Potency Assay Validation for Ranibizumab Using the Vascular Endothelial Growth Factor Inhibition Method

Abstract

Background: Ranibizumab is a monoclonal antibody with a complex structure used to treat ocular diseases caused by abnormalities in the retinal blood vessels. Ensuring its therapeutic efficacy requires rigorous quality control, particularly potency testing. Therefore, developing an accurate and standardized potency assay is essential. The developed and validated potency assay, in accordance with international guidelines, will establish a reliable method for regulatory quality control of ranibizumab in biopharmaceutical laboratories in Thailand.

Objective: To develop and validate a potency assay for ranibizumab using the Vascular Endothelial Growth Factor (VEGF) inhibition method, ensuring its suitability as a standard approach for quality control in support of new drug registration and biosimilar approval in Thailand.

Methods: The experimental study was conducted between October 2022 and September 2023 to develop a relative potency assay for ranibizumab based on VEGF inhibition, with results evaluated against a reference standard. The assay was validated following international guidelines (ICH guideline) by assessing specificity, accuracy, precision, linearity, range, and robustness. The %relative potency was analyzed using geometric mean (GM), geometric standard deviation (GSD), and percent geometric coefficient of variation (%GCV). Statistical differences were assessed using an independent Sample T-test and one-way ANOVA (single factor) to confirm the assay’s reliability and applicability for quality control.

Results: The VEGF inhibition assay demonstrated high specificity and reliability for assessing ranibizumab potency. The relative potency ranged from 80% to 125%, and %Recovery for accuracy was between 100.9% and 104.6%, indicating high accuracy. The %GCV for repeatability and intermediate precision were 6.8% and 6.6%, respectively, both were within the acceptable limits. The assay exhibited appropriate linearity within the 70%–130% range, with a coefficient of determination (R²) of 0.9898. Moreover, variations in HUVEC passage number and cell density did not significantly impact assay performance, confirming its robustness. All validation parameters met the acceptance criteria.

Conclusion: The validation study of the VEGF inhibition assay for ranibizumab potency assessment demonstrates the high reliability and meets international guidelines. The assay can be implemented as a standard method for ranibizumab quality control in regulatory laboratories with high efficiency.