Recommendations for High Parameter Flow Cytometry Assay Development to Understand Cell Therapy ProductsConsiderations for CAR-T Therapy Flow Cytometry Based Analytical Assays

摘要

Research in cell therapies has grown in the past few years with the FDA approval of Yescarta (axicabtagene ciloleucel) in 2017 for relapsed or refractory large B-cell lymphoma and Kymriah (tisagenlecleucel), also in 2017 for relapsed or refractory acute lymphoblastic leukemia, both of which are autologous Chimeric Antigen Receptor (CAR) T therapies. Currently research is underway for allogeneic 'off-the-shelf CAR-T cells with the cell source being the major difference between the two therapies. While autologous CAR-T therapy utilizes patients' cells, allogeneic CAR-T therapy takes advantage of the selection of healthy donors from which the cells can be isolated and, after modification, infused to patients from various genetic backgrounds. The main advantage is the availability of already modified cells that do not require the patient to wait for their own cells to be manufactured. Additionally, it allows manufacturers, to obtain cells from carefully selected donors. The products generated from either of these processes results in a live, constantly changing product that requires sophisticated analytical flow cytometry techniques. Hence, flow cytometry has become an essential component of the CAR-T therapy field and is used for the release assays that are required by the FDA, for exploratory assays to understand the product before and after infusion as well as to track it over time. The analytical strategies used to characterize the autologous and allogeneic CAR-T therapies will be different because of the source of the cells, the development process and the questions that need to be answered. In addition, the attributes may differ based on the disease indication, the target and the Mechanism of Action (MoA) of the product. However, the recommendations for generating flow cytometry-based assays will be the same. Therefore, for the purpose of this review, I will focus only on the autologous platform.