Inhibition of thymidine synthesis by folate analogues induces a Fas–Fas ligand-independent deletion of superantigen-reactive peripheral T cells

Methotrexate (MTX), a folate antagonist with multiple enzymatic targets, is used in the treatment of malignancies as well as in autoimmune and chronic inflammatory diseases, and ZD1694 (tomudex), a water-soluble quinazoline specific inhibitor of thymidylate synthase (TS), is used in the treatment of adenocarcinomas. In this study, we investigated the effects of these folate analogues on superantigen (SAg)-reactive peripheral T cells in vivo. In BALB/c mice, staphylococcal enterotoxin B (SEB)-induced cytokine secretion, IL-2R (CD25) expression and early deletion of a fraction of SEB-reactive V(beta)8(+) T cells were not impaired by either MTX (7 mg/kg/day) or tomudex (5 mg/kg/day). However, both MTX and tomudex prevented V(beta)8-selective T cell expansion and accelerated their peripheral elimination. Administration of thymidine (500 mg/kg/12 h) completely abrogated this effect, indicating that inhibition of TS but not that of other folate-dependent enzymes was the main mechanism involved. Furthermore, a marked increase of apoptotic cells restricted to the V(beta)8(+) T cell subset indicated that proliferation inhibition was associated with apoptosis. In contrast with peripheral V(beta)8(+) T cell deletion, MTX and tomudex did not prevent the increase of V(beta)8(+) thymocytes triggered by SEB. Experiments in C57BL/6-lpr/lpr mice further demonstrated that deletion of V(beta)8(+) T cells induced by folate analogues was independent of Fas-Fas ligand interaction. Our results provide evidence that folate analogues may selectively delete dividing peripheral T cells through TS inhibition, but do not interfere with other events triggered by SAg.