Inhibition of orotidylate decarboxylase by 4(5H)-oxo-1-beta-D-ribofuranosylpyrazolo[3,4-d] pyrimidine-3-thiocarboxamide (APR-TC) in B lymphoblasts. Activation by adenosine kinase.

The nucleoside allopurinol riboside-3-thiocarboxamide (APR-TC; 4-(5H)oxo-1-beta-D-ribofuranosylpyrazolo[3,4,d]pyrimidine-3-thioca rboxamide) demonstrates potent in vitro antiviral activity against various DNA and RNA viruses and cytostatic activity against a variety of cell lines in culture. The IC50 for APR-TC in the splenic derived B lymphoblast cell line, WI-L2, was 0.3 microM. Adenosine kinase-deficient WI-L2 cells were resistant to growth inhibition by APR-TC, indicating that adenosine kinase (EC 2.7.1.20) is responsible for phosphorylation of APR-TC to form the monophosphate derivative (APR-TC-5'P). A 4-hr incubation of cells with 50 microM APR-TC resulted in severe depletion of intracellular pyrimidine nucleotide pools and the accumulation of 3 microM APR-TC-5'P. The cytotoxicity of APR-TC was reversed by uridine, indicating that the active form of this compound inhibits the de novo pyrimidine biosynthetic pathway. Further, APR-TC-treated cells could not utilize the pyrimidine nucleotide precursor [6-14C]orotic acid, suggesting that the UMP synthase complex is the major cellular site of inhibition. In studies utilizing cell-free lysates of WI-L2, chemically prepared APR-TC-5'P provided potent inhibition of the orotidylate decarboxylase activity (ODCase, EC 4.1.1.23) of the UMP synthase complex. APR-TC-5'P was competitive with OMP, and a Ki value of 0.35 nM was determined.