Immunogenicity analysis of a biotherapeutic involves evaluating its ability to trigger an immune response, whether through the production of antibodies in the humoral system or via cellular-based immune reactions. Anti-drug Antibody (ADA) assays, including screening, confirmation, titration, and neutralizing antibody (NAb) assay development, are commonly employed throughout the drug development process. These assays aim to identify and assess antibodies binding to therapeutic products.
In the realm of protein-based biotherapeutics, this immune response spectrum ranges from the detection of clinically insignificant ADA to those with significant implications for drug safety and efficacy. Immune responses to therapeutic protein products profoundly impact both safety and effectiveness, exhibiting a wide range of clinical effects, from minor to severe adverse impacts on patient health. Detecting and analyzing ADA formation serve as invaluable tools in understanding potential patient immune responses.
Development of reliable immunogenicity assays
The evolving standards for antibody immunogenicity assay performance and the absence of standardized and validated tools for testing and reporting NAbs have led to substantial time investment by health authorities and sponsors in resolving regulatory inquiries. To tackle these challenges specific to both cell-based and non-cell-based NAbs assays, a collaborative effort involving experts from the AAPS Therapeutic Product Immunogenicity Community and representatives from the FDA and industry was initiated.
The collaborative team provides comprehensive services for standardization testing and reporting strategies. These include assistance with format selection, determining cut points, establishing assay acceptance criteria, evaluating control precision, and assessing sensitivity, which involves selecting positive controls and monitoring performance. Additionally, the team assists in selecting negative controls, conducting selectivity and specificity analyses, evaluating drug tolerance, assessing target tolerance, examining sample stability, and testing assay robustness.
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General considerations for antibody assay validation dictate that samples obtained from pivotal studies must be analyzed using fully validated assays. Upon submission of the license application, the sponsor is expected to furnish data showcasing the comprehensive validation of these assays. Validation encompasses various assessments to ensure that the ADA immunogenicity testing employed for ADA samples is suitable for its intended purpose. Critical parameters for validation encompass determining the cut-point, assessing sensitivity and drug tolerance, evaluating specificity and selectivity, measuring precision, considering reproducibility where applicable, examining the robustness of certain assay aspects, and determining the in-use stability of critical reagents.
Antibody assay Comparability Criteria: None of the established quantitative standards for evaluating comparability in NAb assays exist. Therefore, sponsors must provide scientific justification when addressing differences and similarities in sensitivity and precision and discussing reproducibility results. This should consider potential impacts on clinical outcomes across methods, studies, or laboratories. When conducting comparability assessments using study samples, criteria may encompass evaluating the positive and negative rates, aiming for ≥80% concordance across samples at appropriate low and high levels. If sufficient data allows, discussing the clinical implications of any observed differences may also be relevant. The cross-validation results should be meticulously outlined within the method validation summary table and expounded upon in the validation report or any related addendum. Additionally, any strategies to mitigate significant disparities in sensitivity, drug tolerance, or potential clinical outcomes should be elaborated upon.
Today, immunogenicity bioanalytical services specialize in immunogenicity assay development and provide a wide array of services, including ADA immunogenicity testing, ADA PK analysis, and ELISA ADA assay. Their MSD cell-based assays enhance analytical capabilities, ensuring precise analysis, while our expertise extends to gene expression analysis, offering comprehensive solutions to meet all research needs.
