In Vitro Functional Assay Using Real-Time Cell Analysis for Assessing Cancer Immunotherapeutic Agents

Immuno-oncology is undoubtedly one of the fastest developing fields in cancer therapy, most recently propelled by successful clinical applications of chimeric antigen receptor (CAR)-based T cell therapy and monoclonal antibodies targeting immune checkpoints, namely, CTLA4, PD-1, and PD-L1. However, while the advancement of the cancer immunotherapy has brought tremendous hope to some patients, there is a pressing need for more predictive in vitro and in vivo translational models to aid in the development of novel cancer immunotherapeutic agents. As a potential solution, the xCELLigence Real-Time Cellular Analyzer (RTCA) system (ACEA Biosciences) was developed as a high-throughput, in vitro platform to monitor and screen the ability of pharmacological agents to modulate the cytolytic activity of the immune cells (i.e., the effector cell) against tumor cells (i.e., the target cell) in a real-time manner. This specialized microtiter plate system uses integrated gold microelectrodes as a sensor to measure the electronic current flow that is impeded by the tumor cells attached on the sensor surface. This measured impedance signal is dictated by cell number, size, and surface attachment strength of the adherent cells. Thus, under specific assay conditions, the xCELLigence RTCA is able to distinguish between the adherent target cancer cells and nonadherent effector immune cells. The RTCA platform has also been successfully adapted to monitor suspension target cells originated from hematological cancers. In this chapter, we outline several workflows for using the xCELLigence RTCA to evaluate the potency of clinically relevant cancer immunotherapeutic agents in modulating immune cell-mediated killing of hematological and solid tumor cells.