AN IMPROVED CIRCULARLY PERMUTED INTERLEUKIN 4-TOXIN IS HIGHLY CYTOTOXIC TO HUMAN RENAL-CELL CARCINOMA-CELLS

We have previously demonstrated that a chimeric protein composed of hu man IL-4 and Pseudomonas exotoxin, termed IL4-PE(4E), is cytotoxic to primary cells derived from human renal cell carcinoma (RCC). To improv e the cytotoxicity of IL4-toxins such as IL4-PE(4E) and IL4-PE38KDEL t o IL-4 receptor (IL-4R) positive tumor cells, a circularly permutated chimeric toxin was prepared by fusing a truncated PE gene encoding PE3 8KDEL 3' to a circularly permutated IL-4 mutant gene encoding IL4 amin o acids 38-129, the linker GGNGG, and IL4 amino acids 1-37. The result ing chimeric protein, termed IL4(38-37)-PE38KDEL, was tested on five R CC cell lines and its cytotoxicity was compared to that of the native IL4-toxins IL4-PE(4E) and IL4-PE38KDEL. IL4(38-37)-PE38KDEL was found to be 5 to 10 times more cytotoxic to all cell cultures tested compare d to either native IL4-toxin. The cytotoxic activity of IL4(38-37)-PE3 8KDEL was competable by excess IL4 and was confirmed by clonogenic ass ay. IL4(38-37)-PE38KDEL bound to IL-4R on RCC cells with 6- to 12-fold higher affinity than IL4-PE38KDEL or IL4-PE(4E), RCC tumor cells were found to lack the common gamma chain (gamma(c)) of the IL-4R reported to be present on immune cells. The stable transfection of RCC cells w ith the gamma(c), chain gene did not significantly change their sensit ivity to IL4(38-37)-PE38KDEL, Taken together, our results indicate tha t the CPIL4-toxin IL4(38-37)-PE38KDEL is highly cytotoxic to human RCC cells due to increased binding affinity to IL-4R while it is not cyto toxic or slightly cytotoxic to T and B cells, monocytic cell. lines, a nd fresh resting or activated bone marrow-derived cells. The gamma(c) does not seem to increase the internalization rate and/or processing o f IL4-toxins in RCC cells, CPIL4-toxin may be a useful agent for the t reatment of human RCC. (C) 1996 Academic Press, Inc.