Advancing Cancer Immunotherapy: Chimeric Antigen Receptor (CAR)-T Cell Engineering through Novel Screening Methods

Cancer immunotherapy represents a paradigm-shifting achievement in oncology. Particularly, chimeric antigen receptor (CAR)-T cell therapy utilizing genetically engineered T cells has produced remarkable clinical responses in hematological malignancies. However, significant challenges still remain including limited efficacy in solid tumors and critical safety concerns. The functionality of CAR-T cells depends on their synthetic receptor, CAR, which redirects T cell specificity and enhances effector functions. Therefore, optimal CAR engineering is crucial for successful development of CAR-T cell therapy. In this review, we discuss the limitations of current CAR screening methods, which primarily assess antigen binding affinity <i>in vitro</i> and often fail to predict T cell function and <i>in vivo</i> therapeutic performance. Advanced cell-based screening platforms have been developed to overcome these limitations. We overview the principles of these CAR screening systems utilizing reporter cell lines. While most are based on the detection of antigen binding properties or CAR-T cell activation markers, we emphasize a FRET-based immunological synapse biosensor as a powerful system that directly assesses CAR activation upon antigen binding. This platform offers significant advantages in speed and scalability for predicting CAR-T cell functionality. We also discuss recent advances in CAR library screening directly in primary T cells, which provides more physiologically relevant data. Such advanced platforms are essential to accelerate the development of safe and effective CAR-T therapy for solid tumors, ultimately expanding the therapeutic potential of this transformative cancer treatment.