Abstract A25: Humanized immunoenzyme with enhanced activity for glucuronide prodrug activation in the tumor microenvironment

Abstract

Abstract Antibody-directed enzyme prodrug therapy (ADEPT) is one of potential therapies to overcome systemic toxicity caused by traditional chemotherapy. Our lab have designed and developed glucuronide prodrugs that can be converted to potent antineoplastic agents by β-glucuronidase (βG). Toxic drugs can be preferentially generated at the tumor site after βG is first targeted to tumor cells as an antibody-enzyme fusion protein (immunoenzyme). For utilization of this therapeutic strategy in clinic, the immunoenzyme need to be compatible with humans. We have recently employed directed evolution to generate a human βG mutant (S2) that enhanced activity at neutral pH. To investigate if the S2 variant would be beneficial for tumor-selective prodrug activation, a humanized cc49 scFv gene with specificity for the TAG-72 tumor-associated antigen was fused to the wild-type hβG or S2 genes to create hcc49-hβG and hcc49-S2, respectively. Here, we show that immunoenzymes have 100-folds lower KD than hcc49 scFv and retain enzymatic activities as wild-type hβG and S2 mutant proteins. In in vitro cytotoxicity assay, the data demonstrated that hcc49 immunoenzymes can preferentially activate glucuronide prodrugs at TAG-72 positive cells, and hcc49-S2 was about 10-fold more effective than hcc49-hβG as indicated by a lower IC50 value. Further in vivo experiment, we will examine if the enhanced immunoenzyme have better efficacy to hydrolyze anticancer glucuronide prodrugs at physiological condition. Citation Information: Clin Cancer Res 2010;16(7 Suppl):A25