Bioinspired Preservation of Natural Killer Cells for Cancer Immunotherapy

Assal, Rami El; Abou‐Elkacem, Lotfi; Tocchio, Alessandro; Pasley, Shannon; Matosevic, Sandro; Kaplan, David L.; Zylberberg, Claudia; Demirci, Utkan

doi:10.1002/advs.201802045

Cited by 36 publications

(35 citation statements)

References 74 publications

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“…NK cells are generally cryopreserved using 10-20% dimethyl sulfoxide (DMSO) solution as a cryoprotectant at a slow freezing rate of −1°C/min (ref. 22 ). Several groups have reported changes in cell viability and/or function in non-expanded (primary) NK cells after cryopreservation.…”

Section: Discussionmentioning

confidence: 99%

Cryopreservation impairs 3-D migration and cytotoxicity of natural killer cells

et al. 2020

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Natural killer (NK) cells are important effector cells in the immune response to cancer. Clinical trials on adoptively transferred NK cells in patients with solid tumors, however, have thus far been unsuccessful. As NK cells need to pass stringent safety evaluation tests before clinical use, the cells are cryopreserved to bridge the necessary evaluation time. Standard degranulation and chromium release cytotoxicity assays confirm the ability of cryopreserved NK cells to kill target cells. Here, we report that tumor cells embedded in a 3-dimensional collagen gel, however, are killed by cryopreserved NK cells at a 5.6-fold lower rate compared to fresh NK cells. This difference is mainly caused by a 6-fold decrease in the fraction of motile NK cells after cryopreservation. These findings may explain the persistent failure of NK cell therapy in patients with solid tumors and highlight the crucial role of a 3-D environment for testing NK cell function.

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Section: Discussionmentioning

confidence: 99%

Cryopreservation impairs 3-D migration and cytotoxicity of natural killer cells

et al. 2020

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“…However, NK cells represent large granular lymphocytes which are more sensitive to cryopreservation than T cells, usually displaying impaired viability and long lag phases after thawing (181). While efforts are made to improve cryopreservation of NK cells, and some NK-92 cell products under commercial development are provided in cryopreserved form (154, 182, 183), we decided to use fresh NK-92/5.28.z cells in exponential growth phase for application in glioblastoma patients.…”

Section: Manufacturing Of Car-nk-92 Cells For Clinical Applicationmentioning

confidence: 99%

CAR-Engineered NK Cells for the Treatment of Glioblastoma: Turning Innate Effectors Into Precision Tools for Cancer Immunotherapy

et al. 2019

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Glioblastoma (GB) is the most common and aggressive primary brain tumor in adults and currently incurable. Despite multimodal treatment regimens, median survival in unselected patient cohorts is <1 year, and recurrence remains almost inevitable. Escape from immune surveillance is thought to contribute to the development and progression of GB. While GB tumors are frequently infiltrated by natural killer (NK) cells, these are actively suppressed by the GB cells and the GB tumor microenvironment. Nevertheless, ex vivo activation with cytokines can restore cytolytic activity of NK cells against GB, indicating that NK cells have potential for adoptive immunotherapy of GB if potent cytotoxicity can be maintained in vivo. NK cells contribute to cancer immune surveillance not only by their direct natural cytotoxicity which is triggered rapidly upon stimulation through germline-encoded cell surface receptors, but also by modulating T-cell mediated antitumor immune responses through maintaining the quality of dendritic cells and enhancing the presentation of tumor antigens. Furthermore, similar to T cells, specific recognition and elimination of cancer cells by NK cells can be markedly enhanced through expression of chimeric antigen receptors (CARs), which provides an opportunity to generate NK-cell therapeutics of defined specificity for cancer immunotherapy. Here, we discuss effects of the GB tumor microenvironment on NK-cell functionality, summarize early treatment attempts with ex vivo activated NK cells, and describe relevant CAR target antigens validated with CAR-T cells. We then outline preclinical approaches that employ CAR-NK cells for GB immunotherapy, and give an overview on the ongoing clinical development of ErbB2 (HER2)-specific CAR-NK cells currently applied in a phase I clinical trial in glioblastoma patients.

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“…We recently reported the cryopreservation of NK cells with combinations of non‐DMSO CPAs. [ 50,51 ] While our results show that NK cells cryopreserved under these conditions retain cytotoxic functions, none of these CPAs are able to penetrate the cell membrane, limiting potential intracellular damage that NK cells experience during the freezing and thawing process, and impairing durable functional responses.…”

Section: Introductionmentioning

confidence: 81%

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

et al. 2020

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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.”

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Bioinspired Preservation of Natural Killer Cells for Cancer Immunotherapy

Cited by 36 publications

References 74 publications

Cryopreservation impairs 3-D migration and cytotoxicity of natural killer cells

Cryopreservation impairs 3-D migration and cytotoxicity of natural killer cells

CAR-Engineered NK Cells for the Treatment of Glioblastoma: Turning Innate Effectors Into Precision Tools for Cancer Immunotherapy

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

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