The effects of leflunomide and CsA on immune responses in vitro and in vivo were investigated. Like CsA, leflunomide inhibited mitogen- or antigen-driven T cell proliferation in vitro. However, leflunomide impaired neither the capability of T cells to produce IL-2 and IL-4, nor the expression of IL-2R, that is, the acquisition of competence. In contrast to CsA, the IL-2-driven growth of secondary T cells was blocked by leflunomide. Cell cycle analyses revealed that activated T cells did not enter S phase of the cell cycle in the presence of leflunomide. Next, the effects of leflunomide and CsA on the T cell response toward the bacterial superantigen (SAg) staphylococcal enterotoxin B (SEB) were analyzed in vivo. SEB-induced early deletion (apoptosis) of a fraction of SEB-reactive V beta 8+ T cells and IL-2R expression were not impaired by either CsA nor leflunomide. On the other hand, both CsA and leflunomide prevented V beta 8-selective clonal T cell expansion and generation of SEB-specific cytolytic activity. In contrast to CsA, leflunomide treatment permitted in vivo SEB-induced production of T cell-derived lymphokines (IL-2 and TNF). Further, leflunomide failed to protect D-galactosamine-sensitized mice from SEB-induced, T cell-mediated lethal shock, whereas CsA was fully protective. Manifestation of SEB-induced T cell anergy was not impaired by leflunomide. Our results provide evidence that CsA and leflunomide differ significantly in their functional properties to suppress immune responses in that both agents inhibit T cell functions linked to clonal expansion, while leflunomide does not inhibit lymphokine secretion and thus permits lymphokine-mediated immune functions.