INHIBITION OF PLATELET-DERIVED GROWTH FACTOR-MEDIATED SIGNAL-TRANSDUCTION AND TUMOR-GROWTH BY N-[4-(TRIFLUOROMETHYL)-PHENYL] 5-METHYLISOXAZOLE-4-CARBOXAMIDE

Many reports have cited coexpression of platelet-derived growth factor (PDGF) and its receptors by tumor cells or cells supporting tumor gro wth, suggesting both autocrine and paracrine mechanisms for PDGF-media ted tumor growth. We found that a small organic molecule, N-[4-(triflu oromethyl)phenyl] 5-methylisoxazole-4-carboxamide (SU101, leflunomide) , inhibited PDGF-mediated signaling events, including receptor tyrosin e phosphorylation, DNA synthesis, cell cycle progression, and cell pro liferation. SU101 inhibited PDGF-stimulated tyrosine phosphorylation o f PDGF receptor (PDGFR) beta in C6 (rat glioma) and NIH3T3 cells engin eered to overexpress human PDGFR beta (3T3-PDGFR beta). SU101 blocked both PDGF- and epidermal growth factor (EGF)-stimulated DNA synthesis. Previously, this compound was shown to inhibit pyrimidine biosynthesi s by interfering with the enzymatic activity of dihydroorotate dehydro genase. In the current study, EGF-stimulated DNA synthesis was restore d by the addition of saturating quantities of uridine, whereas PDGF-in duced DNA synthesis was not, suggesting that the compound demonstrated some selectivity for the PDGFR pathway that was independent of pyrimi dine biosynthesis. Selectivity was further demonstrated by the ability of the compound to block the entry of PDGF-stimulated cells into the S phase of the cell cycle, without affecting cell cycle progression of EGF-stimulated cells. In cell growth assays, SU101 selectively inhibi ted the growth of PDGFR beta-expressing cell lines more efficiently th an it inhibited the growth of PDGFR beta-negative cell lines. SU101 in hibited the s.c., i.p., and intracerebral growth of a panel of cell li nes including cells from glioma, ovarian, and prostate origin. In cont rast, SU101 failed to inhibit the in vitro or s.c. growth of A431 and KB tumor cells, both of which express EGF receptor but not PDGFR beta. SU101 also inhibited the growth of D1B and L1210 (murine leukemia) ce lls in syngeneic immunocompetent mice, without causing adverse effects on the immune response of the animals. In an i.p. model of tumor grow th in syngeneic immunocompetent mice, SU101 prevented tumor growth and induced long-term survivors in animals implanted with 7TD1 (murine B- cell hybridoma) tumor cells. Because PDGFR beta was detected on most o f the tumor cell lines in which in vivo growth was inhibited by SU101, these data suggest that SU101 is an effective inhibitor of PDGF-drive n tumor growth in vivo.