GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation

Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology. Ausführliche Beschreibung

Gespeichert in:

Autor*in:	Tittlbach, Hannes [verfasserIn] Schneider, Andrea Strobel, Julian Zimmermann, Robert Maas, Stefanie Gebhardt, Bernd Rauser, Georg Mach, Michael Mackensen, Andreas Winkler, Thomas H. Winkler, Julia

Format:	E-Artikel
Sprache:	Englisch

Erschienen:	2017

Schlagwörter:	Adoptive immunotherapy B cells GMP-compliant manufacturing

Anmerkung:	© The Author(s) 2017

Übergeordnetes Werk:	Enthalten in: Journal of translational medicine - London : BioMed Central, 2003, 15(2017), 1 vom: 07. Nov.
Übergeordnetes Werk:	volume:15 ; year:2017 ; number:1 ; day:07 ; month:11

Links:	Volltext

DOI / URN:	10.1186/s12967-017-1330-5

Katalog-ID:	SPR028965485

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520			\|a Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology.
650		4	\|a Adoptive immunotherapy \|7 (dpeaa)DE-He213
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700	1		\|a Rauser, Georg \|4 aut
700	1		\|a Mach, Michael \|4 aut
700	1		\|a Mackensen, Andreas \|4 aut
700	1		\|a Winkler, Thomas H. \|4 aut
700	1		\|a Winkler, Julia \|4 aut
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matchkey_str	article:14795876:2017----::mpoutooprfehmnlmhctsotedpiernfrnainsfealgnihm
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allfields	10.1186/s12967-017-1330-5 doi (DE-627)SPR028965485 (SPR)s12967-017-1330-5-e DE-627 ger DE-627 rakwb eng Tittlbach, Hannes verfasserin aut GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © The Author(s) 2017 Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology. Adoptive immunotherapy (dpeaa)DE-He213 B cells (dpeaa)DE-He213 GMP-compliant manufacturing (dpeaa)DE-He213 Schneider, Andrea aut Strobel, Julian aut Zimmermann, Robert aut Maas, Stefanie aut Gebhardt, Bernd aut Rauser, Georg aut Mach, Michael aut Mackensen, Andreas aut Winkler, Thomas H. aut Winkler, Julia aut Enthalten in Journal of translational medicine London : BioMed Central, 2003 15(2017), 1 vom: 07. Nov. (DE-627)369084136 (DE-600)2118570-0 1479-5876 nnns volume:15 year:2017 number:1 day:07 month:11 https://dx.doi.org/10.1186/s12967-017-1330-5 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 15 2017 1 07 11
spelling	10.1186/s12967-017-1330-5 doi (DE-627)SPR028965485 (SPR)s12967-017-1330-5-e DE-627 ger DE-627 rakwb eng Tittlbach, Hannes verfasserin aut GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © The Author(s) 2017 Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology. Adoptive immunotherapy (dpeaa)DE-He213 B cells (dpeaa)DE-He213 GMP-compliant manufacturing (dpeaa)DE-He213 Schneider, Andrea aut Strobel, Julian aut Zimmermann, Robert aut Maas, Stefanie aut Gebhardt, Bernd aut Rauser, Georg aut Mach, Michael aut Mackensen, Andreas aut Winkler, Thomas H. aut Winkler, Julia aut Enthalten in Journal of translational medicine London : BioMed Central, 2003 15(2017), 1 vom: 07. Nov. (DE-627)369084136 (DE-600)2118570-0 1479-5876 nnns volume:15 year:2017 number:1 day:07 month:11 https://dx.doi.org/10.1186/s12967-017-1330-5 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 15 2017 1 07 11
allfields_unstemmed	10.1186/s12967-017-1330-5 doi (DE-627)SPR028965485 (SPR)s12967-017-1330-5-e DE-627 ger DE-627 rakwb eng Tittlbach, Hannes verfasserin aut GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © The Author(s) 2017 Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology. Adoptive immunotherapy (dpeaa)DE-He213 B cells (dpeaa)DE-He213 GMP-compliant manufacturing (dpeaa)DE-He213 Schneider, Andrea aut Strobel, Julian aut Zimmermann, Robert aut Maas, Stefanie aut Gebhardt, Bernd aut Rauser, Georg aut Mach, Michael aut Mackensen, Andreas aut Winkler, Thomas H. aut Winkler, Julia aut Enthalten in Journal of translational medicine London : BioMed Central, 2003 15(2017), 1 vom: 07. Nov. (DE-627)369084136 (DE-600)2118570-0 1479-5876 nnns volume:15 year:2017 number:1 day:07 month:11 https://dx.doi.org/10.1186/s12967-017-1330-5 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 15 2017 1 07 11
