THE INFLUENCE OF LEFLUNOMIDE ON CELL-CYCLE, IL-2-RECEPTOR (IL-2-R) AND ITS GENE-EXPRESSION

Leflunomide is a novel immunomodulatory drug shown to be very effectiv e in animal models of autoimmune diseases and transplantation rejectio n, as well as in human rheumatoid arthritis. Leflunomide's main metabo lite, A77 1726, has been shown to be reversibly antiproliferative in v itro. Pursuing this, we performed cell cycle analysis by flow cytometr y of a B-cell lymphoma line and found that at concentrations > 2.5 mu M cells accumulated in the early S-phase. In order to determine A77 17 26's effects on cell activation, human peripheral blood lymphocytes (P BL) were cultured in the presence of PHA or OKT 3 antibody. Flow cytom etric evaluation of IL-2 and transferrin receptor expression exhibited a dose-dependent inhibition of these activation markers (10-100 mu M) . Further, using the polymerase chain reaction, we investigated the ab ility of leflunomide to impair the transcription of the IL-2-R gene. W e found that A77 1726 did not significantly decrease IL-2-R alpha-chai n mRNA expression regardless of stimulation. It seems that leflunomide 's main metabolite did not affect IL-2-R at the level of gene transcri ption, and thus its effects could be due to impairment of post-transla tional events. Taken together, these studies demonstrate that leflunom ide: (1) impairs activation of quiescent lymphocytes (reduction of IL- 2-R expression), without inhibiting IL-2-R mRNA, formation; (2) revers ibly inhibits proliferation by holding resting lymphocytes in the G(0) -phase and activated cells in the early S-phase of the cell cycle. Thu s, this compound may exert its effects through influencing two importa nt aspects of an immune response, i.e. activation and proliferation of lymphocytes.