The objective of the present investigation was to examine the effect of alpha-difluoromethylornithine (DFMO), an irreversible inhibitor of ornithine decarboxylase, in combination with the immunosuppressant cyclosporin A (CsA) on cytolytic T lymphocytes (CTL) induction in vitro and in vivo. Treatment with DFMO (0.2 mg/ml) or CsA (10 ng/ml) alone in vitro inhibited mitogen-induced CTL generation by 56% and 51%, respectively. Similarly, DFMO or CsA treatment alone inhibited alloantigen-induced CTL generation by 50% and 62%, respectively. Combination treatment with DFMO and CsA reduced mitogen- and alloantigen-mediated CTL induction by 79% and 90%, respectively. In vivo, DFMO treatment alone did not inhibit alloantigen induced CTL generation. However, DFMO potentiated the immunosuppressive effects of CsA in vivo on CTL induction. DFMO treatment reduced activated lymphocyte putrescine and spermidine levels by 81% and 91%, respectively. Combination treatment with DFMO and CsA, at concentrations that effectively inhibited CTL induction, did not further deplete polyamine levels beyond those levels observed with DFMO alone. CsA treatment with or without DFMO did reduce detectable levels of interleukin 2 (IL-2) activity. DFMO treatment alone did not impair IL-2 production. These results indicate that CsA and DFMO may inhibit different processes required for CTL induction, IL-2 production and polyamine biosynthesis. Therefore, inhibitors of polyamine biosynthesis may be useful in lowering the doses of CsA required to inhibit CTL induction.