Immunotherapy is emerging as a promising approach in cancer treatment, allowing specific targeting of cancer cells based on surface marker expression. In vitro characterization of the effectiveness of the treatment is a necessary step before moving into clinical trials, but currently technologies rely on endpoint assays that are unable to capture the complexity and dynamics of the in vivo immune response. Here we present the adaptation of an impedance-based technology to measure immune-mediated cell killing of B lymphoid and lymphoma tumor cells. Impedance technology is able to detect alterations in cell adhesion, morphology and cell number of adherent cells by measuring changes in conductance of microelectrodes embedded in 96 and 384-wells cell culture plates. The variation is reported through an impedance-related parameter termed Cell Index (CI). The platform enables label-free and continuous monitoring of cell viability, leaving the same cells still available for secondary end point assays. To adapt such adhesion-based technology to monitor non-adherent B cells, we developed a strategy where the bottom of the wells is coated with an anti-CD40 antibody. The coating allows specific adhesion and retention of B cells and measurement of changes in impedance that are proportional to cell number. Using primary T lymphocytes expressing Chimeric Antigen Receptors (CAR) against CD19 or unrelated targets, we were able to demonstrate specific killing of tumor B cell lines and primary cells also at very low effector:target ratios. The results were also confirmed by flow cytometry. Taken together our results demonstrate the high sensitivity of this high throughput approach when compared to end point assays like 51Cr release or flow cytometry, improving the workflow for therapy validation and optimization in the context of personalized medicine.

Citation Format: Chelsea Xue, Fabio Cerignoli, Vladimir Senyukov, Piotr Pierog, Thomas Miller, Amy Shaw, Biao Xi, Lincoln Muir, Yama Abassi, Sadik Kassim. Evaluation of a novel impedance-based assay for label free and dynamic assessment of CAR-T constructs efficacy against B-cell leukemia and lymphoma [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr B090.