Tumor-derived cell lines are indispensable tools for understanding the contribution of activated signaling pathways to the cancer phenotype and for the design and testing of targeted signal therapies. In our study, we characterize 10 colorectal carcinoma cell lines for the presence of mutations in the wnt, Ras/MAPK, PI3K and p53 pathways. The mutational spectrum found in this panel of cell lines is similar to that detected in primary CRC, albeit with higher frequency of mutation in the b-catenin and B-Raf genes. We have monitored activation of the wnt and Ras/MAPK pathways in these cells and analyzed their sensitivity to selective signaling inhibitors. Using b-catenin subcellular distribution as a marker, we show that cells harboring APC mutations have low-level activated wnt signaling, which can be blocked by the extracellular wnt inhibitor DKK-1, suggesting autocrine activation of this pathway; proliferation of these cells is also blocked by DKK-1. In contrast, cells with b-catenin mutations are unresponsive to extracellular wnt inhibition. Constitutive phosphorylation of MAPK is present in the majority of the cell lines and correlates with B-Raf but not K-Ras mutations; correspondingly, the proliferation of cells harboring mutations in B-Raf, but not K-Ras, is exquisitely sensitive inhibition of the MAPK pathway. We find no correlation between PI3K mutation or loss of PTEN expression and increased sensitivity to PI3K inhibitors. Our study discloses clear-cut differences in responsiveness to signaling inhibitors between individual mutations within an activated signaling pathway and suggests likely targets for signal-directed therapy of colorectal carcinomas. '
UICCKey words: colorectal cancer; cell lines; mutations; wnt signaling; signal therapiesThe progression of colorectal cancer from adenomas to carcinomas is characterized by a series of genetic changes, which lead to the activation of oncogenes and to the inactivation of tumor suppressor genes, perturbing the self-renewal, proliferative, adhesive and survival characteristics of the cells. 1,2 Colorectal cancers are characterized by mutations in 4 major pathways: wnt, Ras/MAPK, p53 and DNA repair.The APC gene is a key regulator of the wnt pathway. APC mutations are detected early in the adenoma-carcinoma progression, 3 are the hallmark of Familial Adenomatous Polyposis (FAP) and also occur with high frequency (up to 80%) in sporadic CRC. Mutations of the APC gene cluster in the central region of the protein and result in truncations, which delete most of the b-catenin binding sites, the axin-binding domain and the microtubule-binding regions. These deletions have been associated with activation of the canonical wnt signaling pathway and cytosolic/nuclear accumulation of b-catenin 4 but may also play an important, b-catenin independent, role in the loss of cell polarity observed in many colorectal carcinomas. 5 Underlying the relevance of the wnt pathway in the aetiogenesis of CRC, mutations of other components in the canonical wnt pathway also occur with...