guanosine-diphosphate and tiazofurin

The ras oncogene product (p21ras, Ras) is a GTP-binding protein and is thought to transduce signals regulating cellular proliferation and differentiation. The active form Ras-GTP is inactivated by hydrolyzing bound GTP to GDP. Tiazofurin, a specific inhibitor of IMP dehydrogenase, decreased cellular GTP pools and downregulated c-ras gene expression, leading to differentiation (Olah, E. et al., Proc. Natl. Acad. Sci. USA 85: 6533-6537, 1988; Weber et al., Cancer Commun. 3:61-66, 1991). To clarify the link between the action of tiazofurin on metabolic alterations and the induction of differentiation, we examined the effect of tiazofurin on the ratio of active Ras-GTP to total Ras in K562 cells in culture. Cells were labeled for 6 h with [32P]Pi in phosphate-free RPMI 1640. Tiazofurin (100 or 200 microM) was added to cells, and samples were taken at 0, 2, 4, 6 and 12 h of incubation. Cell lysates were immunoprecipitated with monoclonal anti-p21 antibody (Y13-259), then developed on thin layer chromatography. GTP and GDP bound to Ras were visualized by autoradiography. Tiazofurin treatment decreased Ras-GTP concentration in a time- and dose-dependent fashion. In the untreated K562 cells the Ras-GTP concentration was 26.3 +/- 1.4, and tiazofurin (200 microM) decreased it at 6 h to 16.6 +/- 2.9 and at 12 h to 10.6 +/- 2.1%. Inhibition of the GTP salvage pathway with hypoxanthine (100 or 200 microM) enhanced the tiazofurin-induced decrease of Ras-GTP, whereas addition of guanosine (100 microM) prevented the Ras-GTP decrease.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Antimetabolites, Antineoplastic; Cell Differentiation; Guanosine; Guanosine Diphosphate; Guanosine Triphosphate; Humans; Hypoxanthine; Hypoxanthines; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Oncogene Protein p21(ras); Protein Binding; Ribavirin; Tumor Cells, Cultured

Since taxol (NSC 125975) and tiazofurin (NSC 286193) attack at two different sites in microtubular synthetic processes, we tested the rationale that the two drugs might be synergistic in human ovarian (OVCAR-5), pancreatic (PANC-1) and lung carcinoma (H-125) cells and in rat hepatoma 3924A cells. In human OVCAR-5, PANC-1, H-125 and rat 3924A cells, for taxol the anti-proliferative IC50 was 0.05, 0.06, 0.03 and 0.04 microM, respectively; for tiazofurin IC50 = 8.3, 2.3, 1.8 and 6.9 microM. Thus, the concentrations for taxol required for IC50 for inhibiting cell proliferation were 166-, 38-, 60- and 173-fold lower than those for tiazofurin. Taxol and tiazofurin proved synergistic in all four cell lines tested. The synergism of taxol with tiazofurin should have implications in the clinical treatment of human solid tumors with particular relevance to ovarian, pancreatic, lung and hepatocellular carcinomas.

Topics: Animals; Antineoplastic Agents; Antineoplastic Agents, Phytogenic; Carcinoma; Carcinoma, Adenosquamous; Cell Division; Drug Screening Assays, Antitumor; Drug Synergism; Female; Guanosine Diphosphate; Guanosine Triphosphate; Humans; Liver Neoplasms, Experimental; Lung Neoplasms; Ovarian Neoplasms; Paclitaxel; Pancreatic Neoplasms; Rats; Ribavirin; Spindle Apparatus; Tumor Cells, Cultured