IN-VITRO AND IN-VIVO CHARACTERIZATION OF BR96 SFV-PE40 - A SINGLE-CHAIN IMMUNOTOXIN FUSION PROTEIN THAT CURES HUMAN BREAST-CARCINOMA XENOGRAFTS IN ATHYMIC MICE AND RATS

BR96 sFv-PE40 is a single-chain immunotoxin fusion protein targeted to the Le(Y) Ag, which is expressed in many different human carcinomas a s well as in normal gastrointestinal epithelium of humans and certain animals, including athymic rats but not mice. In vitro binding analysi s determined that BR96 sFv-PE40 was similar in affinity to BR96 Fab. B R96 sFv-PE40 internalizes rapidly, similar to BR96 IgG. H3396 cells, d erived from metastatic hu man breast carcinoma, have been established as tumor xenografts in estradiol-supplemented athymic m ice and rats. H3396 tumor xenografts established in athymic mice (up to 350 mm(3)) a nd rats (up to 100 mm(3)) completely regressed after i.v. administrati on of BR96 sFv-PE40, given as 0.625 mg/kg (1.975 mg/m(2)) every 4th da y for a total of five doses (mice) or 0.25 mg/kg (1.475 mg/m(2)) every 4th day for a total of four doses (rats). The tumors remained regress ed for the duration of the study (>85 days post-implant), which repres ents >10 doubling times, indicating that the animals were cured. There was no toxicity in rats receiving a curative dose of 0.25 mg/kg, alth ough liver and lung toxicity could be detected at a 16 times higher do se, 4 mg/kg or 23.6 mg/m(2). We conclude, therefore, that BR96 sFv-PE4 0 can cure tumor xenografts at well tolerated doses and also in the pr esence of Le(Y) expression in normal tissues.