Methotrexate inhibits the first committed step of purine biosynthesis in mitogen-stimulated human T-lymphocytes: a metabolic basis for efficacy in rheumatoid arthritis?
The immunosuppressive and anti-inflammatory eects of lowdose methotrexate (MTX) have been related directly to inhibition of folate-dependent enzymes by polyglutamated derivatives, or indirectly to adenosine release and}or apoptosis and clonal deletion of activated peripheral blood lymphocytes in S-phase. In this study of phytohaemagglutinin-stimulated primary human Tlymphocytes we show that MTX (20 nM to 20 lM) was cytostatic not cytotoxic, halting proliferation at G " . This stasis of blastogenesis was associated with an inhibition of purine ribonucleotide synthesis but a stimulation of pyrimidine biosynthesis, the normal mitogen-induced expansion of ATP and GTP pools over 72 h being restricted to concentrations of unstimulated T-cells, whereas the increment in UTP pools exceeded that of controls. Decreased incorporation of H"%CO’ or ["%C]glycine into purine ribonucleotides, with no radiolabel accumulation in any de noao synthetic intermediate but enhanced H"%CO’ incorporation into UTP, supported these MTX-related eects. Exaggerated ["%C]hypoxanthine salvage (which normalized the purine and UTP pools) confirmed the increased availability of 5-phospho