Effects of 6-azauridine on nucleotides, orotic acid, and orotidine in L5178Y mouse lymphoma cells in vitro.

Abstract

Murine lymphoma L5178Y cells in culture were exposed to 5 µm 6-azauridine (6-aza-UR) for up to 8 hr. The nucleotides in the acid-soluble extracts were determined by high-pressure liquid chromatography. The concentrations of uracil and cytosine nucleotides decreased rapidly in the treated cells. Adenosine 5′-triphosphate and adenosine 5′-diphosphate increased steadily, whereas guanosine 5′-diphosphate decreased for a few hr before returning to the normal level. Orotic acid and orotidine, but no detectable amount of orotidine monophosphate (OMP), accumulated in treated cells. In homogenates of the cells harvested from ascitic fluids of mice, the presence of a membrane-bound phosphatase with activity for OMP was demonstrated, which may be responsible for the lack of OMP accumulation. When incubated with the homogenate, [7-14C]OMP was converted to [7-14C]orotidine in the absence of inorganic phosphate, but it was converted to [7-14C]orotic acid in the presence of phosphate, suggesting the occurrence of an orotidine phosphorylase. The increase in adenosine 5′-triphosphate levels and the rapid decrease in the pyrimidine nucleotides observed after treatment with 6-aza-UR may be responsible for the observation (Brenckman et al. , Biochem. Biophys. Res. Commun., 52: 1368–1373, 1973) that 6-aza-UR potentiates the activity of 1-β-d-arabinofuranosylcytosine only when 6-aza-UR is added prior to 1-β-d-arabinofuranosylcytosine.