We have previously demonstrated that functional high-affinity interleukin-4 receptors (IL-4R) are expressed on human renal cell carcinoma (RCC) cells (N. I. Obiri et al., J. Clin. Invest. 91, 88, 1993). In the present study, we examined the cytotoxic effect (determined by inhibition of protein synthesis) of a chimeric protein composed of human IL-4 and Pseudomonas exotoxin (PE) on human RCC tumor samples obtained from patients undergoing nephrectomy. The chimeric gene encoding hIL4-PE4E was constructed by fusing a cDNA clone for human IL-4 to the 5' end of a mutated cDNA encoding a full-length PE molecule. This gene was expressed in Escherichia coli, and large quantities of this recombinant protein were isolated to more than 95% purity. This chimeric protein, hIL4-PE4E, was highly cytotoxic to all six RCC cell lines examined. The concentration of hIL4-PE4E at which 50% inhibition of protein synthesis was obtained ranged from < 1 ng/ml (12 pM) to 10 ng/ml (120 pM) in five of the six isolates of RCC and 40-70 ng/ml in one other. A mutant chimeric protein which can bind to IL-4R but lacks the ADP ribosylation activity of PE was not cytotoxic to the RCC cells. The cytotoxic effect of hIL4-PE4E was IL-4R mediated because a fourfold molar excess of IL-4 abrogated the cytotoxic effect of hIL4-PE4E. A neutralizing monoclonal antibody to IL-4 also abrogated the cytotoxic effect of hIL4-PE4E. hIL4-PE4E showed very little cytotoxic activity to a normal human umbilical vein endothelial cell line (ID50 = 1000 ng/ml) and a human fibroblast cell line (ID50 approximately 400 ng/ml). Nonactivated human peripheral blood lymphocytes (PBL) were also insensitive to hIL4-PE4E (ID50, approximately 500 ng/ml), whereas phytohemagglutinin-activated PBL were highly susceptible to the cytotoxic effect of hIL4-PE4E (ID50, approximately 4 ng/ml). These data indicate that hIL4-PE4E may be a useful agent for the treatment of human RCC without affecting normal and resting immune cells.