Recommendations for the Development and Validation of Neutralizing Antibody Assays in Support of Biosimilar Assessment

Abstract

Abstract. The American Association of Pharmaceutical Scientists (AAPS) biosimilar focus group on nonclinical and clinical assays has developed this manuscript to guide the industry on best practices and testing strategies when developing neutralizing antibody (NAb) assays for biosimilar programs. The immunogenicity assessment to biosimilar and originator drug products is one of the key aspects of clinical programs for biosimilars to demonstrate biosimilarity. Establishing that there are no clinically meaningful differences in immune response between a proposed product and the originator product is a key element in the demonstration of biosimilarity. It is critical to collect, evaluate, and compare the safety and immunogenicity data from the clinical pharmacology, safety, and/or efficacy studies especially when the originator drug product is known to have potential for immune-mediated toxicity. This manuscript aims to provide a comprehensive review and recommendations on assay formats, critical reagents, approaches to method development, and validation of the neutralizing antibody assays in extrapolation within the scope of biosimilar drug development programs. Even if there are multiple options on the development and validation of NAb assays for biosimilar programs, the type of drug and its MoA will help determine the assay format and technical platform for NAb assessment (e.g., cell-based or non-cell-based assay). We recommend to always perform a one-assay approach as it is better to confirm the biosimilarity using one-assay for NAb. If a one-assay approach is not feasible, then a twoassay format may be used. This manuscript will provide all the details necessary to develop NAb assays for biosimilars.

Comparative Immunogenicity Assessment of Biosimilars

“…In addition to the choice of screening and confirmatory assays, multiple options for detection of neutralizing antibodies (NAbs) in ADA positive samples also exist [43,44]. Depending on the biologic's mechanism of action (MoA), the selected assay may be cell-based or non-cell-based (Table 1).…”

“…Depending on the biologic's mechanism of action (MoA), the selected assay may be cell-based or non-cell-based (Table 1). While the non-cell-based (competitive ligand binding) approach offers greater precision and sensitivity and an easier set up in terms of establishing validation parameters, these assays are considered appropriate only when the biologic's MoA involves blocking of a soluble target or cellular receptor [3,11,26,44]. Additionally, cell-based approaches may be favored by certain regulatory authorities as they may offer better insight into the functional and biological activity of the product [11,26,44].…”

“…While the non-cell-based (competitive ligand binding) approach offers greater precision and sensitivity and an easier set up in terms of establishing validation parameters, these assays are considered appropriate only when the biologic's MoA involves blocking of a soluble target or cellular receptor [3,11,26,44]. Additionally, cell-based approaches may be favored by certain regulatory authorities as they may offer better insight into the functional and biological activity of the product [11,26,44]. However, cell-based experiments should not be considered the default option if sufficient sensitivity, precision and robustness cannot be achieved; in these situations, further dialogue with regulators may be necessary to define an appropriate non-cell-based method [3,26].…”

“…Similar to ADA assays, NAb assays are validated according to assay-specific control signals [44]. Additionally, during the development of the reference product, the NAb assay is likely to have been developed according to key MoAs that contribute most therapeutic efficacy as well as contemporaneous technology limitations; yet, with advancements in technology and knowledge of biological processes, it is accepted that NAb assays for biosimilars can be based on a distinct MoA, which should be a major efficacy contributor but also the most sensitive MoA for detecting differences between products [44]. As such, historic NAb data are also of limited value when assessing a biosimilar's immunogenicity profile.…”

“…Currently, no specific regulatory guidelines are available for the conduct of NAb confirmatory or titration assays as it is widely accepted that the NAb assay is a qualitative (positive or negative) experiment used for further characterization of ADA assays, which already include a confirmatory step. However, if unexpected or nonspecific responses are observed during qualitative experiments, further practical considerations for NAb confirmation (e.g., use of an agonist molecule to assess signal inhibition) or titration experiments will be required [44].…”