What are Anti-drug antibodies (ADAs) assays?
Biotherapeutics use a variety of mechanisms of action, such as the interfering cell-signaling pathway, to inhibit the specific proteins necessary for disease progression. Despite their curative promise, these biologicals may trigger an immune reaction, resulting in anti-drug antibodies (ADAs) that bind to specific regions of the therapeutic protein. Among these, neutralizing antibodies (NAbs) are mainly concerned with blocking or completely neutralizing the pharmacological potential by interacting with the areas crucial to their function. The clinical implications of ADAs, particularly NAbs, range from a reduction in efficacy, which can be managed with alternative treatments, to severe side reactions such as hypersensitivity or intervention with endogenous protein. Regulatory agencies, such as the FDA and EMA, mandate the thorough monitoring and characterization of ADAs to evaluate a drug’s immunogenicity, safety, and efficacy in clinical trials. Their guidelines advocate a multi-tiered approach, starting with a Screening Assay to detect ADA presence, followed by a Confirmatory Assay to ascertain specificity. Confirmed positive samples are further analyzed using Titer Assays to assess their capacity to inhibit drug function.
Advanced analytical options, such as isotype differentiation, provide a profound understanding of the immune reaction. To produce reliable data, assays must be carefully designed to be sensitive, specific, precise, and robust. For the detection of ADA, various assay formats are used, e.g., Enzymatic-linked immunosorbent assay (ELISA), electrochemiluminescence assay (ECL), radioimmunoassay (RIA), radioimmunoprecipitation assay (RIPA), and surface plasmon resonance assay (SPR). These technologies are unique in their ability to capture and monitor immune responses. As biotherapeutics continue to be used more and more, strict monitoring of ADAs and their analysis remains important for ensuring continued safety and medicinal efficacy.
Importance of ADA on medicines safety and potency
ADAs produced by biological therapeutics can significantly influence pharmacokinetics, pharmacodynamics, clinical efficacy, and tolerance safety and require thorough immunogenicity ADA risk assessment during clinical evaluation. Various assay methods, including ELISA, radioimmunoassay, and electrochemistry, with a bridging assay, are currently the standard ADA assay screening and confirmation method. The present widely used immunoassay layout, where the product is labeled with a unique hapten or tag, enables the detection of all ADA isotypes (IgG, IgM, IgA, etc.) throughout the species by forming an immune complex. The first step in identifying the ADA immunogenicity testing method is to screen and confirm the presence of the ADAs. This phase is followed by developing an NAb assay to determine the neutralizing antibody binding assay, which better reflects the second in vivo mechanism of action.
Measurement of immunogenicity is essential for both preclinical and clinical development of drugs, as ADAs may trigger unwanted immune responses that compromise efficacy and tolerance safety. As clinical ADA assays require a drug-specific reagent and a strict three-tiered method, preclinical assays propose greater flexibility guided by ICH S6 recommendations for assessing altered pharmacodynamics or immune-mediated reactions. For simplifying preclinical testing, generic ADA assays provide a cost-effective alternative using commercially available reagents, reducing growth time and resources. Moreover, lean ADA assay provides productive options for early investigations, particularly in cases where appropriate controls are not available, in addition to simplifying the ADA immunogenicity test and accelerating drug development.
Emerging technologies for ADA assays: Useful in clinical trials
Advances in immunogenicity assay development by improving drug tolerance, sensitivity, and automation are emerging as novel ADA assays. The Gyrolab platform combines microfluidics and automation to perform the ADA Immunogenicity Test with minimal sample volume and reagent consumption. However, their short incubation times may restrict the detection of low-affinity antibodies, impacting assay validation. Immune-PCR (iPCR) enhances drug tolerance by exploiting high dilution rates and high sensitivity to detect ADA even in the presence of a circulating drug. During iPCR, it is essential to improve sensitivity and drug tolerance; it relies on a proprietary reagent and requires extensive training for analysts. The stages that admire SQI SquidLite and Genalyte Maverick enable simultaneous ADA detection and isotyping, increased throughput, and elimination of the need for several assays. However, they demand high device costs and vendor-dependent optimization. LC/MS immunogenicity bioanalytical services provide multiplexing and specificity advantages for developing the NAb assay, but obstacles remain similar to endogenous Ig intervention and interlaboratory variability. Generic ADA assays, identical to immune complex ELISAs and universal ADA ECL immunoassays, provide a cost-effective option for nonclinical analysis, reducing the need for drug-specific reagents and expediting the timely selection of drug molecules. While the technologies mentioned above improve performance, the individual has a single limitation and requires careful selection based on the requirements of the study, the characteristics of the molecules, and regulatory requirements.
Conclusion
Industry and regulators broadly accept the interpretation of immunogenicity information from ADA assays to approve biotherapeutics. During the development of technologies that enhance ADA isotyping, sensitivity, and drug tolerance, data from market products suggest that highly drug-tolerant assays do not always correlate with clinical efficacy. Regulatory authorities demand profound perceptions of immunogenic responses to ensure continued safety. The choice of technologies under consideration here provides the power to overcome the limitations of the ADA assay and provide a more complete picture of the anti-drug antibody.