allfieldsGer	10.1186/s12967-017-1330-5 doi (DE-627)SPR028965485 (SPR)s12967-017-1330-5-e DE-627 ger DE-627 rakwb eng Tittlbach, Hannes verfasserin aut GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © The Author(s) 2017 Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology. Adoptive immunotherapy (dpeaa)DE-He213 B cells (dpeaa)DE-He213 GMP-compliant manufacturing (dpeaa)DE-He213 Schneider, Andrea aut Strobel, Julian aut Zimmermann, Robert aut Maas, Stefanie aut Gebhardt, Bernd aut Rauser, Georg aut Mach, Michael aut Mackensen, Andreas aut Winkler, Thomas H. aut Winkler, Julia aut Enthalten in Journal of translational medicine London : BioMed Central, 2003 15(2017), 1 vom: 07. Nov. (DE-627)369084136 (DE-600)2118570-0 1479-5876 nnns volume:15 year:2017 number:1 day:07 month:11 https://dx.doi.org/10.1186/s12967-017-1330-5 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 15 2017 1 07 11
allfieldsSound	10.1186/s12967-017-1330-5 doi (DE-627)SPR028965485 (SPR)s12967-017-1330-5-e DE-627 ger DE-627 rakwb eng Tittlbach, Hannes verfasserin aut GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation 2017 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier © The Author(s) 2017 Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology. Adoptive immunotherapy (dpeaa)DE-He213 B cells (dpeaa)DE-He213 GMP-compliant manufacturing (dpeaa)DE-He213 Schneider, Andrea aut Strobel, Julian aut Zimmermann, Robert aut Maas, Stefanie aut Gebhardt, Bernd aut Rauser, Georg aut Mach, Michael aut Mackensen, Andreas aut Winkler, Thomas H. aut Winkler, Julia aut Enthalten in Journal of translational medicine London : BioMed Central, 2003 15(2017), 1 vom: 07. Nov. (DE-627)369084136 (DE-600)2118570-0 1479-5876 nnns volume:15 year:2017 number:1 day:07 month:11 https://dx.doi.org/10.1186/s12967-017-1330-5 kostenfrei Volltext GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 15 2017 1 07 11
language	English
source	Enthalten in Journal of translational medicine 15(2017), 1 vom: 07. Nov. volume:15 year:2017 number:1 day:07 month:11
sourceStr	Enthalten in Journal of translational medicine 15(2017), 1 vom: 07. Nov. volume:15 year:2017 number:1 day:07 month:11
format_phy_str_mv	Article
institution	findex.gbv.de
topic_facet	Adoptive immunotherapy B cells GMP-compliant manufacturing
isfreeaccess_bool	true
container_title	Journal of translational medicine
authorswithroles_txt_mv	Tittlbach, Hannes @@aut@@ Schneider, Andrea @@aut@@ Strobel, Julian @@aut@@ Zimmermann, Robert @@aut@@ Maas, Stefanie @@aut@@ Gebhardt, Bernd @@aut@@ Rauser, Georg @@aut@@ Mach, Michael @@aut@@ Mackensen, Andreas @@aut@@ Winkler, Thomas H. @@aut@@ Winkler, Julia @@aut@@
publishDateDaySort_date	2017-11-07T00:00:00Z
hierarchy_top_id	369084136
id	SPR028965485
language_de	englisch
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These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. 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author	Tittlbach, Hannes
spellingShingle	Tittlbach, Hannes misc Adoptive immunotherapy misc B cells misc GMP-compliant manufacturing GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation
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topic_title	GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation Adoptive immunotherapy (dpeaa)DE-He213 B cells (dpeaa)DE-He213 GMP-compliant manufacturing (dpeaa)DE-He213
topic	misc Adoptive immunotherapy misc B cells misc GMP-compliant manufacturing
topic_unstemmed	misc Adoptive immunotherapy misc B cells misc GMP-compliant manufacturing
topic_browse	misc Adoptive immunotherapy misc B cells misc GMP-compliant manufacturing
format_facet	Elektronische Aufsätze Aufsätze Elektronische Ressource
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hierarchy_parent_title	Journal of translational medicine
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title	GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation
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title_full	GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation
