Cryopreservation of NK and T Cells Without DMSO for Adoptive Cell-Based Immunotherapy

Dimethylsufoxide (DMSO) being universally used as a cryoprotectant in clinical adoptive cell-therapy settings to treat hematological malignancies and solid tumors is a growing concern, largely due to its broad toxicities. Its use has been associated with significant clinical side effects-cardiovascular, neurological, gastrointestinal, and allergic-in patients receiving infusions of cell-therapy products. DMSO has also been associated with altered expression of natural killer (NK) and T-cell markers and their in vivo function, not to mention difficulties in scaling up DMSO-based cryoprotectants, which introduce manufacturing challenges for autologous and allogeneic cellular therapies, including chimeric antigen receptor (CAR)-T and CAR-NK cell therapies. Interest in developing alternatives to DMSO has resulted in the evaluation of a variety of sugars, proteins, polymers, amino acids, and other small molecules and osmolytes as well as modalities to efficiently enable cellular uptake of these cryoprotectants. However, the DMSO-free cryopreservation of NK and T cells remains difficult. They represent heterogeneous cell populations that are sensitive to freezing and thawing. As a result, clinical use of cryopreserved cell-therapy products has not moved past the use of DMSO. Here, we present the state of the art in the development and use of cryopreservation options that do not contain DMSO toward clinical solutions to enable the global deployment of safer adoptively transferred cell-based therapies.