Inhibition of mannose incorporation into glycoproteins and dolichol-linked intermediates of sarcoma 180 cells by 6-methylmercaptopurine ribonucleoside

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Abstract
6‐Methylmercaptopurine ribonucleoside (6‐MMPR), an inhibitor of purine nucleotide biosynthesis de novo, was used as a model compound to evaluate the relationship between the levels of intracellular guanosine triphosphate (GTP) and the formation of cellular glycoproteins and their dolichol‐oligosaccharide precursors in Sarcoma 180 cells. Previous studies using the purine antimetabolite, 6‐thioguanine (6‐TG), demonstrated a relationship between the drug‐induced decrease in GTP levels and the incorporation of radiolabelled mannose and fucose into cellular glycoproteins; estimation of the importance of these cell‐surface alterations to the cytotoxicity produced by this agent was complicated by the incorporation of 6‐TG nucleotides into cellular DNA and RNA. In this report, evidence is presented to show that the toxicity of 6‐MMPR to Sarcoma 180 cells is associated with the effects of this agent on the intracellular pools of purine nucleotides. GTP functions in part in the activation of the sugar mannose, a step necessary for the biosynthesis of glycoproteins from nucleotide sugar precursors. Thus, 6‐MMPR, which blocks the de novo pathway of purine nucleotide biosynthesis, caused a pronounced decrease in the intracellular pools of GTP in Sarcoma 180 cells; this phenomenon was accompanied by a marked reduction in the incorporation of radiolabelled mannose into cellular glycoproteins and their dolichol‐linked oligosaccharide precursors. In contrast, the incorporation of glucosamine, a sugar not metabolically activated by GTP, into glycoproteins, and of leucine into protein, were depressed only after prolonged incubation with either 6‐MMPR or 6‐TG. Adenine restored purine nucleotide pools depleted by 6‐MMPR and partially prevented both the reduction in mannose incorporation into glycoprotein and the cytotoxic effects of this antimetabolite. Guanosine partially reversed the effects of 6‐MMPR on intracellular GTP pools and mannose incorporation but not the depression of ATP pools produced by this antimetabolite. However, guanosine did not reverse the cytotoxicity of 6‐MMPR but instead enhanced its toxicity. The findings are consistent with the possibility of membrane changes being involved in the cytotoxicity of 6‐MMPR, but clearly other factors are involved as well.