Optimization of Large-Scale Expansion and Cryopreservation of Human Natural Killer Cells for Anti-Tumor Therapy

Min, Bumki; Choi, Heechul; Her, Jung Hyun; Kim, Hyojin; Jung, M.; Lee, Eun‐Kyoung; Cho, Sung Yoo; Hwang, Yu Kyeong; Shin, Eui‐Cheol

doi:10.4110/in.2018.18.e31

Cited by 43 publications

(36 citation statements)

References 29 publications

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“…Fresh medium with 500 IU/mL IL-2 was fed into NK cell cultures every two days to maintain a concentration at 0.5–2 × 10 6 cells/mL for 3-week culture. NK cells were thawed in water bath (37 °C) and used immediately for experiments without recovery period [90]. Cell count and viability were assessed by propidium iodide (PI) staining using an automatic cell counter (Digital Bio, Seoul, Republic of Korea).…”

Section: Methodsmentioning

confidence: 99%

Cryopreserved Human Natural Killer Cells Exhibit Potent Antitumor Efficacy against Orthotopic Pancreatic Cancer through Efficient Tumor-Homing and Cytolytic Ability

Min²,

et al. 2019

Cancers

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Pancreatic cancer is known to be highly aggressive, and desmoplasia-induced accumulation of extracellular matrix (ECM), which is a hallmark of many pancreatic cancers, severely restricts the therapeutic efficacy of both immunotherapeutics and conventional chemotherapeutics due to the ECM functioning as a major physical barrier against permeation and penetration. In the case of cell-based immunotherapeutics, there are several other bottlenecks preventing translation into clinical use due to their biological nature; for example, poor availability of cell therapeutic in a readily usable form due to difficulties in production, handling, shipping, and storage. To address these challenges, we have isolated allogeneic natural killer (NK) cells from healthy donors and expanded them in vitro to generate cryopreserved stocks. These cryopreserved NK cells were thawed to evaluate their therapeutic efficacy against desmoplastic pancreatic tumors, ultimately aiming to develop a readily accessible and mass-producible off-the-shelf cell-based immunotherapeutic. The cultured NK cells post-thawing retained highly pure populations of activated NK cells that expressed various activating receptors and a chemokine receptor. Furthermore, systemic administration of NK cells induced greater in vivo tumor growth suppression when compared with gemcitabine, which is the standard chemotherapeutic used for pancreatic cancer treatment. The potent antitumor effect of NK cells was mediated by efficient tumor-homing ability and infiltration into desmoplastic tumor tissues. Moreover, the infiltration of NK cells led to strong induction of apoptosis, elevated expression of the antitumor cytokine interferon (IFN)-γ, and inhibited expression of the immunosuppressive transforming growth factor (TGF)-β in tumor tissues. Expanded and cryopreserved NK cells are strong candidates for future cell-mediated systemic immunotherapy against pancreatic cancer.

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Section: Methodsmentioning

confidence: 99%

Cryopreserved Human Natural Killer Cells Exhibit Potent Antitumor Efficacy against Orthotopic Pancreatic Cancer through Efficient Tumor-Homing and Cytolytic Ability

Min²,

et al. 2019

Cancers

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“…Peripheral blood mononuclear cells were collected from healthy donors (n=3; 2 males and 1 female; average age, 36.6) via lymphapheresis for 2-4 batch productions in February 2019 from Seoul National University Hospital (Seoul, Korea). NK cells present in the mononuclear cells were enriched and expanded with Cellgro SCGM medium (CellGenix) containing human plasma and interleukin-2 for approximately three weeks as previously described (21,22). When the NK cell cultivation was completed, the NK-CdM was harvested by centrifugation at 400 × g for 3 min at 4˚C to remove the NK cells.…”

