Sensitive Monitoring Of CAR+ T Cells In Peripheral Blood By Flow Cytometry And Real-Time qPCR

By Stefanie Biedermann, Laura Pluisch, Inga Herfort, Ardeshir Ansari, Elisabeth Pötschke, Iris Bürger, Christian Dose, and Christiane Siewert

Autologous T cells, which are genetically modified to express a second-generation chimeric antigen receptor (CAR), are currently being investigated as a treatment for various human hematopoietic and other malignancies. Initial clinical data suggest that the persistence and expansion of CAR-T cells following autologous T cell transfer are positively correlated with effective disease clearance and protection against recurrence. Consequently, quantifying these cells in the peripheral blood of patients is a valuable tool for monitoring treatment efficacy.

Multicolor flow cytometry provides an opportunity to analyze both the presence and phenotype of circulating CAR-T cells, including the expression of CAR proteins on the cell surface during patient follow-up. This method allows for a detailed examination of CAR-T cells and offers insights into their functional status and potential therapeutic impact. On the other hand, real-time quantitative polymerase chain reaction (qPCR) is a more sensitive method for detecting transgenes in CAR+ T cells. In this poster, we present the results obtained during the assay verification of CAR qPCR. We correlated CAR-T cell detection by flow cytometry and qPCR in spike-in experiments with CAR-T cells targeting different antigens, such as CD19, CD20, or a combination of both.

Explore how this correlation helps validate the effectiveness of using both methods together for accurate monitoring of CAR-T cell therapy.