Synergistic effect of purine derivatives on the toxicity of pyrazofurin and 6‐azauridine towards cultured mammalian cells

A novel synergistic effect of several purine derivatives such as adenine, adenosine, hypoxanthine, and guanine on the toxicity of nucleoside analogs pyrazofurin and 6-azauridine towards cultured Chinese hamster ovary (CHO) cells has been observed. The presence of the above purine derivatives enhanced the toxicity of pyrazofurin and 6-azauridine, in a dose dependent manner. The growth inhibitory effects of these nucleoside analogs either alone or in combination with the purine derivatives were reversed by uridine and cytidine, providing evidence that the synergistic effect of the purine derivatives was exerted at the level of pyrimidine nucleotide biosynthesis. Studies with mutant cells lacking various purine phosphorylating enzymes show that phosphorylation of purine derivatives through reactions utilizing phosphoribosylpyrophosate (PRPP) is essential for observing the synergistic response. It is suggested that the above purine derivatives (including adenosine, via conversion to hypoxanthine) exert their synergistic effects by depleting the cellular pool of PRPP by two separate mechanisms (direct utilization and feedback inhibition of its synthesis), which as a result becomes rate limiting in the synthesis of orotidine monophosphate (OMP). The reduced levels of OMP, which is a competing substrate with pyrazofurin- and 6-azauridine-5'-monophosphates for binding to the target enzyme OMP decarboxylase, could then account for the inhibition of the enzyme at lower concentrations of these analogs.