Involvement of deregulated epiregulin expression in tumorigenesis in vivo through activated Ki-Ras signaling pathway in human colon cancer cells

To identify the genes located downstream of the activated Ki-Ras signaling pathways in human colon cancer cells, a PCR-based cDNA subtraction library was constructed between HCT116 cells and HCT116-derived activated Ki-ms-dis rupted cells (HKe3), One of the genes in HCT116 that was evidently up-regul ated was epiregulin, a member of the epidermal growth factor family that is expressed in many kinds of human cancer cells. HKe3-stable transfectants e xpressing activated Ki-Ras regained over-expression of epiregulin, To furth er elucidate the biochemical structure and significance of epiregulin expre ssion in tumorigenesis, HKe3-stable transfectants expressing epiregulin (e3 -pSE cells) were established. Epiregulin existed as highly glycosylated mem brane-bound Forms, and TPA rapidly induced ectodomain shedding of epireguli n, Furthermore, the conditioned medium of e3-pSE cells showed more DNA synt hesis for 32D cells expressing epidermal growth factor receptor (DER) cells than that of HKe3. Although anchorage-independent growth in soft agar was not observed for e3-pSE cells, tumorigenicity in nude mice was observed evi dently, and their growth rate was correlated with each amount of exogenous epiregulin expression. These results suggested that activated Iii-Ras will be one of the factors contributing to the overexpression of epiregulin in h uman colon cancer cells, and that epiregulin will play a critical role in h uman tumorigenesis in vivo.