(T0930-01-02) Advancing Biomarker Assay Validation: From Supportive Secondary Endpoint to Primary Endpoint for Regulatory Decision Making in Generalized Myasthenia Gravis

Purpose: The evolving role of Acetylcholine Receptor (AChR) antibodies as a pharmacodynamic (PD) biomarker in the development of efgartigimod for generalized Myasthenia Gravis (gMG) has necessitated significant adaptations to our assay validation strategy to meet regulatory expectations. Initially, this biomarker was used during the development of the IV product (Vyvgart) to confirm the mechanism of action (MOA) of efgartigimod in reducing disease-specific autoantibodies. For the development of the SC product (Vyvgart Hytrulo) the biomarker transitioned into the primary endpoint to demonstrate treatment response comparability between SC and IV products. This poster delves deeper into the impact of this change in context of use (CoU) on the suitability of the assay validation package and the additional validation efforts needed to meet the requirements for biomarker use in regulatory decision making.

Methods: A commercially available AChR antibody radioimmunoassay (RIA) intended for diagnostic purposes was customized by introducing additional calibrators and spiked QC samples at five different levels and was validated in a bioanalytical (BA) lab to support its intended use as a relative quantitative PD biomarker.

Results: The following validation parameters were assessed during the initial validation study: precision, spike recovery/accuracy, parallelism, and drug tolerance. Long term storage and process stability and selectivity in hemolyzed and lipemic serum were not assessed at this point, as we referred to the kit insert and literature to indicate the commonly accepted stability of immunoglobulins. The results obtained using this method were able to support a label claim that the approved IV product decreases AChR antibodies with a similar pattern to the reduction in total IgG levels relative to baseline. As such it was concluded that the method was sufficiently validated for its intended use as a relative quantitative PD biomarker. However, for the submission and approval of the SC product, the AChR antibodies were evaluated as primary endpoint to show treatment response comparability between the SC and IV product. During review of the BLA dossier of the SC product, the FDA concluded that the original method was not sufficiently validated for this CoU. The regulators cited a lack of data to demonstrate process and storage stability, as well as no data demonstrating selectivity in the presence of hemolytic or lipemic interference. Consequently, additional validation experiments were performed using incurred study samples. Room temperature and freeze-thaw stability were assessed with study samples representing a range of concentrations measured in the clinical study. Long-term stability was demonstrated using 136 incurred samples, and absence of hemolytic and lipemic interference were assessed using study samples spiked with either lysed whole blood or Intralipid. After demonstrating process and long term stability as well as selectivity in hemolyzed and lipemic samples, the method was considered sufficiently validated for its intended use and was able to support a label claim that there was no clinically significant difference between the IV and SC product based on a reduction in AChR antibodies.

Conclusion: This study underscores that biomarker method validation depends on the intended use of the biomarker. For certain purposes, the FDA requires full validation of the method in line with Bioanalytical Method Validation (BMV) guidelines. This case study exemplifies a CoU that necessitates validation according to BMV guidelines and highlights that references to kit inserts or literature may not replace experimental data. This request for additional experimental data required negotiation with the FDA on acceptable response times and major flexibility from the external BA vendor to perform unscheduled activities within short turnaround times.