The cytotoxicity of adriamycin (ADM) combined with hyperthermia (Hyp) and/or dipyridamole (DP) was investigated using B16 melanoma cells in vitro and in vivo. When ADM was combined with Hyp at 43 degrees C or DP in a dose of 5 microM or with both, drug cytotoxicity enhanced the inhibition of colony formation and cell growth, and the effect was maximal when Hyp and DP were combined. Incubation with DP alone for 24 h, following ADM exposure, enhanced the toxicity even further (p less than 0.01). Measurement of intracellular levels of ADM in the B16 melanoma cells showed that Hyp and DP increased accumulation of ADM, and that DP, but not Hyp, significantly suppressed the ADM excretion (p less than 0.05). Growth of B16 melanoma implanted into subcutaneous tissue of the foot of C57BL mice was inhibited to a greater extent by the combination treatment of ADM and DP, ADM and Hyp, and also that of ADM, Hyp and DP, compared to the findings in cases of ADM or Hyp alone. As this trimodality treatment is effective both in vitro and in vivo, further study on possible clinical advantage is warranted.