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

Tittlbach H, Schneider A, Strobel J, Zimmermann R, Maas S, Gebhardt B, Rauser G, Mach M, Mackensen A, Winkler T, Winkler J (2017)

Publication Type: Journal article

Publication year: 2017

Journal

Book Volume: 15

Pages Range: 228

Journal Issue: 1

DOI: 10.1186/s12967-017-1330-5

Abstract

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.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.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 × 108 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.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.

Authors with CRIS profile

Involved external institutions

Miltenyi Biotec GmbH Germany (DE)

How to cite

APA:

Tittlbach, H., Schneider, A., Strobel, J., Zimmermann, R., Maas, S., Gebhardt, B.,... Winkler, J. (2017). GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation. Journal of Translational Medicine, 15(1), 228. https://dx.doi.org/10.1186/s12967-017-1330-5

MLA:

Tittlbach, Hannes, et al. "GMP-production of purified human B lymphocytes for the adoptive transfer in patients after allogeneic hematopoietic stem cell transplantation." Journal of Translational Medicine 15.1 (2017): 228.

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