Interleukin 2 (IL‐2) administration is known to induce marked eosinophilia. To evaluate the potential role of eosinophils as anti‐tumor effectors and to understand the direct or indirect effects of IL‐2 on eosinophils, the physical and functional characteristics of eosinophils obtained during IL‐2 therapy were compared with those of eosinophils obtained from the same patients before IL‐2 administration, or from healthy donors. The treatment schedule consisted of subcutaneous (s.c.) injections of IL‐2, and was performed in 7 patients with small‐cell lung cancer (SCLC) in advanced stage. A marked increase of hypodense cells in peripheral blood was found to correlate with eosinophil activation in patients undergoing IL‐2 therapy. Cyto‐toxic activity of eosinophils against allogeneic tumor cells (SCLC, K562 and melanoma lines), as assessed by direct and antibody(Ab)‐dependent cellular cytotoxicity (ADCC), was markedly increased during IL‐2 therapy. Conversely, eosinophils obtained before treatment, like those of healthy donors, lacked any activity against tumor cells. Sera from IL‐2‐treated, but not from untreated, patients, significantly improved the in vitro survival and anti‐tumor cytotoxicity of eosinophils from healthy donors. Comparable effects were obtained with eosinophils cultured with interleukin 5 (IL‐5), granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) and, to a lesser extent, by tumor necrosis factor‐α (TNFα), while no direct activity was mediated by IL‐2. A 91% inhibition of eosinophil ADCC was found after pre‐incubation of the sera of IL‐2‐treated patients with anti‐IL‐5 but not with anti‐GM‐CSF or anti‐TNFα Ab. IL‐5 mRNA expression was detected in peripheral‐blood lymphocytes (PBL) obtained 4 hr after IL‐2 injection during the second and third week of IL‐2 therapy. Phenotypic analysis of eosinophils from IL‐2‐treated patients showed enhanced expression of activation markers, including Fcγ RII (CD32), HLA‐DR, CR3 (CD I lb) and CRI (CD35). These findings suggest that a significant cytotoxicity against tumor cells can be mediated by eosinophils after indirect, IL‐5‐mediated in vivo activation by IL‐2, and that eosinophils may be involved in the anti‐tumor response(s) induced in vivo by IL‐2.