As a world-leading authority in the field of immunogenicity testing, BioAgilytix understands the complexities that must be addressed in Antidrug Antibodies (ADA) analysis, Neutralizing Antibody (NAb) assay development and validation, isotyping, and more. Our experts will help you determine the optimal method for immunogenicity assessment, and select the optimal analytical equipment platform to yield high-quality data results.
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Immunogenicity is a significant concern for biologic drugs as it can affect both safety and efficacy. The consequences of product immunogenicity vary from no evidence of clinical effect to severe, life-threatening responses. Therefore the assessment of undesirable immunogenicity of therapeutic proteins and peptides is a key element in biological drug development. In the context of immune-mediated adverse effects of biological drugs, it is critical to understand the interplay of the adaptive and innate immune responses.
In recent years, there has been growing consensus regarding the data needed to assess product immunogenicity. Understanding the incidence, kinetics and magnitude of anti-drug antibody (ADA), its neutralizing ability, cross-reactivity with endogenous molecules or other marketed biologic drugs, and the related clinical impact, may enhance clinical management of patients treated with biologic drugs.
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Dr. Arno Kromminga discusses the emerging technologies that are making analytical and clinical data interpretation of immunogenicity more challenging, and the issues that must be addressed in the comparative immunogenicity analysis of an innovative therapeutic and biosimilar.
Our team’s scientific, medical, and technical expertise supports all aspects to assess this key aspect for the approval of the protein drugs. We understand that the measurement of adverse immune reactions is complex as it depends on many drug- and disease-specific aspects. The selection of appropriate assays and an assay strategy has to be selected specifically for each project. A generic approach for the assessment of immunogenicity is not feasible. Challenges in appropriate assay selection include the evaluation of the presence of anti-drug antibodies prior to the initiation of the treatment (pre-existing antibodies) or cross-reactivity of ADA of different isotypes.
Immunogenicity data are interpreted in the context of PK and PD data, but the clinical manifestations of the patients also must be taken into account. Sometimes the loss or reduction of drug efficacy can be observed after a long period of time, measured by disease activity scores or by MRI or other clinical parameters.
Therefore, ADA should not only be measured during drug development in pre-clinical or clinical studies, but also post-approval as a routine monitoring parameter in specialized clinical laboratories.
The cascade of immunogenicity testing is performed following a tiered approach starting with a screening assay, followed by a confirmatory assay and characterization assay.
Ultimately, all biopharmaceuticals are immunogenic and may induce anti-drug antibodies (ADA). This can impact patient safety and drug efficacy, and therefore specific and sensitive ADA detection is a key element of drug development programs, and a regulatory requirement.
Our ADA assays are fully validated in compliance with GLP regulations for the use in pre-clinical and / or clinical studies, with ADA analysis is done in a tiered-based approach. The analytical cascade starts with a sensitive ADA screening resulting in a 5% false positivity rate for clinical use. Other values for the screening cut point may be applied for non-clinical purposes. The determination of the cut point, with the help of statistical methods, is a highly complex and crucial step in this process.
Positively screened samples must be re-analyzed in a confirmatory assay. Confirmed positive ADA samples may be further characterized for their neutralizing capacity, binding affinity, isotyping, and other characteristics.
In addition to the analysis of anti-drug antibodies (ADA) and the determination of the neutralizing capacities of ADA, it may be necessary to further characterize the nature of the ADAs.
In some cases it might be helpful to know the isotype of the ADA. IgM ADA may be early markers of ADA formation. In other cases, such as oral or inhaled administration, it might be helpful to know whether the ADAs are of IgA isotypes. Importantly, the presence of IgE antibodies indicates an allergic reaction against the drug. In addition, the measurement of IgG subclasses may be supportive for the biological activities of ADA since mainly IgG1 and IgG3 are involved in complement activation and are more prone to NK cell recognition.
Furthermore, the measurement of binding affinities of ADA is informative for the ADA interpretation. Since immunoassays are end-point analyses, the availability of kinetic parameters enables a comparative analysis of ADA and their biological relevance.
Neutralizing antibodies (NAb) are anti-drug antibodies (ADA) with neutralizing capacities. While screening assays by definition are intended to measure ADA in total, the further characterization of ADA following a tiered-based assay strategy is required to understand the relevance of ADA formation.
We provide different assay formats, techniques, and methods for the measurement of the neutralizing capacity of ADA. Neutralizing antibodies (NAb) can be measured by cell-based assays or by competitive ligand binding (inhibitory) assays (CLBA). The selection of the most appropriate assay format depends on different factors such as the mode of action of the drug. While a cell-based assay usually reflects more realistically the situation in vivo, CLBAs are often (but not always) more robust and demonstrate lower variances. On the other hand, gene reporter assays and CLBAs often show comparable variances.
In our current understanding, cell-based assays are the first choice for NAb analysis. CLBAs can be used if the data are provided that CLBAs show equal or even better results. There is an ongoing scientific debate about the most appropriate assay format and we appreciate any input on this topic.
The assessment of immunogenicity can leverage a variety of analytical equipment platforms. Each of these platforms possesses unique strengths, weaknesses and therefore, optimal uses. BioAgilytix has experience immunogenicity assay development across the bioanalytical lifecycle requiring high drug tolerance levels. We can recommend strategies and provide contract services for assessing immunogenicity using a variety of technologies. The platforms we typically leverage are listed below.
ELISA
MSD-ECL
Gyrolab
Biacore
ImmunoCAP
Flow Cytometry
(FACS Analysis)
Luminex
Immunogenicity and bioanalytical assessments are becoming more challenging as the immune system is exposed to more complex biologics. Learn more about this evolving topic in a recent article from one of our experts.
Our experts understand the complexities that must be addressed in immunogenicity testing, including isotyping, ADA analysis, NAb assays, and more, and will help you determine the optimal analytical equipment platform to yield high-quality data results. Specifically, our immunogenicity services include labeling of antibody reagents, ADA purification, and oversight of surrogate positive control antibody generation. Our scientists are experienced in supporting immunogenicity studies in a variety of species including rodents, humans, and non-human primates.
Our Immunogenicity Expertise Includes:
See how our PK expertise complements our immunogenicity services by evaluating absorption, distribution, metabolism and excretion characteristics of a product.
Learn how we will translate our experience working with over 500 biomarkers in singleplex and multiplex formats to your project’s success.
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