EFFECTS OF LEFLUNOMIDE AND OTHER IMMUNOSUPPRESSIVE AGENTS ON T-CELL PROLIFERATION IN-VITRO

Leflunomide and its active metabolite, A771726, are structurally unrel ated to immunosuppressive agents currently under investigation Previou s in vitro studies have revealed that leflunomide primarily inhibits i nterleukin-2-stimulated T cell proliferation. In the current study, we have extended our previous work and demonstrate that leflunomide prev ents T cell progression induced by phytohemagglutinin into the S phase of the cell cycle. To discriminate further the action on T cells of l eflunomide from other immunosuppressive agents, we performed kinetic s tudies where leflunomide was added either after the initiation of mixe d lymphocyte cultures (MLC) or after interleukin-2 stimulation of CTLL -4 cell proliferation. These studies revealed that leflunomide acted c omparably to rapamycin, but was distinct from brequinar sodium in the MLC, and from cyclosporine and mycophenolic acid in both MLC and CTLL- 4. Although previous biochemical studies indicated that leflunomide ca n inhibit src-family tyrosine kinase activity, more recent studies hav e suggested that leflunomide can also inhibit pyrimidine synthesis. Ou r data demonstrate that the ability of leflunomide (25-100 mu M) to in hibit MLC and CTLL-4 cell proliferation is partially antagonized by ur idine (25-100 mu M), and support the hypothesis that leflunomide inhib its pyrimidine synthesis in T cells. Unique molecular mechanisms of im munosuppression suggest that drug combinations may result in synergist ic immunosuppression. Our in vitro studies revealed synergistic inhibi tion of T cell proliferation with the combinations of leflunomide with cyclosporine or with rapamycin. We have extended those studies to qua ntitate inhibition of MLC by the combinations of leflunomide and brequ inar sodium or mycophenolic acid.