ExPeCT™ Expansion and Persistence of CAR T Assay

For Hematology/Oncology patients this assay provides interventional data which may provide information on the expansion and persistence of the genetically modified T-cells used as therapeutic treatment for specific lymphomas and leukemias (Chimeric Antigen Receptor T-cell therapy or CAR T). However, some patients fail CAR-T therapy due to lack of cell expansion or persistence. Eurofins Viracor has developed and validated a multiplexed quantitative real-time PCR (qPCR) assay targeting the single chain variable fragment (scFv) FMC63 for the longitudinal monitoring of CD19-directed CAR T-cell therapy. 

Ideally, a pre-transfusion samples of the CAR T cells in the infusion bag and patient’s blood should be collected to establish a baseline and then monitored longitudinally. Subsequently several sources recommend routine monitoring for the first year and then annually up to five years.   

The successes with chimeric antigen receptor (CAR) T cell therapy involving patients with pre-B cell acute lymphoblastic leukemia (ALL) or B cell lymphomas have revolutionized anticancer therapy, providing a potentially curative option for patients who are refractory to standard treatments. FDA approvals of anti-CD19 CAR T cell products for both ALL and certain types of B cell lymphoma (the first approved gene therapies in the USA). CD19-targeted CAR-T cells, which have been engineered to recognize the CD19 (specifically the FMC63 scFv region) cell surface molecule of malignant B cells, show remarkable efficacy in patients with B cell acute lymphoblastic leukemia. Some patients relapse due to the CAR-T therapy not persisting in the body, leading to antigen loss, tumor escape (frameshift mutation), or exhaustion. 

In the clinical setting, long-term remission in patients with hematological malignancies is associated with sustained persistence of CAR T cells.  In the clinical oncology setting, CAR T cell expansion and persistence correlates with response and attaining remission in patients.  Because of this observation, establishing a reliable and direct method of tracking CAR T cell numbers is particularly important, not only in the estimation of effectiveness of CAR T cell therapy, but in terms of safety evaluation in post-infusion monitoring. Serious side effects of clinical CAR T cell therapy have been noted, one of the most severe being cytokine release syndrome (CRS) and is associated with CAR T cell expansion in vivo.  In addition, a major concern for recipients of allogenic CAR T cells is GvHD.  

The persistence/expansion of CAR T cells is key for determining anti-tumor efficacy, which depends on several factors such as structure of CAR T therapeutic, the manufacturing process, the lymphodepletion regimen, the infused cell phenotype, tumor burden, and disease. Persistence of CAR T-cell population is considered a critical factor of anti-tumor efficacy (therapy effectiveness).  Understanding the limitations of anti-CD19 CAR T cell therapy is critical to realizing the full potential of this treatment approach.  Growing experience with these agents has revealed that remissions will be brief in a substantial number of patients owing to poor CAR T cell persistence and/or cancer cell resistance resulting from antigen loss or modulation.  The Viracor ExPeCT™ anti-CD19 (FMC63) CAR T expansion and persistence assay is a monitoring tool to evaluate and validate the efficacy of anti-CD19 FMC63 CAR T therapy. 

CAR T cells are regulated as a gene therapy product under the FDA’s framework for biological products.

Viracor's ExPeCT anti-CD19 (FMC63) CAR T Expansion and Persistence Monitoring qPCR assay is not expected to cross-react with human genomic DNA or commensal organisms present in whole blood specimens, as determined by in silico specificity analysis. Specificity against human genomic DNA was demonstrated experimentally during validation, with no CAR vector signal detected in negative whole blood specimens.