An anti-Ras cancer potential of PP1, an inhibitor specific for Src family kinases: In vitro and in vivo studies

BACKGROUND We previously found that both PAK, a Rac/CDC42-activated Ser/Thr kinase, and its binding partner PIX are required for malignant transformat ion caused by oncogenic Pas mutants, such as v-Ha-Ras. Furthermore, oncogen ic Pas requires an autocrine pathway to activate PAK. This pathway involves at least two distinct receptor kinases: EGF receptor (ErbB1) and ErbB2. In terestingly, both of these kinases are known to activate Src family kinases that phosphorylate CAT, another binding partner of PIX. PURPOSE The major aim of this study was to determine whether Src family kin ases are required for both Pas-induced PAK activation and malignant transfo rmation. For this purpose, we used PP1, an inhibitor specific for Src famil y kinases, which does not inhibit either EGF receptor or ErbB2. METHODS AND RESULTS We studied the effect of PP1 on the anchorage-dependent growth of normal and v-Ha-Ras transformed NIH 3T3 fibroblasts, PAK activat ion and anchorage-independent growth of Pas transformants, and development of Pas-induced sarcomas in nude mice. We found that PP1 (10 nM) strongly in hibits PAK activity in Pas transformants. PP1 at this concentration is know n to inhibit c-Fyn kinase, but not c-Src kinase, and none of the three know n Src family kinases (c-Src, c-Fyn, and c-Yes) expressed in fibroblasts is activated by v-Ha-Ras. Thus, it is most likely that the primary target of t his drug is an as yet unidentified Ras-activated Tyr (Y) kinase or kinases, which we call "Ray." Although PP1 has no effect on their anchorage-depende nt growth, it significantly inhibits their anchorage-independent growth in soft agar, as well as a rapid growth of Pas-induced sarcomas in mice. CONCLUSION Like EGF receptor and ErbB2, a member of Src family kinases (mos t likely a new Src-related kinase called "Ray") is essential for the Ras-in duced activation of PAK and the malignant transformation both in vitro and in vivo. These findings suggest that PP1 and other inhibitors specific for Src family kinases are potentially useful for the treatment of Pas-associat ed cancers.