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Composite Tissue Allotransplantation in Swine Across a Full MHC Barrier Using a Mixed Hematopoietic Chimerism Protocol
Angelo A. Leto Barone, MD, Radbeh Torabi, Mark A. Randolph, BS, Raimon Duran-Struuck, DVM, Robert J. Hawley, Ph.D., Ben M. Horner, MD, Abraham Matar, Rebecca Crepeau, Alexander Albritton, Christopher Mallard, Yuqing Tang, Aseda Tena, Joseph R. Scalea, MD, Josef M. Kurtz, Ph.D., Christene A. Huang, Ph.D., David H. Sachs, M.D., Curtis L. Cetrulo, Jr., M.D..
Harvard Medical School, Massachusetts General Hospital, Transplantation Biology Research Center, Boston, MA, USA.
Composite tissue allotransplantation (CTA) is currently limited by acute and chronic rejection, and the morbidity of immunosuppression. Using MHC-defined miniature swine, we have investigated various approaches to induce immune tolerance to achieve CTA acceptance across both minor and major histocompatibility (MHC) barriers. Animals exhibiting transient mixed chimerism following haploidentical hematopoietic cell transplantation have demonstrated acceptance to all components of a CTA with the exception of skin. When stable mixed chimerism is obtained, the skin component is also accepted. The aim of this study was to induce stable mixed chimerism across a full MHC barrier through bone marrow transplantation (BMT) to facilitate long-term acceptance of a simultaneously transplanted CTA.
CTA/BMT were performed across a full MHC barrier (swine leukocyte antigen SLAcc→SLAdd). Recipients underwent low-dose (100cGy) total body irradiation, T-cell depletion with CD3-Immunotoxin and a 45-day course of Cyclosporine (CyA). A gracilis myocutaneous CTA was transplanted along with BMT (Fig. A). CyA was maintained between 400-800 ng/mL for 30 days and tapered off by day 45. Flap viability was assessed daily by clinical exam and serial biopsies. Chimerism and serum alloantibody against donor MHC were assessed by flow cytometry, and immunoresponse was measured by Mixed-Lymphocyte-Reaction and Cell-Mediated-Lymphocytotoxicity assays. Control animals did not undergo the conditioning regimen and received CTA.
Controls receiving a CTA without conditioning rejected all components of the flap by day 6 post-transplantation (PTD6). In the first experimental no detectable chimerism was observed. Erythema in the flap occurred on PTD8, suggesting a rejection crisis of the skin component of the CTA, while the muscular component remained normal on biopsy. Surprisingly, the skin recovered to 100% viability by PTD13 and all components of the flap appeared viable up to PTD43. Upon cessation of CyA, the epidermis sloughed by PTD45, and was followed by loss of all components of the flap by PTD71. In the second, myeloid chimerism was observed at different timepoints (Fig.B). All components of the flap are accepted up to PTD43 in this animal (Fig.C) and monitoring of additional timepoints is ongoing, as immunosuppression is tapered.
We are currently developing a clinically-relevant concomitant CTA/BMT transplant strategy in a preclinical large animal model. The conditioning regimen used prolonged survival of a CTA across a full MHC barrier. Prolonged survival of skin and muscle was observed in one animal up to PTD43. A second animal’s CTA has all components currently viable to PTD40, and is currently ongoing. Improved T-cell depletion and increased bone marrow cell doses favorable to production of transient or stable mixed chimerism are under investigation.
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