Preparation of cytokine-activated NK cells for use in adoptive cell therapy in cancer patients: Protocol optimization and therapeutic potential

Monique M. Van Ostaijen-Ten Dam, Henk Jan Prins, Gerharda H. Boerman, Carly Vervat, Daniela Pende, Hein Putter, Arjan Lankester, Maarten J.D. Van Tol, Jaap J. Zwaginga, Marco W. Schilham

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Cell-based immunotherapy using donor-derived natural killer (NK) cells after allogeneic hematopoietic stem cell transplantation may be an attractive treatment of residual leukemia. This study aimed to optimize clinical grade production of a cytokine- activated NK-cell product. NK cells were isolated either by double depletion (CD3-, CD19-) or by sequential depletion and enrichment (CD3-, CD56+) via CliniMACS from leukapheresis material and cultured in vitro with interleukin (IL)-2 or IL-15. Both NK cell isolation procedures yielded comparable recovery of NK cells and levels of T-cell contamination. After culture with cytokines, the CD3-CD56+ procedure resulted in NK cells of higher purity, that is, less T cells and monocytes, higher viability, and a slightly higher yield than the CD3-CD19- procedure. CD69, NKp44, and NKG2A expression were higher on CD3-CD56+ products, whereas lysis of Daudi cells was comparable. Five days of culture led to higher expression of CD69, NKp44, and NKp30 and lysis of K562 and Daudi cell lines. Although CD69 expression and lysis of Daudi cells were slightly higher in cultures with IL-2, T-cell contamination was lower with IL-15. Therefore, further experiments were performed with CD3-CD56+ products cultured with IL-15. Cryopreservation of IL-15-activated NK cells resulted in a loss of cytotoxicity (>92%), whereas thawing of isolated, uncultured NK cells followed by culture with IL-15 yielded cells with about 43% of the original lytic activity. Five-day IL-15-activated NK cells lysed tumor target cell lines and primary leukemic blasts, providing the basis for NK cell-based immunotherapeutic strategies in a clinical setting.

Original languageEnglish
Pages (from-to)90-100
Number of pages11
JournalJournal of Immunotherapy
Volume39
Issue number2
DOIs
Publication statusPublished - 1 Jan 2016

Cite this

Van Ostaijen-Ten Dam, Monique M. ; Prins, Henk Jan ; Boerman, Gerharda H. ; Vervat, Carly ; Pende, Daniela ; Putter, Hein ; Lankester, Arjan ; Van Tol, Maarten J.D. ; Zwaginga, Jaap J. ; Schilham, Marco W. / Preparation of cytokine-activated NK cells for use in adoptive cell therapy in cancer patients : Protocol optimization and therapeutic potential. In: Journal of Immunotherapy. 2016 ; Vol. 39, No. 2. pp. 90-100.
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abstract = "Cell-based immunotherapy using donor-derived natural killer (NK) cells after allogeneic hematopoietic stem cell transplantation may be an attractive treatment of residual leukemia. This study aimed to optimize clinical grade production of a cytokine- activated NK-cell product. NK cells were isolated either by double depletion (CD3-, CD19-) or by sequential depletion and enrichment (CD3-, CD56+) via CliniMACS from leukapheresis material and cultured in vitro with interleukin (IL)-2 or IL-15. Both NK cell isolation procedures yielded comparable recovery of NK cells and levels of T-cell contamination. After culture with cytokines, the CD3-CD56+ procedure resulted in NK cells of higher purity, that is, less T cells and monocytes, higher viability, and a slightly higher yield than the CD3-CD19- procedure. CD69, NKp44, and NKG2A expression were higher on CD3-CD56+ products, whereas lysis of Daudi cells was comparable. Five days of culture led to higher expression of CD69, NKp44, and NKp30 and lysis of K562 and Daudi cell lines. Although CD69 expression and lysis of Daudi cells were slightly higher in cultures with IL-2, T-cell contamination was lower with IL-15. Therefore, further experiments were performed with CD3-CD56+ products cultured with IL-15. Cryopreservation of IL-15-activated NK cells resulted in a loss of cytotoxicity (>92{\%}), whereas thawing of isolated, uncultured NK cells followed by culture with IL-15 yielded cells with about 43{\%} of the original lytic activity. Five-day IL-15-activated NK cells lysed tumor target cell lines and primary leukemic blasts, providing the basis for NK cell-based immunotherapeutic strategies in a clinical setting.",
keywords = "Acute leukemia, Clini-MACS, Clinical protocol, Good manufacturing practice, Interleukin-15, Interleukin-2, Natural killer cells",
author = "{Van Ostaijen-Ten Dam}, {Monique M.} and Prins, {Henk Jan} and Boerman, {Gerharda H.} and Carly Vervat and Daniela Pende and Hein Putter and Arjan Lankester and {Van Tol}, {Maarten J.D.} and Zwaginga, {Jaap J.} and Schilham, {Marco W.}",
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Van Ostaijen-Ten Dam, MM, Prins, HJ, Boerman, GH, Vervat, C, Pende, D, Putter, H, Lankester, A, Van Tol, MJD, Zwaginga, JJ & Schilham, MW 2016, 'Preparation of cytokine-activated NK cells for use in adoptive cell therapy in cancer patients: Protocol optimization and therapeutic potential' Journal of Immunotherapy, vol. 39, no. 2, pp. 90-100. https://doi.org/10.1097/CJI.0000000000000110