author_sort	Tittlbach, Hannes
journal	Journal of translational medicine
journalStr	Journal of translational medicine
lang_code	eng
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author_browse	Tittlbach, Hannes Schneider, Andrea Strobel, Julian Zimmermann, Robert Maas, Stefanie Gebhardt, Bernd Rauser, Georg Mach, Michael Mackensen, Andreas Winkler, Thomas H. Winkler, Julia
container_volume	15
format_se	Elektronische Aufsätze
author-letter	Tittlbach, Hannes
doi_str_mv	10.1186/s12967-017-1330-5
title_sort	gmp-production of purified human b lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation
title_auth	GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation
abstract	Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology. © The Author(s) 2017
abstractGer	Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology. © The Author(s) 2017
abstract_unstemmed	Background We have recently shown that memory B cells from murine CMV immune donor animals adoptively transferred into immunodeficient mice were highly effective in protecting from a viral infection indicating a therapeutic potential of virus specific memory B cells. These preclinical data provided evidence that a cell-based strategy supporting the humoral immune response might be effective in a clinical setting of immunodeficiency after allogeneic hematopoietic stem cell transplantation. As adoptive transfer of B cells has not been used before in a clinical setting it was necessary to establish a technology for the generation of good manufacturing practice (GMP)-grade B cell products. Methods Starting from the leukapheresis product of healthy blood donors, B cells were purified by two different separation strategies using GMP-grade microbeads and the CliniMACS system. A one-step protocol was used for positive enrichment of B lymphocytes with anti-CD19 microbeads. In a two-step enrichment protocol, first T lymphocytes were depleted by anti-CD3 microbeads and the remaining fraction was positively selected by anti-CD19 microbeads. Results The purity and recovery after enrichment of B lymphocytes from the leukapheresis material in both separations strategies was not statistically different. However, contamination of the B-cell product with T cells was significantly lower after the two-step protocol (0.16%, range 0.01–0.43% after two-step separation and 0.55%, range 0.28–0.85% after one-step separation, p < 0.05). Therefore, a combined CD3 depletion and CD19 enrichment was used for the production of GMP-conform B-cell products from the leukapheresis material of 17 healthy stem cell donors. The absolute B-cell numbers obtained in the final product was 4.70 ± 3.64 × $ 10^{8} $ with a purity of 95.98 ± 3.31% B lymphocytes and a recovery of 18.9 ± 10.6%. Importantly, the contamination with $ CD3^{+} $ T cells was extremely low in the final B- cell products (0.10 ± 0.20%). Purified B cells exhibited normal antibody production after in vitro stimulation and showed excellent viability after cryopreservation. Conclusions A GMP-grade B-cell product can be obtained with high purity and very low T-cell contamination using the two-step enrichment protocol based on CliniMACS® technology. © The Author(s) 2017
collection_details	GBV_USEFLAG_A SYSFLAG_A GBV_SPRINGER SSG-OLC-PHA GBV_ILN_11 GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_31 GBV_ILN_39 GBV_ILN_40 GBV_ILN_60 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_206 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2003 GBV_ILN_2005 GBV_ILN_2009 GBV_ILN_2011 GBV_ILN_2014 GBV_ILN_2055 GBV_ILN_2111 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700
container_issue	1
title_short	GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation
url	https://dx.doi.org/10.1186/s12967-017-1330-5
remote_bool	true
author2	Schneider, Andrea Strobel, Julian Zimmermann, Robert Maas, Stefanie Gebhardt, Bernd Rauser, Georg Mach, Michael Mackensen, Andreas Winkler, Thomas H. Winkler, Julia
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up_date	2024-07-03T22:47:32.847Z
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