Specific Activation of Glucuronide Prodrugs by Antibody-targeted Enzyme Conjugates for Cancer Therapy¹

Cancer chemotherapy may be improved by increasing antineoplastic drug specificity for tumor cells. We have synthesized a glucuronide prodrug that can be enzymatically converted to an antineoplastic agent at tumor cells that are able to bind β-glucuronidase-monoclonal antibody conjugates. The glucuronide prodrug BHAMG, the tetra-n-butyl ammonium salt of (p-di-2-chloroethylaminophenyl-β-d-glucopyranosid) uronic acid, was 150 times less toxic than the parent drug, N,N-di-(2-chloroethyl)-4-hydroxyaniline, to HepG2 human hepatoma cells and over 1000-fold less toxic than the parent drug to AS-30D rat hepatoma cells in vitro. In the presence of β-glucuronidase, BHAMG was activated and became as toxic as the parent drug N,N-di-(2-chloroethyl)4-hydroxyaniline. A conjugate (RH1-βG) was formed by linking β-glucuronidase to a monoclonal antibody which binds to an antigen expressed on the surface of AS-30D cells. The concentration of BHAMG causing 50% inhibition of AS-30D cellular protein synthesis was reduced over 1000-fold, from >770 µm to <0.74 µm after these cells were preincubated with RH1-βG. Specificity of BHAMG activation at antigen-positive cells was shown by monoclonal antibody RH1 blocking of RH1-βG conversion of BHAMG to toxic drug and by the inability of BHAMG to be converted to active drug when antigen-negative control cells were preincubated with RH1-βG. Our results show that the targeted-β-glucuronidase activation of BHAMG can increase the specificity of chemotherapy for rat hepatoma in vitro and suggest that the targeted activation of glucuronide prodrugs may be useful for cancer therapy.