A chemotherapy regimen consisting of dacarbazine, vincristine, bleomycin and lomustine in combination with natural leukocyte interferon (IFN)-a was administered to 48 metastatic melanoma patients with very favorable results. The frequencies of peripheral blood lymphocyte subsets during this treatment were monitored by flow cytometry in order to elucidate the underlying antitumor mechanism. Analysis of 467 peripheral blood samples revealed a significant decrease during treatment in absolute values of all lymphocyte subsets (CD4, CD8, CD56) as well as in total lymphocyte count. All the values returned to normal after discontinuation of treatment. Significant alterations in proportions of lymphocyte subsets were detected. Notably, the CD4+/CD8+ ratio showed changes related to response, treatment duration, and disease progression. Exceptionally high CD4+/CD8+ ratios (up to 14) were observed soon after the start of treatment in some of the patients later achieving a complete response. During the course of treatment, in 76% of the patients receiving at least a 2-month treatment, the CD4+/CD8+ ratio decreased to less than 1.0. Peculiarly, the nadir CD4+/CD8+ ratio was consistently observed 1-6 months before clinically detectable disease progression, suggesting a relation between CD4+/CD8+ ratio and tumor control.