IN-VITRO BIOLOGICAL CHARACTERIZATION AND ANTIANGIOGENIC EFFECTS OF PD-166866, A SELECTIVE INHIBITOR OF THE FGF-1 RECEPTOR TYROSINE KINASE

Through direct synthetic efforts, we discovered a small molecule that is a nanomolar inhibitor of the human fibroblast growth factor-1 recep tor(FGFR) tyrosine kinase. PD 166866, a member of a new structural cla ss of tyrosine kinase inhibitors, the 6-aryl-pyrido[2,3-d]pyrimidines, was identified by screening a compound library with assays that measu re protein tyrosine kinase activity. PD 166866 inhibited human full-le ngth FGFR-1 tyrosine kinase with an IC50 value of 52.4 +/- 0.1 nM and was further characterized as an ATP competitive inhibitor of the FGFR- 1. In contrast, PD 166866 had no effect on c-Src, platelet-derived gro wth factor receptor-beta, epidermal growth factor receptor or insulin receptor tyrosine kinases or on mitogen-activated protein kinase, prot ein kinase C and CDK4 at concentrations as high as 50 mu M. PD 166866 was a potent inhibitor of basic fibroblast growth factor (bFGF)-mediat ed receptor autophosphorylation in NIH 3T3 cells expressing endogenous FGFR-1 and in L6 cells overexpressing the human FGFR-1 tyrosine kinas e, confirming a tyrosine kinase-mediated mechanism. PD 166866 also inh ibited bFGF-induced tyrosine phosphorylation of the 44- and 42-kDa (ER K 1/2) mitogen-activated protein kinase isoforms in L6 cells, presumab ly via inhibition of bFGF-stimulated FGFR-1 tyrosine kinase activation . PD 166866 did not inhibit platelet-derived growth factor, epidermal growth factor or insulin-stimulated receptor autophosphorylation in va scular smooth muscle, A431 or NIHIR cells, respectively, further suppo rting its specificity for the FGFR-1. In addition, daily exposure of P D 166866 to L6 cells at concentrations from I to 100 nM resulted in a concentration-related inhibition of bFGF-stimulated cell growth for 8 consecutive days with an IC50 value of 24 nM. In contrast, PD 166866 h ad little effect on platelet-derived growth factor-BE-stimulated growt h of L6 cells or serum-stimulated vascular smooth muscle cell prolifer ation. Finally, PD 166866 was found to be a potent inhibitor of microv essel outgrowth (angiogenesis) from cultured artery fragments of human placenta. These results highlight the discovery of PD 166866, a new n anomolar potent and selective small molecule inhibitor of the FGFR-1 t yrosine kinase with potential use as antiproliferative/antiangiogenic agent for such therapeutic targets as tumor growth and neovascularizat ion of atherosclerotic plaques.