MECHANISTIC STUDY OF THE ROLE OF TYROSINE KINASE SIGNAL

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Activation of Src and Ras has been demonstrated be closely associated with the pathogenesis and metastatic potential of many human tumors. However, the role of these oneogenes in the drug sensitivity and invasion process remain to be clarified. We examined the role of Src and Ras on these processes, using HAG-1 human epithelial cell lines transfected with v-src and activated H-ras. First we examined the potential role of Ras and Rac, a small GTPase binding protein which acts downstream of Ras, by introducing adenoviral vector containing dominant negative Ras and Rac (DN/Ras and DN/Rac) into these cells. Both DN/Ras and DN/Rac reduced the invasive potential of src-transfected cells. Moreover, DN/Rac suppressed completely the tumorigenic potentials of src-transfected cells in nude mice. These results suggest that Src, Ras, Rac, all appeared to participate in the invasion processes, and that Rac may act downstream of these signaling pathways of invasion and tumorigenicity. Next we have ex … More amined the effect of activated Src on the sensitivity to taxotere, an anticancer drug targeting microtubules. As compared with parental HAG-1 cell line, v-src-transfected HAG/src3-l cells became 7.0-fold sensitive to taxotere. The taxotere sensitivity in HAG/src3-1 cells was reversed by herbimycin A (HA), indicating that Src tyrosine kinase augments sensitivity to taxotere. Treatment of HAG/src3-1 cells with taxotere resulted in phosphorylation of Bel-2 and subsequent induction of apoptotic cell death, whereas neither Bcl-2 phosphorylation nor apoptosis occurred in parental or c-H-ras-transfected HAG-1 cells. The Bcl-2 protein is overexpressed in v-src-transfected cell line. Treatment of HAG/src3-1 cells with HA reduced the expression and phosphorylation of Bcl-2, and abrogated taxotere-induced apoptosis, suggesting a potential role for Src tyrosine kinase in the taxotere-induced apoptotic events. H-7, and wortmannin, PI-3 kinaseinhibitor, neither altered taxoteresensitivity nor inhibited taxotere-induced apoptosis in these cells. These data indicate that the ability of activated Src to increase taxotere sensitivity would be mediated by apoptotic events occurring through Src to downstream signal transduction pathways toward Bcl-2 phosphorylation, but not by activated Ras, PI-3 kinase or protein kinase C. Less