Section: Methodsmentioning

confidence: 99%

Anti‑photoaging and anti‑oxidative activities of natural killer cell conditioned medium following UV‑B irradiation of human dermal fibroblasts and a reconstructed skin model

et al. 2019

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Conditioned media from various sources comprise numerous growth factors and cytokines and are known to promote the regeneration of damaged tissues. Among these, natural killer cell conditioned medium (NK-CdM) has been shown to stimulate collagen synthesis and the migration of fibroblasts during the wound healing process. With a long-term aim of developing a treatment for skin photoaging, the ability of NK-CdM to prevent ultraviolet-B (UV-B) damage was assessed in neonatal human dermal fibroblasts (NHDFs) and an in vitro reconstructed skin model. The factors present in NK-CdM were profiled using an antibody array analysis. Protein and mRNA levels in UV-B exposed NHDFs treated with NK-CdM were measured by western blotting and quantitative reverse transcription-PCR, respectively. The total antioxidant capacity of NK-CdM was determined to assess its ability to suppress reactive oxygen species. The anti-photoaging effect of NK-CdM was also assessed in a 3D reconstituted human full skin model. NK-CdM induced proliferation of UV-B-treated NHDFs, increased procollagen expression, and decreased matrix metalloproteinase (MMP)-1 expression. NK-CdM also exhibited a potent antioxidant activity as measured by the total antioxidant capacity. NK-CdM inhibited UV-B-induced collagen degradation by inactivating MAPK signaling. NK-CdM also elicited potential anti-wrinkle effects by inhibiting the UV-B-induced increase in MMP-1 expression levels in a 3D reconstituted human full skin model. Taken together, the suppression of both UV-B-induced MMP-1 expression and JNK activation by NK-CdM suggests NK-CdM as a possible candidate anti-skin aging agent.

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“…NK cell-based cures have been promising and groundbreaking and have been clinically tested in customized chemotherapy. In cell-mediated immunotherapy, an important goal is obtaining a large number of NK cells by efficiently proliferating NK cells ( 39 40 ). Although oral administration of high doses of silk peptide inhibited NK cell frequency, oral administration of intermediate doses (750 and 1,500 mg/kg body weight/day) significantly increased the frequency of NK cells ( Fig.…”

Section: Discussionmentioning

confidence: 99%

Oral Administration of Silk Peptide Enhances the Maturation and Cytolytic Activity of Natural Killer Cells

Jang

Oh²,

Baek³

et al. 2018

Immune Netw

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Silk peptide, the hydrolysate of silk protein derived from cocoons, has been employed as a biomedical material and is believed to be safe for human use. Silk peptide display various bioactivities, including anti-inflammatory, immune-regulatory, anti-tumor, anti-viral, and anti-bacterial. Although earlier investigations demonstrated that silk peptide stimulates macrophages and the production of pro-inflammatory cytokines, its effect on natural killer (NK) cell function has not yet been explored. In this study, we initially confirmed that silk peptide enhances NK cell activity in vitro and ex vivo. To assess the modulatory activity of silk peptide on NK cells, mice were fed various amounts of a silk peptide-supplemented diet for 2 months and the effects on immune stimulation, including NK cell activation, were evaluated. Oral administration of silk peptide significantly enhanced the proliferation of mitogen- or IL-2-stimulated splenocytes. In addition, oral silk peptide treatment enhanced the frequency and degree of maturation of NK cells in splenocytes. The same treatment also significantly enhanced the target cell cytolytic activity of NK cells, which was determined by cell surface CD107a expression and intracellular interferon-γ expression. Finally, oral administration of silk peptide stimulated T helper 1-type cytokine expression from splenic lymphocytes. Collectively, our results suggest that silk peptide potentiates NK cell activity in vivo and could be used as a compound for immune-modulating anti-tumor treatment.

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Optimization of Large-Scale Expansion and Cryopreservation of Human Natural Killer Cells for Anti-Tumor Therapy

Cited by 43 publications

References 29 publications

Cryopreserved Human Natural Killer Cells Exhibit Potent Antitumor Efficacy against Orthotopic Pancreatic Cancer through Efficient Tumor-Homing and Cytolytic Ability

Cryopreserved Human Natural Killer Cells Exhibit Potent Antitumor Efficacy against Orthotopic Pancreatic Cancer through Efficient Tumor-Homing and Cytolytic Ability

Anti‑photoaging and anti‑oxidative activities of natural killer cell conditioned medium following UV‑B irradiation of human dermal fibroblasts and a reconstructed skin model

Oral Administration of Silk Peptide Enhances the Maturation and Cytolytic Activity of Natural Killer Cells

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