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A monoclonal antibody against human beta-glucuronidase for application in antibody-directed enzyme prodrug therapy
H J Haisma1, M Van Muijen, G Scheffer
1Department of Medical Oncology, Free University Hospital, Amsterdam, The Netherlands.
Abstract:
The selectivity of anticancer agents may be improved by antibody-directed enzyme prodrug therapy (ADEPT). The immunogenicity of antibody-enzyme conjugates and the low tumor to normal tissue ratio calls for the use of a human enzyme and the development of a monoclonal antibody (MAb) against that enzyme for rapid clearance of the conjugate from the circulation. We isolated beta-glucuronidase from human liver. BALB/c mice were immunized with the roughly purified human liver beta-glucuronidase and we obtained an MAb designated 105. Immunoblotting showed reactivity with native tetrameric human beta-glucuronidase. MAb 105 neither bound to enzyme from bovine liver, rat liver, or mouse liver nor reacted with other human lysosomal enzymes. The antibody appeared to be useful to further purify human beta-glucuronidase from human liver or human placenta to homogeneity by affinity chromatography. MAb 105 did not inhibit the activity of human beta-glucuronidase. When human beta-glucuronidase was injected i.v. into BALB/c mice, the newly generated MAb 105 could indeed accelerate the clearance of the enzyme with a 50% drop in its activity within 5 min.
Insights
Researchers developed a new monoclonal antibody (MAb 105) targeting human beta-glucuronidase. This antibody enhances the clearance of the enzyme, a key component in antibody-directed enzyme prodrug therapy (ADEPT) for cancer treatment.
Area of Science:
- Biochemistry
- Immunology
- Oncology
Background:
- Antibody-directed enzyme prodrug therapy (ADEPT) offers improved anticancer agent selectivity.
- Challenges in ADEPT include immunogenicity of conjugates and low tumor-to-normal tissue ratios, necessitating human enzymes and specific antibodies for rapid clearance.
Purpose of the Study:
- To develop a monoclonal antibody (MAb) against human beta-glucuronidase for potential use in ADEPT.
- To characterize the specificity and utility of the generated MAb for enzyme purification and pharmacokinetic modulation.
Main Methods:
- Isolation of beta-glucuronidase from human liver.
- Immunization of BALB/c mice with human beta-glucuronidase to generate monoclonal antibodies.
- Characterization of MAb 105 using immunoblotting and assessment of its reactivity with different enzyme sources and other human lysosomal enzymes.
- Evaluation of MAb 105's ability to purify human beta-glucuronidase via affinity chromatography.
- Assessment of MAb 105's effect on human beta-glucuronidase activity and its capacity to accelerate enzyme clearance in vivo.
Main Results:
- A monoclonal antibody, designated MAb 105, was successfully generated against human liver beta-glucuronidase.
- MAb 105 demonstrated high specificity, reacting only with native tetrameric human beta-glucuronidase and not with enzymes from other species or other human lysosomal enzymes.
- The antibody was effective in purifying human beta-glucuronidase to homogeneity using affinity chromatography.
- MAb 105 did not inhibit the enzymatic activity of human beta-glucuronidase.
- In vivo studies showed that MAb 105 significantly accelerated the clearance of human beta-glucuronidase from circulation in BALB/c mice.
Conclusions:
- MAb 105 is a specific and effective antibody for targeting human beta-glucuronidase.
- This antibody holds potential for improving ADEPT by facilitating rapid clearance of antibody-enzyme conjugates.
- MAb 105 can be utilized for both purification of human beta-glucuronidase and modulation of its pharmacokinetics.