Development of a method to measure ADAs against CAR T cells

CYTO 2024 -- Chimeric antigen receptors (CARs) are engineered proteins expressed on the surface of modified T cells in order to direct the binding of resulting CAR T cells to targets expressed on malignant cells. The potential immunogenicity, including humoral and cellular-type responses, of CAR T therapies has been now broadly recognized and noted as possibly impactful on both efficacy and safety outcomes, impacting the persistence and efficacy of the treatment. Anti-drug antibodies (ADAs) are a potentially fatal immune response to many biological drugs, including CAR T cell cancer therapy. The predictability of the onset and severity of ADA production is still in the early stages of understanding. Measuring ADAs against new CAR T cell models during drug development could be a valuable indicator in determining unwanted immunogenicity. Currently, methodologies to detect antibody response to biotherapeutic compounds (ADA assays) have been well established. ADA assays are typically viewed as semi-quantitative. Commonly, a titer is reported based on the assay-specific cut-point value defined based on the statistical analysis of the treatment-native samples. The cellular nature of the CAR expression offers an opportunity to apply an alternative approach, as flow cytometry. Flow cytometry is an advantageous alternative to the currently used serological ELISA methods. Flow cytometry allows ADA detection against the CAR in its native cell membrane environment while bridging ELISA assays use only the labelled CAR fragments. In the absence of interactions from other elements of the membrane, the CAR may structurally differ, impacting the validity of the immunogenicity assessment.