Nanoparticle‐Mediated Intracellular Protection of Natural Killer Cells Avoids Cryoinjury and Retains Potent Antitumor Functions

Abstract

The ability of natural killer (NK) cells to mediate potent antitumor immunity in clinical adoptive transfer settings relies, in large part, on their ability to retain cytotoxic function following cryopreservation. To avoid potential systemic toxicities associated with infusions of NK cells into patients in the presence of dimethylsulfoxide (DMSO), interest in alternative cryoprotective agents (CPAs) with improved safety profiles has grown. Despite the development of various sugars, amino acids, polyols, and polyampholytes as cryoprotectants, their ability to promote protection from intracellular cryodamage is limited because they mostly act outside of the cell. Though ways to shuttle cryoprotectants intracellularly exist, NK cells' high aversity to manipulation and freezing has meant they are highly understudied as targets for the development of new cryopreservation approaches. Here, the first example of a safe and efficient platform for the intracellular delivery of non‐DMSO CPAs to NK cells is presented. Biocompatible chitosan‐based nanoparticles are engineered to mediate the efficient DMSO‐free cryopreservation of NK cells. NK cells cryopreserved in this way retain potent cytotoxic, degranulation, and cytokine production functions against tumor targets. This not only represents the first example of delivering nanoparticles to NK cells, but illustrates the clinical potential in manufacturing safer allogeneic adoptive immunotherapies “off the shelf.”

Ice Inhibition for Cryopreservation: Materials, Strategies, and Challenges

“…More recently, in order to avoid intracellular ice damage, Matosevic and co‐workers achieved high‐efficiency DMSO‐free cryopreservation of natural killer (NK) cells by using nanoparticle‐mediated intracellular trehalose delivery. [ 159 ] As schematically illustrated in Figure 11D, chitosan–tripolyphosphate (CS–TPP) nanoparticles were assembled by ionic gelation. In the acidic environment of the cell interior, the CS–TPP nanoparticles can successfully release trehalose, owing to the pH‐responsive property.…”

Chimeric antigen receptor natural killer (CAR-NK) cell design and engineering for cancer therapy

“…Numerous groups are trying to optimize the ingredients of the freezing medium and procedures for expanded CAR-NK cells and have achieved encouraging outcomes [ 162 , 190 , 191 ]. A recent approach uses nanoparticle-mediated intracellular protection of NK cells which can avoid cryoinjury and maintain the killing potential of NK-92 cells, which could completely replace DMSO as a freezing protector [ 192 ]. This technique still has to be tried on and optimized for primary NK cells.…”

New Orders to an Old Soldier: Optimizing NK Cells for Adoptive Immunotherapy in Hematology

“…A DMSO-free freezing medium composed of poly-lysine, dextran and ectoine improves post-thaw cytotoxicity [ 106 ]. Nanoparticle-mediated intracellular protection was shown to avoid cryoinjury [ 107 ].…”