Immunogenicity assays are essential in clinical drug development, particularly for biotherapeutic products. Despite the regulatory focus on clinical requirements, nonclinical immunogenicity assessment remains critical under specific circumstances. According to ICH S6, nonclinical immunogenicity should be evaluated when there is significant pharmacodynamic activity alteration or when drug exposure changes without pharmacodynamic markers or immune-mediated response indicators.
Minimum Strategy Consensus by the European Bioanalysis Forum (EBF)
The EBF established a working group in 2016, resulting in a consensus on minimal strategies for nonclinical immunogenicity evaluation. Key parameters include SCP evaluation, sensitivity, drug tolerance, and precision:
1. Cut-off Values: For screening thresholds, a False Positive Rate (FPR) of 0.1-1% is recommended without confirmatory testing. A 0.1% FPR helps exclude false positives near the SCP signal, effectively detecting relevant nonclinical immune responses. Adhering to the 3R principle, the SCP can be determined using fewer individual samples, ideally 15-30, generating 60 data points (e.g., 30 samples analyzed twice or 15 samples analyzed four times). Pre-dosing sample FPR confirmation is unnecessary due to its limited nonclinical relevance.
2. Sensitivity: Sensitivity should be assessed within a single analysis batch. The MRD should ideally be below 100, with sensitivity under 1000 ng/mL. According to NMPA 2021 guidelines, sensitivity should reach 250-500 ng/mL, with sub-100 ng/mL sensitivity being preferable if achievable. LPC concentration can be set near 2-3 times the SCP signal without extensive statistical analysis.
3. Precision: Precision should be evaluated across three analysis batches, each including six sets of independently prepared positive controls at three concentration levels (HPC, MPC, LPC) and six sets of negative controls (NC).
4. Drug Tolerance Levels: Drug tolerance should be evaluated at LPC (or 1000 ng/mL positive control) levels, ensuring the method can detect ADA at expected drug levels in nonclinical studies.
Optional validation parameters include selectivity, specificity, titers, and stability, tailored to individual project needs.
Addressing Industry Challenges
The "minimum strategy" faces practical implementation challenges, particularly due to the industry's longstanding reliance on clinical guidance for nonclinical immunogenicity studies. For example, the industry often hesitates to adopt strategies like "screening only with a 0.1-1% FPR without confirmatory tests." Nonetheless, defining cut-off values, sensitivity, drug tolerance, and precision remains crucial.
Recommendations for Nonclinical Immunogenicity Assays
Given the recognized lack of translatability between nonclinical and clinical data, the EBF advises the following:
1. Case-by-Case Assessment: Nonclinical anti-drug antibody (ADA) assessments should be based on scientific rationale and data utility, rather than being a default requirement.
2. Pharmacokinetic/Pharmacodynamic (PK/PD) Studies: These studies, conducted before GLP toxicology, may provide insights into immune responses but have limited value.
3. Toxicology Studies: The primary goal is to demonstrate sufficient exposure and assess resulting pathologies and their reversibility, defining no effect levels and safety margins for clinical use. ADA may or may not impact TK/PD or cause observed pathologies.
4. Decision Making: The likelihood of generating an immune response in vivo should guide assay development activities, balancing business risks and sponsor timelines.
Immunogenicity Strategy Approaches:
- Bank samples and develop/validate assays only if TK/PD and safety impacts are observed.
- Develop assays and initiate validation/testing upon observed impacts on TK/PD and safety.
- Proactively develop and validate assays based on business risks and resources.
A lean approach can be taken for assay validation, avoiding the full requirements of clinical assays and applying 3R principles. Screening assays with stringent cut points (0.1-1% FPR) or signal/noise approaches can suffice for sample analysis.
By adopting these optimized strategies, the nonclinical immunogenicity evaluation can become more efficient and aligned with scientific and regulatory expectations, ultimately enhancing the development of biotherapeutic products.
Keywords:
Immunogenicity assays
Nonclinical immunogenicity
Anti-drug antibodies (ADA)
Pharmacokinetics (PK)
Pharmacodynamics (PD)
Toxicology
Sensitivity
Precision
European Bioanalysis Forum (EBF)
ICH S6 guidelines
