Abstract
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.
Highlights
Natural killer (NK) cells are important effector cells in the immune response to cancer
We confirm that NK cells retain their ability to induce target cell death in a degranulation assay, but we find a decrease of cytotoxic function in a chromium-release assay after cryopreservation, following a good manufacturing practice conform protocol, which was developed and approved for the cryopreservation of T cells intended for clinical use[17,18]
We find that the fraction of motile NK cells in 3-D collagen gels is decreased by sixfold after cryopreservation, while the small remaining population of motile cells retains its cytotoxic function in a 3-D environment
Summary
Natural killer (NK) cells are important effector cells in the immune response to cancer. Standard degranulation and chromium release cytotoxicity assays confirm the ability of cryopreserved NK cells to kill target cells. We report that tumor cells embedded in a 3-dimensional collagen gel, 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. We confirm that NK cells retain their ability to induce target cell death in a degranulation assay, but we find a decrease of cytotoxic function in a chromium-release assay after cryopreservation, following a good manufacturing practice conform protocol, which was developed and approved for the cryopreservation of T cells intended for clinical use[17,18]. We find that the fraction of motile NK cells in 3-D collagen gels is decreased by sixfold after cryopreservation, while the small remaining population of motile cells retains its cytotoxic function in a 3-D environment
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