Expansion and Activation of Natural Killer Cells for Cancer Immunotherapy

Duck Cho; Dario Campana

doi:10.3343/kjlm.2009.29.2.89

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Cho and Campana: Expansion and Activation of Natural Killer Cells for Cancer Immunotherapy

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Korean J Leg Med 2009;29(2):89-96.

Published online: 14 February 2009

DOI: https://doi.org/10.3343/kjlm.2009.29.2.89

Expansion and Activation of Natural Killer Cells for Cancer Immunotherapy

Duck Cho, M.D.¹, Dario Campana, M.D.^1,^2,

¹Department of Oncology, St. Jude Children's Research Hospital, Memphis, TN, USA

²Department of Pathology, St. Jude Children's Research Hospital, Memphis, TN, USA

Corresponding author: Dario Campana, M.D., Ph.D. Department of Oncology, St. Jude Children's Research Hospital, 262 Danny Thomas Place, Memphis, TN 38105, USA Tel: +901-4595 2528, Fax: +901-4595 5947 E-mail: dario.campana@stjude.org

^∗This work was supported by grants CA113482 and CA21765 from the National Cancer Institute, grants from the Assisi Foundation and from the Fondation de Gouverneurs de l'Espoir, and by the American Lebanese Syrian Associated Charities (ALSAC).

Received 18 February 200925 March 2009 Accepted 27 March 2009

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Natural killer (NK) cells can kill a wide range of cancer cells and are a promising tool for cell therapy of cancer. NK cells cytotoxicity is regulated by a balance between stimulatory and inhibitory signals. Interleukin-2 is known to increase NK cell cytotoxicity. Although many cytokines have been studied in efforts to induce durable NK cell expansions, most reports indicate a rather modest effect and the requirement for additional stimuli. We found that contact with the K562 myeloid leukemia cell line, genetically modified to express a membrane-bound form of interleukin-15 and the ligand for the costimulatory molecule 4-1BB, induced vigorous expansion of NK cells from peripheral blood. Based on these findings, we developed a method for large-scale clinical-grade expansion of NK cells. This method is currently used to expand allogeneic NK cells for infusion in patients with leukemia and solid tumors. We here summarize methods for expansion and activation of NK cells from human peripheral blood mononuclear cells as well as clinical-scale methods to produce NK cells for immunotherapy under Current Good Manufacturing Practices (cGMP) conditions.

Keywords: NK cells, Cell therapy, Acute myeloid leukemia, Acute lymphoblastic leukemia, Chimeric receptors

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Fig. 1.

Schematic representation of protocols using expanded NK cells at St Jude Children's Research Hospital. The leukapheresis product obtained from a haploidentical donor is mixed with irradiated K562-mb15-41BBL cells. After 7 days of culture, most cells recovered are activated NK cells. After T-cell depletion using the CliniMACS system, NK cells are infused in patients with NK-sensitive malignancies such as acute myeloid leukemia (AML), Ewing sarcoma or rhabdomyosarcoma. For patients whose neoplasia is less sensitive to NK cytotoxicity, such as B-lineage acute lymphoblastic leukemia (ALL) or B-cell non-Hodgkin lymphoma (B-NHL), expanded NK cells are transduced with an anti-CD19 chi-meric receptor before infusion.

Table 1.

Selected protocols for expansion and activation of NK cells

Protocol	Median fold expansion after culture (% of CD56+CD3- cells)			Reference
Protocol	Day 7	Day 14	Day 21	Reference
PBMC from donor, OKT3 (10 ng/mL), IL-2 (500 IU/mL)	-	-	193 (55)	[21]
PBMC from myeloma, OKT3 (10 ng/mL), IL-2 (500 IU/mL)	-	-	1,625^∗ (65)	[20]
Non-adherent PBMC from ALL, RPMI8866, IL-2+IL-15	-	40^† (62-95)	-	[49]
PBMC from donor, HFWT, IL-2 (200 IU/mL)	-	58-401^‡ (77.4-85.6)	-	[28]
NK cell enriched from donor, IL-2 (100 IU/mL)+IL-15 (10 IU/mL)+ PHA (100 μg/mL)+ionomycin (1 μM/mL)	-	80-200	-	[29]
PBMC from donor, K562-mIL15-41BBL, IL-2 (10-100 IU/mL)	21.6 (62.9)	152 (90)	277 (96.8)	[43]

^∗ Measured on day 20;

^† day 10;

^‡ day 10-21

Abbreviations: PBMC, peripheral blood mononuclear cells; ALL, acute lymphoblastic leukemia.

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