Improved characteristics of a human beta-glucuronidase-antibody conjugate after deglycosylation for use in antibody-directed enzyme prodrug therapy.

Abstract

Antibody-directed enzyme prodrug therapy (ADEPT) aims at the specific activation of relatively nontoxic prodrugs into active drugs at the tumor site. One of the enzymes described to be useful in ADEPT is human beta-glucuronidase (GUS), which is expected to have low immunogenicity in patients. A major obstacle for the use of GUS, however, is its rapid glycan-specific hepatic clearance. The carbohydrates of GUS have been modified by subsequent treatment with NaIO4 and NaBH4 to improve its retention in the circulation. The modification of GUS did not decrease the enzyme activity. In vitro it was demonstrated that a conjugate prepared with a pancarcinoma specific monoclonal antibody (mAb) 323/A3 and the modified enzyme (mGUS), when bound to tumor cells, was capable of complete prodrug activation. In vivo, the 323/A3-mGUS conjugate was cleared faster from the circulation of BALB/c mice (t1/2 = 9 h) than mAb 323/A3 (t1/2 = 32 h), but it was retained in the circulation much longer than an immunoconjugate prepared with native GUS (t1/2 = 24 min). In nude mice bearing subcutaneous OVCAR-3 tumors the distribution of 323/A3-mGUS was qualitatively comparable to that of mAb 323/A3. The 323/A3-mGUS conjugate showed specific localization in the tumor but to a lesser extent than mAb 323/A3 (2.7% vs 6.4% injected dose per gram at 1 day after iv injection). A favorable tumor-to-blood ratio of > 2 was observed for the conjugate at 7 days after administration, which is necessary for tumor-specific prodrug activation.