2558: Optimization of intra-thymic transplantation of donor-derived thymic epithelial cells to promote lasting regulation of anti-donor reactivity

Abstract

2558: Optimization of intra-thymic transplantation of donor-derived thymic epithelial cells to promote lasting regulation of anti-donor reactivity Sara Alfadil, Kim Mi-Jeong, Marcos Iglesias Lozano, Byoung Chol Oh, W. P. Andrew Lee, Gerald Brandacher, Thomas Serworld, and Giorgio Raimondi Johns Hopkins University School of Medicine, Vascularized Composite Allotransplantation (VCA) Laboratory, Baltimore, MD; Joslin Diabetes Center, Boston, MA, USA Background Targeting the process of central (thymic) selection of developing T lymphocytes is the key mechanism for transplant tolerance induction mediated bybone marrow transplantation (BM-Tx)-based protocols. However, they are not amenable to most transplant recipients. Thymic Epithelial Cells (TEC), a population of stromal cells residing in the thymus, exert a major contribution to central selection. Unfortunately, donor TEC do not develop following BM-Tx protocols. Therefore, we propose a new strategy that voids the need for heavy pre-conditioning and is based on generating a donor-recipient “Hybrid Thymus,” through donor TEC engraftment, to re-engineer the thymic microenvironment and promote lasting central tolerance. Methods We developed an improved protocol for isolating mouse TEC via a combination of negative and positive selection. Allogeniec (Allo)-TEC purified from BALB/c were injected into the right thymic lobe of C57BL/6, Recipients were divided in groups based on treatment with CTLA4-Ig C/¡ anti-CD40L C/¡ T depletion. 14, 28, and 45 days post-injection Allo-TEC survival was assessed and the selection and reactivity of T cells was analyzed via flow cytometry and CFSEbased mixed lymphocyte reaction. Results Our optimized purification protocol yields an average 70% TEC purity. Unmanipulated recipients promptly rejected Allo-TEC, indicating the existence of effective immunity in the thymus. However, CTLA4-Ig and MR1 co-administration exerted a significant (but not lasting) protection. In this latter group, the percentage of peripheral Vb11C T cells (inversely correlated with central selection) on d24 was significantly lower, indicating functional activity of the engrafted donor TEC. Addition of transient T depletion enabled longterm engraftment of Allo-TEC that, on d45, were in a normal architecture within the thymus integrated with recipient thymocytes. However, antidonor T cell reactivity remained unaltered, suggesting the need for engraftment in both thymic lobes for proper regulation. Conclusion Our preliminary data show that the thymic engraftment, survival, and function of allo-TEC can be promoted via costimulation blockade C T depletion. These exciting results indicate that engineering a donor-recipient Hybrid Thymus is feasible and has the potential to promote a dominant regulation of alloreactivity that could be conducive to transplant tolerance induction. CONTACT Sara Alfadil salfadi1@jhmi.edu © 2016 Sara Alfadil, Kim Mi-Jeong, Marcos Iglesias Lozano, Byoung Chol Oh, W. P. Andrew Lee, Gerald Brandacher, Thomas Serworld, and Giorgio Raimondi. Published with license by Taylor & Francis. 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. The moral rights of the named author(s) have been asserted. VASCULARIZED COMPOSITE ALLOTRANSPLANTATION 2016, VOL. 3, NOS. 1–2, 46 http://dx.doi.org/10.1080/23723505.2016.1234206