TYROSINE KINASE INHIBITORS INHIBIT MULTIPLE STEPS OF THE CELL-CYCLE OF VASCULAR SMOOTH-MUSCLE CELLS

Protein tyrosine kinase (PTK) inhibitors have been reported to inhibit proliferation of vascular smooth muscle cells (SMC), To elucidate the mode of this inhibition, the effects on the cell cycle of cultured vascular SMC of three PTK inhibitors with different modes of action (methyl 2,5-dihydroxycinnamate, genistein, and herbimycin A) were studied. Rat aortic SMC were synchronized to the GO phase of the cell cycle and then released to proceed through the cell cycle by the addition of platelet-derived growth factor (PDGF), and [H-3]thymidine incorporation into DNA was measured. The three PTB: inhibitors all inhibited PDGF-induced DNA synthesis in a dose-dependent fashion, with IC50 values of 4.7 +/- 1.4 mu M for methyl 2,5-dihydroxycinnamate, 6.7 +/- 2.5 mu M for genistein, and 0.17 +/- 0.07 mu M for herbimycin A. Time course studies suggested that the agents inhibited early G1 phase but not the G0-G1 transition. The lack of effect on the G0-G1 transition was also supported by the finding that the agents did not inhibit the ligand-induced autophosphorylation of PDGF receptor nor the induction of c-fos mRNA at concentrations which were sufficient to inhibit DNA synthesis. PTK inhibitors inhibited progression of the S phase when they were added to SMC that had been arrested at the G1-S border with hydroxyurea, Methyl 2,5-dihydroxycinnamate also blocked the M phase when it was added to SMC cultured in the presence of 10% fetal calf serum, while genistein and herbimycin A did not inhibit the M phase under the same experimental conditions. In accordance with our previous observation, methyl 2,5-dihydroxycinnamate impaired microtubule networks and formation of the mitotic spindle during the M phase. Our findings indicated that PTK inhibitors inhibit multiple steps of the vascular SMC cell cycle. (C) Academic Press, Inc.