be almost the same, the occurrence of Ras mutation is largely different in human each tissue. For example, APC and BRACA1 genes are associated with a high risk of developing tissue specific cancer. APC mutation caused familial adenomatous polyposis. Affected people have a nearly 100% lifetime risk of developing colorectal cancer, whereas other tumor types are rare. Such genes are therefore considered as tissue-specific cancer genes. Whether Ras genes have tissue specificity has not been clarified. In this study, we thought to elucidate whether mutated Ras isoform has tissue specificity and there is a difference in the function among three mutated Ras isoforms by using human normal epithelial cell lines (prostate, small airway, mammary, skin and bronchus). In clinical data, skin and bronchus frequently occurred H-Ras mutation, others occurred K-Ras mutation. Firstly, we induced immortalization of these cell lines by introduction of human papillomavirus E6/E7 and mutant c-myc (T58A). These cell lines could stably culture and were expressed cytokeratin 5, which is basal cell marker of epidermis. We analyzed the carcinogenesis efficiency by soft-agar assay when the cells infected with retrovirus of three mutated Ras in five immortalized cell lines. As a result, carcinogenesis efficiency was different for each cell line. These results were mostly consistent with clinical data. Also, we identified downstream signaling of Ras (PI3K-AKT and MAPK signaling) involved in carcinogenesis by performing Western blot analysis when the cells infected with mutated Ras, which induced highest carcinogenesis efficiency in each cell lines. As a result, the protein expression level that associated with growth-related PI3K/AKT and MAPK signaling affected differently in each mutated Ras isoform. These results indicated that mutated Ras isoform have tissue specificity and specific functions involved in carcinogenesis. Taken together, we concluded that these events present the possibility of new therapeutic target for mutated Ras tumors. Citation Format: Minami Kumazaki, Yusuke Yamamoto. Context-dependent transformation with activated Ras isoforms in human normal epithelial cells [abstract]. In: Proceedings of the AACR Special Conference on Targeting RAS-Driven Cancers; 2018 Dec 9-12; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(5_Suppl):Abstract nr A05.