Desensitization and Prevention of Antibody-mediated Rejection in Vascularized Composite Allotransplantation by Syngeneic Hematopoietic Stem Cell Transplantation

AbstractBackgroundCandidates for vascularized composite allotransplantation (VCA) are frequently sensitized, putting them at risk for antibody-mediated rejection (AMR). Current desensitization strategies are imperfect and require a living-donor setting. Here we investigated the impact of sensitization on and the efficacy of a desensitization protocol utilizing syngeneic hematopoietic stem cell transplantation (HSCT) to prevent AMR in VCA.MethodsSkin transplants from Dark Agouti (DA) to Lewis rats were performed for sensitization. Orthotopic hind-limb transplants from DA donors were performed to sensitized and non-sensitized recipients, and the animals were treated with either daily tacrolimus or no immunosuppression. A desensitization protocol consisting of total body irradiation (TBI), fludarabine and syngeneic HSCT was applied to sensitized animals. Graft rejection was monitored by clinical assessment and histological analysis. Serum levels of donor-specific antibodies (DSA IgG) were measured using flow-cytometry.ResultsSensitized recipients exhibited accelerated rejection by 5.5±1.2 days without immunosuppression and 10.2±3.6 days with daily tacrolimus, compared to 8.7±1.2 days and >30 days in non-sensitized recipients, respectively. Serum levels of DSA IgG were markedly elevated (37.3±3.34-fold from baseline) in sensitized recipients after VCA and correlated with histologic evidence of rejection and C4d deposition. Desensitization significantly reduced DSA compared to sensitized-controls (2.6±0.5-fold vs 6.0±1.2-fold, p30 days without evidence of C4d deposition (n=6).ConclusionsIn summary, sensitization leads to accelerated rejection of VCA, and syngeneic HSCT combined with conventional immunosuppression effectively reduces DSA and improves allograft survival in sensitized rats. Background Candidates for vascularized composite allotransplantation (VCA) are frequently sensitized, putting them at risk for antibody-mediated rejection (AMR). Current desensitization strategies are imperfect and require a living-donor setting. Here we investigated the impact of sensitization on and the efficacy of a desensitization protocol utilizing syngeneic hematopoietic stem cell transplantation (HSCT) to prevent AMR in VCA. Methods Skin transplants from Dark Agouti (DA) to Lewis rats were performed for sensitization. Orthotopic hind-limb transplants from DA donors were performed to sensitized and non-sensitized recipients, and the animals were treated with either daily tacrolimus or no immunosuppression. A desensitization protocol consisting of total body irradiation (TBI), fludarabine and syngeneic HSCT was applied to sensitized animals. Graft rejection was monitored by clinical assessment and histological analysis. Serum levels of donor-specific antibodies (DSA IgG) were measured using flow-cytometry. Results Sensitized recipients exhibited accelerated rejection by 5.5±1.2 days without immunosuppression and 10.2±3.6 days with daily tacrolimus, compared to 8.7±1.2 days and >30 days in non-sensitized recipients, respectively. Serum levels of DSA IgG were markedly elevated (37.3±3.34-fold from baseline) in sensitized recipients after VCA and correlated with histologic evidence of rejection and C4d deposition. Desensitization significantly reduced DSA compared to sensitized-controls (2.6±0.5-fold vs 6.0±1.2-fold, p30 days without evidence of C4d deposition (n=6). Conclusions In summary, sensitization leads to accelerated rejection of VCA, and syngeneic HSCT combined with conventional immunosuppression effectively reduces DSA and improves allograft survival in sensitized rats. These authors contributed equally to this work, Howard D. Wang, Samuel A.J. Fidder. Corresponding Author: Gerald Brandacher, MD, Associate Professor, Ross Research Building, Room 749A, 720 Rutland Avenue, Baltimore, MD 21205, Email: brandacher@jhmi.edu Authorship Contributions Howard D. Wang: Study design, data acquisition and analysis, manuscript composition and revision Samuel A.J. Fidder: Study design, data acquisition and analysis, manuscript composition and revision Devin T. Miller: Study design, data acquisition and analysis, manuscript composition Georg J. Furtmüller: Data acquisition and manuscript revision Ali Reza Ahmadi: Study design, data acquisition and analysis, manuscript revision Felix Nägele: Data acquisition and manuscript revision Joseph Lopez: Data acquisition and manuscript revision Amy Quan: Data acquisition and manuscript revision Joshua Budihardjo: Data acquisition and manuscript revision Denver M. Lough: Data acquisition and analysis, manuscript revision Burcu Akpinarli: Data acquisition Joanna Etra: Data acquisition Dalibor Vasilic: Study design, manuscript revision Giorgio Raimondi: Study conception and design, manuscript revision W.P. Andrew Lee: Study conception and design, manuscript revision Richard Montgomery: Study conception and design, manuscript revision Zhaoli Sun: Study conception and design, manuscript revision Gerald Brandacher: Study conception and design, manuscript revision Disclosure: The authors of this manuscript declare no conflicts of interest Funding: This work was partially supported by the Plastic Surgery Foundation and the Russel Scholarship from the Johns Hopkins Military and Veterans Health Institute. Copyright © 2018 Wolters Kluwer Health, Inc. All rights reserved.

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