BACKGROUND: Colorectal cancer (CRC) metastasis is a leading cause of cancer-related deaths in the United States. The molecular mechanisms underlying this complex, multi-step pathway are yet to be completely elucidated. Recent reports have stressed the importance of intra-tumoral heterogeneity in the development of a metastatic phenotype. The purpose of this study was to characterize the intra-tumoral phenotypic heterogeneity between two iso-clonal human colon cancer sublines HCT116 and HCT116b on their ability to undergo metastatic colonization and survive under growth factor deprivation stress (GFDS).
MATERIALS AND METHODS: HCT116 and HCT116b cells were transfected with green fluorescence protein and subcutaneously injected into BALB/c nude male mice. Once xenografts were established, they were excised and orthotopically implanted into other male BALB/c nude mice using microsurgical techniques. Animal tissues were studied for metastases using histochemical techniques. Microarray analysis was performed to generate gene signatures associated with each subline. In vitro assessment of growth factor signaling pathway was performed under GFDS for 3 and 5 days.
RESULTS: Both HCT116 and HCT116b iso-clonal variants demonstrated 100% primary tumor growth, invasion and peritoneal spread. However, HCT116 was highly metastatic with 68% metastasis observed in liver and/or lungs compared to 4% in HCT116b. Microarray analysis revealed an upregulation of survival and metastatic genes in HCT116 cells compared to HCT116b cells. In vitro analysis showed that HCT116 upregulated survival and migratory signaling proteins and downregulated apoptotic agents under GFDS. However, HCT116b cells effectively showed the opposite response under stress inducing cell death.
CONCLUSIONS: We demonstrate the importance of clonal variation in determining metastatic potential of colorectal cancer cells using the HCT116/HCT116b iso-clonal variants in an orthotopic metastatic mouse model. Determination of clonal heterogeneity in patient tumors can serve as useful tools to identify clinically relevant biomarkers for diagnostic and therapeutic assessment of metastatic colorectal cancer.
Animals, Carcinoma, Cell Survival, Clone Cells, Colonic Neoplasms, Gene Expression Profiling, Gene Expression Regulation, Neoplastic, Genetic Heterogeneity, HCT116 Cells, Humans, Liver Neoplasms, Lung Neoplasms, Male, Mice, Mice, Nude, Neoplasm Proteins, Neoplasm Transplantation, Organ Specificity, Signal Transduction
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Chowdhury, Sanjib; Ongchin, Melanie; Sharratt, Elizabeth; Dominguez, Ivan; Wang, J.; Brattain, Michael G.; and Rajput, Ashwani, "Intra-tumoral heterogeneity in metastatic potential and survival signaling between iso-clonal HCT116 and HCT116b human colon carcinoma cell lines." (2013). Journal Articles: Eppley Institute. Paper 11.