Development and validation of novel enzyme activity methods to assess inhibition of matrix metalloproteinases (MMPs) in human serum by antibodies against enzyme therapeutics

• Published in: Journal of pharmaceutical and biomedical analysis Vol. 70; pp. 408 - 414
• Main Authors: Edkins, Thomas J., Alhadeff, Jack A., Kwok, Vincent, Kalensky, Charles, Rock, Marie T., Vidmar, Thomas J., Del Tito, Benjamin J.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.11.2012; Elsevier
• Subjects: Analysis; Analytical, structural and metabolic biochemistry; antibodies; Antibodies, Bacterial - blood; Antibody Specificity; Biological and medical sciences; Biological Assay - methods; Biological Assay - standards; blood serum; Calibration; Clostridium histolyticum; Clostridium histolyticum - enzymology; collagenase; Cross Reactions; Cross-reactivity of anti-drug antibodies; enzyme activity; Enzyme Stability; Enzyme therapeutics; Fundamental and applied biological sciences. Psychology; gelatinase A; General pharmacology; Humans; Hydrogen-Ion Concentration; ice; interstitial collagenase; Matrix Metalloproteinase 1 - analysis; Matrix Metalloproteinase 1 - immunology; Matrix Metalloproteinase 13 - analysis; Matrix Metalloproteinase 13 - immunology; Matrix Metalloproteinase 2 - analysis; Matrix Metalloproteinase 2 - immunology; Matrix Metalloproteinase 3 - analysis; Matrix Metalloproteinase 3 - immunology; Matrix Metalloproteinase 8 - analysis; Matrix Metalloproteinase 8 - immunology; Matrix Metalloproteinase Inhibitors - pharmacology; Matrix metalloproteinases; Matrix Metalloproteinases - analysis; Matrix Metalloproteinases - immunology; Medical sciences; Microbial Collagenase - immunology; Microbial Collagenase - therapeutic use; MMP enzyme activity inhibition assay method development and validation; Pharmacology. Drug treatments; Recombinant Proteins - analysis; Recombinant Proteins - antagonists & inhibitors; Recombinant Proteins - immunology; Reference Standards; Reproducibility of Results; stromelysin 1; Temperature; therapeutics; Time Factors; Enzyme therapeutics; Matrix metalloproteinases; Cross-reactivity of anti-drug antibodies; MMP enzyme activity inhibition assay method development and validation; Human; Drug; Validation; Biological fluid; Enzyme; Antibody; Metalloendopeptidases; Peptidases; Treatment; Enzymatic activity; Cross reaction; Development; Hydrolases
• ISSN: 0731-7085; 1873-264X
• DOI: 10.1016/j.jpba.2012.06.012

This paper summarizes the development and validation of five enzyme activity methods to assess the specific inhibition of human endogenous matrix metalloproteinases MMP-1 (interstitial collagenase), MMP-2 (gelatinase A), MMP-3 (stromelysin 1), MMP-8 (collagenase 2) and MMP-13 (collagenase 3) by anti-Collagenase Clostridium histolyticum (CCH) antibodies in human serum. These MMPs are of interest since antibodies against a therapeutic enzyme may cross-react with, and inactivate, the MMPs. The validated methods utilize spiked exogenous individual MMPs added to serum to determine if the serum inhibits MMP enzyme activity. Factors evaluated and optimized during development include pH, reaction time and temperature, inhibitor concentration for the positive control, and substrate and serum concentration. Characteristics established during validation for each MMP activity inhibition method included intra- and inter-assay precision and recovery, recovery in the pooled normal human serum samples, bench-top stability at room temperature and on wet ice, and assay cut-point determination. Precision results ranged from ∼1 to 12% CV, recoveries of the activities of the exogenous MMPs ranged from ∼84 to 90% and cut-point values ranged from 67 to 91%.