Preparation of cytokine-activated NK cells for use in adoptive cell therapy in cancer patients : Protocol optimization and therapeutic potential. / Van Ostaijen-Ten Dam, Monique M.; Prins, Henk Jan; Boerman, Gerharda H.; Vervat, Carly; Pende, Daniela; Putter, Hein; Lankester, Arjan; Van Tol, Maarten J.D.; Zwaginga, Jaap J.; Schilham, Marco W.

In: Journal of Immunotherapy, Vol. 39, No. 2, 01.01.2016, p. 90-100.

Research output: Contribution to journalArticleAcademicpeer-review

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T1 - Preparation of cytokine-activated NK cells for use in adoptive cell therapy in cancer patients

T2 - Protocol optimization and therapeutic potential

AU - Van Ostaijen-Ten Dam, Monique M.

AU - Prins, Henk Jan

AU - Boerman, Gerharda H.

AU - Vervat, Carly

AU - Pende, Daniela

AU - Putter, Hein

AU - Lankester, Arjan

AU - Van Tol, Maarten J.D.

AU - Zwaginga, Jaap J.

AU - Schilham, Marco W.

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N2 - Cell-based immunotherapy using donor-derived natural killer (NK) cells after allogeneic hematopoietic stem cell transplantation may be an attractive treatment of residual leukemia. This study aimed to optimize clinical grade production of a cytokine- activated NK-cell product. NK cells were isolated either by double depletion (CD3-, CD19-) or by sequential depletion and enrichment (CD3-, CD56+) via CliniMACS from leukapheresis material and cultured in vitro with interleukin (IL)-2 or IL-15. Both NK cell isolation procedures yielded comparable recovery of NK cells and levels of T-cell contamination. After culture with cytokines, the CD3-CD56+ procedure resulted in NK cells of higher purity, that is, less T cells and monocytes, higher viability, and a slightly higher yield than the CD3-CD19- procedure. CD69, NKp44, and NKG2A expression were higher on CD3-CD56+ products, whereas lysis of Daudi cells was comparable. Five days of culture led to higher expression of CD69, NKp44, and NKp30 and lysis of K562 and Daudi cell lines. Although CD69 expression and lysis of Daudi cells were slightly higher in cultures with IL-2, T-cell contamination was lower with IL-15. Therefore, further experiments were performed with CD3-CD56+ products cultured with IL-15. Cryopreservation of IL-15-activated NK cells resulted in a loss of cytotoxicity (>92%), whereas thawing of isolated, uncultured NK cells followed by culture with IL-15 yielded cells with about 43% of the original lytic activity. Five-day IL-15-activated NK cells lysed tumor target cell lines and primary leukemic blasts, providing the basis for NK cell-based immunotherapeutic strategies in a clinical setting.

AB - Cell-based immunotherapy using donor-derived natural killer (NK) cells after allogeneic hematopoietic stem cell transplantation may be an attractive treatment of residual leukemia. This study aimed to optimize clinical grade production of a cytokine- activated NK-cell product. NK cells were isolated either by double depletion (CD3-, CD19-) or by sequential depletion and enrichment (CD3-, CD56+) via CliniMACS from leukapheresis material and cultured in vitro with interleukin (IL)-2 or IL-15. Both NK cell isolation procedures yielded comparable recovery of NK cells and levels of T-cell contamination. After culture with cytokines, the CD3-CD56+ procedure resulted in NK cells of higher purity, that is, less T cells and monocytes, higher viability, and a slightly higher yield than the CD3-CD19- procedure. CD69, NKp44, and NKG2A expression were higher on CD3-CD56+ products, whereas lysis of Daudi cells was comparable. Five days of culture led to higher expression of CD69, NKp44, and NKp30 and lysis of K562 and Daudi cell lines. Although CD69 expression and lysis of Daudi cells were slightly higher in cultures with IL-2, T-cell contamination was lower with IL-15. Therefore, further experiments were performed with CD3-CD56+ products cultured with IL-15. Cryopreservation of IL-15-activated NK cells resulted in a loss of cytotoxicity (>92%), whereas thawing of isolated, uncultured NK cells followed by culture with IL-15 yielded cells with about 43% of the original lytic activity. Five-day IL-15-activated NK cells lysed tumor target cell lines and primary leukemic blasts, providing the basis for NK cell-based immunotherapeutic strategies in a clinical setting.

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KW - Clini-MACS

KW - Clinical protocol

KW - Good manufacturing practice

KW - Interleukin-15

KW - Interleukin-2

KW - Natural killer cells

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