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ACHIEVING TOLERANCE IN A MISMATCHED VCA TRANSPLANT WHILE REDUCING THE RISK OF GVHD: The Goal of Transient Chimerism
Bruce J. Swearingen, MD1, Jeff Chang, MD1, David W. Mathes, MD1, Tiffany Butts, N/A2, Scott Graves, PhD2, Rainer Storb, MD2.
1University of Washington Medical Center, Seattle, WA, USA, 2The Fred Hutchinson Cancer Research Center Division of Transplantation Biology, Seattle, WA, USA.
Background: Transplantation of vascularized composite allografts (VCA) to reconstruct devastating facial injuries and lost extremities offer the opportunity to truly restore both form and function. Due to the necessity of life-long immunosuppression, the clinical application of these techniques is limited. One promising method of inducing tolerance to an organ allograft is the development of mixed chimerism. We have previously demonstrated that a non-myeloablative stem cell transplant can lead to tolerance in a mismatched dog model. However, the application of this protocol has been limited by graft-versus-host disease (GVHD). We have observed several animals that, after an initial period of donor cell engraftment, lost their stem cell allograft but remained tolerant to the VCA. Conversely, animals that retained persistent donor cell chimerism inevitably developed GVHD. The hypothesis for this study was that our non-myeloablative hematopoietic stem cell transplant protocol could be used to induce tolerance to a recipient VCA without the need for persistent donor cell chimerism. To more closely mimic the clinical setting, our protocol was modified to an extended tapered cessation of immunosuppression.
Methods: 5 Haploidentical canine recipients (median weight and age: 12.1kg and 10.9 months) received a non-myeloablative conditioning regimen of 350 cGy TBI, mobilized donor stem cells (PBMC) and VCA transplantation followed by a short course of immunosuppression (MMF for 56 days and Cyclosporine for 70 days). Peripheral blood chimerism was evaluated by PCR techniques weekly. Peripheral blood cytokine expression was evaluated by flow cytometry. VCA rejection was followed clinically and confirmed histologically after routine biopsies. 3 haploidentical canine recipients (median weight 13.3kg) were then transplanted following a modified immunosuppression protocol. Chimerism and cytokine expression were evaluated.
Results: All 5 animals tolerated the conditioning regimen. One dog rejected the PBMC at 35 days post transplantation and went on to reject the VCA transplant following the cessation of immunosuppression. One dog fully engrafted and converted to 100% donor chimerism and long-term tolerance to the VCA but developed GVHD. 3 dogs demonstrated a prolonged period of transient chimerism (7 to 10 weeks post-transplant) and went on to reject their donor stem cells after the cessation of immunosuppression
without acute rejection of their donor VCAs. One of these dogs was euthanized for persistent fevers at post-operative day 147 with no sign of rejection. The remaining two had long-term acceptance of their VCA (>200 days) with no evidence of acute rejection but have recently demonstrated evidence of chronic rejection. No dog developed GVHD. In the modified protocol, all 3 animals tolerated the conditioning regimen. All dogs fully engrafted and converted to 100% donor chimerism and long-term tolerance to the VCA. One dog developed GVHD upon cessation of immunosuppression.
Conclusions: We demonstrate that our non-myeloablative protocol allows for tolerance to the VCA with selective rejection of the PRBMC and reduction of GVHD risks while persistent donor chimerism can lead to GVHD.
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