Human Natural Killer Cells Cryopreserved without DMSO Sustain Robust Effector Responses

Area of Science:

• Immunology
• Cell Biology
• Cryobiology

Background:

• Natural killer (NK) cell immunotherapy offers safety and allogeneic potential for adoptive transfer.
• Cryopreservation of NK cell products is essential for decentralized manufacturing and logistical deployment.
• Dimethyl sulfoxide (DMSO), a common cryoprotective agent (CPA), has toxicity and can induce molecular/genetic changes in immune cells.

Purpose of the Study:

• To investigate alternative cryoprotective agents (CPAs) to replace DMSO for human NK cell cryopreservation.
• To assess the feasibility of cryopreserving NK cells using non-cell-penetrating and cell-penetrating CPAs without DMSO.
• To evaluate the impact of non-DMSO cryopreservation on NK cell viability, cytotoxicity, and activation markers.

Main Methods:

• Human peripheral blood-derived NK cells were cryopreserved using DMSO-free formulations with cell-penetrating and non-cell-penetrating CPAs.
• Post-thaw NK cell viability was assessed.
• NK cell cytotoxicity assays were performed.
• Surface markers of NK cell activation were analyzed.

Main Results:

• Cryoprotection using cell-penetrating CPAs maintained human NK cell viability comparable to DMSO.
• NK cells cryopreserved without DMSO exhibited comparable cytotoxicity to DMSO-preserved cells.
• Non-DMSO-cryopreserved NK cells displayed similar surface markers of activation as DMSO-preserved cells.

Conclusions:

• Cell-penetrating CPAs show potential as effective alternatives to DMSO for human NK cell cryopreservation.
• This study presents the first evidence for developing non-DMSO CPA formulations for future cell therapy regimens.
• Non-DMSO cryopreservation strategies could enhance the logistical feasibility and safety of NK cell-based therapies.