Ex Vivo Generation of Donor Antigen-Specific Immunomodulatory Cells: A Comparison Study of Anti-CD80/86 mAbs and CTLA4-lg Costimulatory Blockade

M Watanabe; Makiko Kumagai-Braesch; M Yao; S Thunberg; D Berglund; F Sellberg; C Jorns; S Lind Enoksson; J Henriksson; T Lundgren; M Uhlin; E Berglund; B-G Ericzon

doi:10.1177/0963689718794642

Ex Vivo Generation of Donor Antigen-Specific Immunomodulatory Cells: A Comparison Study of Anti-CD80/86 mAbs and CTLA4-lg Costimulatory Blockade

Cell Transplant. 2018 Nov;27(11):1692-1704. doi: 10.1177/0963689718794642. Epub 2018 Sep 27.

Authors

M Watanabe^{1

2}, Makiko Kumagai-Braesch^{1

2}, M Yao^{1

2}, S Thunberg³, D Berglund⁴, F Sellberg⁴, C Jorns^{1

2}, S Lind Enoksson³, J Henriksson^{1

2}, T Lundgren^{1

2}, M Uhlin^{1

3}, E Berglund^{1

2}, B-G Ericzon^{1

2}

Affiliations

¹ Division of Transplantation Surgery, CLINTEC, Karolinska Institutet, Stockholm, Sweden.
² Department of Transplantation Surgery, Karolinska University Hospital, Huddinge, Sweden.
³ Department of Clinical Immunology, Karolinska University Hospital, Stockholm, Sweden.
⁴ Department of Immunology, Genetics and Pathology, Section of Clinical Immunology, Uppsala University, Uppsala, Sweden.

Abstract

Adoptive transfer of alloantigen-specific immunomodulatory cells generated ex vivo with anti-CD80/CD86 mAbs (2D10.4/IT2.2) holds promise for operational tolerance after transplantation. However, good manufacturing practice is required to allow widespread clinical application. Belatacept, a clinically approved cytotoxic T-lymphocyte antigen 4-immunoglobulin that also binds CD80/CD86, could be an alternative agent for 2D10.4/IT2.2. With the goal of generating an optimal cell treatment with clinically approved reagents, we evaluated the donor-specific immunomodulatory effects of belatacept- and 2D10.4/IT2.2-generated immunomodulatory cells. Immunomodulatory cells were generated by coculturing responder human peripheral blood mononuclear cells (PBMCs) (50 × 10⁶ cells) with irradiated donor PBMCs (20 × 10⁶ cells) from eight human leukocyte antigen-mismatched responder-donor pairs in the presence of either 2D10.4/IT2.2 (3 μg/10⁶ cells) or belatacept (40 μg/10⁶ cells). After 14 days of coculture, the frequencies of CD4⁺ T cells, CD8⁺ T cells, and natural killer cells as well as interferon gamma (IFN-γ) production in the 2D10.4/IT2.2- and belatacept-treated groups were lower than those in the control group. The percentage of CD19⁺ B cells was higher in the 2D10.4/IT2.2- and belatacept-treated groups than in the control group. The frequency of CD4⁺CD25⁺CD127^lowFOXP3⁺ T cells increased from 4.1±1.0% (preculture) to 7.1±2.6% and 7.3±2.6% (day 14) in the 2D10.4/IT2.2- and belatacept-treated groups, respectively (p<0.05). Concurrently, delta-2 FOXP3 mRNA expression increased significantly. Compared with cells derived from the no-antibody treated control group, cells generated from both the 2D10.4/IT2.2- and belatacept-treated groups produced lower IFN-γ and higher interleukin-10 levels in response to donor-antigens, as detected by enzyme-linked immunospot. Most importantly, 2D10.4/IT2.2- and belatacept-generated cells effectively impeded the proliferative responses of freshly isolated responder PBMCs against donor-antigens. Our results indicate that belatacept-generated donor-specific immunomodulatory cells possess comparable phenotypes and immunomodulatory efficacies to those generated with 2D10.4/IT2.2. We suggest that belatacept could be used for ex vivo generation of clinical grade alloantigen-specific immunomodulatory cells for tolerance induction after transplantation.

Keywords: CTLA4-Ig; FOXP3; Immunomodulatory cells; anti-CD80/86 antibodies; donor antigen specificity; ex vivo generation.