The IKK/NFkB & B-catenin pathway is critical for migration, invasion and metastasis in colorectal cancer Free

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The transcription factors, nuclear factor kappa B (NFκB) and β-catenin coordinate the expression of genes that control cell proliferation, survival, and transformation. Recently our laboratory has delineated a direct role of the AKT/IκB kinase (IKK) pathway in promoting angiogenic/metastatic gene expression in colorectal cancer (CRC) by inducing inappropriate constitutive activation of NFκB and β-catenin. The IKK complex consists of the two highly related kinase subunits IKKα and IKKβ, as well as a third structural subunit, IKKγ. As our previous work has shown separate distinct functions of the highly related IKKα and IKKβ kinase subunits, we investigated the effect of their individual loss in regulating the invasive and metastatic potential of the RKO CRC cell line. We used RNA interference (RNAi) to obtain clones with stable specific knockdown of IKKα (RA14) or IKKβ (RB3). We also stably over-expressed the IκBα super-repressor (RKO SR) to specifically inhibit NFκB. The basal and tumor necrosis factor-induced NFκB-dependent promoter activity is dramatically reduced in all three mutant lines as compared to the RKO parental and scrambled RNAi control cell lines. The basal and Wnt-induced β-catenin-dependent promoter activity is notably reduced only in the IKKα RNAi cell line, RA14. All three mutant RKO lines have drastically reduced ability to migrate as well as to invade through matri-gel in vitro when compared to the RKO parent and control cell lines. Exciting preliminary in vivo experiments in a colonic orthotopic mouse model, indicate that inhibiting NFκB by the IκBα super-repressor (RKO SR) has no effect on primary tumor formation but completely blocks RKO metastasis to the liver and that specific loss of IKKβ (RB3) may alter the site of metastasis. The effect of the specific loss of IKKα (RA14) on RKO tumorigenesis and metastasis is being currently investigated. We are also interrogating the specific differences in the gene expression profiles of the mutant RKO cell lines to investigate their contribution to the reduced/altered invasive and metastatic potential of these mutants. These results demonstrate that the specific inhibition of NFκB, IKKα, or IKKβ allows for the identification of the separate distinct functions of these pathway components which are required for CRC tumorigenesis and metastasis.