oa Investigation of genes that are related to colorectal cancer and liver metastasis

Progression of colorectal cancer (CRC) culminates in liver metastasis. To identify genes that are involved in the metastatic phenotype, cDNA microarrays were used to analyze mRNA expression profiles of CC531 rat colon adenocarcinoma cells for changes related to their homing into the liver. Briefly, CC531 cells were intraportally implanted into the liver of Wag-Rij rats and re-isolated after 3, 6, 9, 14 and 21 days. Altogether 2919 genes from ca. 25000 genes were deregulated as a response to metastasis formation into the liver. Candidates from promising gene families were chosen for further analysis using different in vitro models and human CRC samples. These gene families included the claudins and the insulin-like growth factor binding proteins (IGFBPs). For claudins, it was shown that the initial phase of rat CRC cell homing into the liver involves a transient down-regulation of several claudins. Also it was shown that siRNA silencing of Cldn1 and Cldn4 increased migration and reduced colony formation, with these phenotypes being consistent with metastatic homing. Correspondingly, human CRC samples showed increased Cldn1 and Cldn4 expression in UICC stages l-lll, and significantly reduced expression in stage IV and in liver metastasis. Additionally, the IHC analysis revealed that the expression of Cldn1, Cldn4 and Cldn3 in liver metastasis specimens was lower than in the corresponding primary carcinomas. Taken together, metastasizing cells require a transient reduction in claudin expression for liberation from the primary tumor and then initialization metastatic growth in the liver. For IGFBPs it was shown that deregulation of IGFBP3 and IGFBP7 is related to the metastatic behavior of CRC cells. In CRC patients, a significant correlation between the expression levels of these genes was noticed but no relation to overall survival was registered. Further, the patients' age was inversely correlated with the expression of IGFBP3. Silencing of IGFBP3 and/or IGFBP7 in human and rat colorectal cancer cell lines reduced proliferation, colony formation, and for IGFBP3, also migration. These results indicate that IGFBP3 and 7 cannot be simply assigned to the group of tumor suppressors, but have additional properties, which become evident only in the context of cancer progression and metastasis formation.