Megan Sykes, M.D.

Mixed Chimerism to Achieve Transplant Tolerance

Bone marrow transplantation (BMT) from diabetes-resistant strains has already been shown to prevent insulitis and diabetes in experimental mouse models of type I diabetes. However, the routine use of BMT for the treatment of autoimmune diabetes has been prohibited so far, largely because of the unacceptable toxicity associated with the conditioning needed to achieve engraftment, and because of the complications associated with allo-BMT. We have recently described a method of inducing tolerance that uses allo-BMT and costimulatory blockade to induce high levels of allogeneic stem cell engraftment and permanent deletional tolerance, without requiring host T cell depletion or myeloablative conditioning. We have now shown that mixed chimerism, induced by a non-myeloablative, non-T cell depleting conditioning regimen is beneficial in pre-diabetic and diabetic NOD mice by 1) reversing the autoimmune T cell response and thus preventing the development and recurrence of disease; and 2) inducing tolerance toward transplanted allogeneic donor islets. We will further evaluate the potential of mixed bone marrow chimerism induced with non-toxic protocols as a treatment for diabetes in the nonobese diabetic (NOD) mouse model, which closely resembles human diabetes. Mechanistic studies will provide insight into this means of curing diabetes.

References:

1. Wekerle T, Sayegh MH, Hill J, Zhao Y, Chandraker A, Swenson KG, Zhao G and Sykes M. Extrathymic T cell deletion and allogeneic stem cell engraftment induced with costimulatory blockade is followed by central T cell tolerance. Journal of Experimental Medicine. 1998; 187:2037-2044.

2. Yang Y-G, Dey BR, Sergio JJ, Pearson DA and Sykes M. Donor-derived Interferon g is required for inhibition of acute graft-versus-host disease by Interleukin 12. J.Clin. Invest. 1998; 102:2126-2135.

3. Sykes M, Preffer F, McAfee S, Saidman SL, Weymouth D, Andrews D, Colby C, Sackstein R, Sachs DH and Spitzer TR. Mixed lymphohematopoietic chimerism and graft-vs-lymphoma effects are achievable in adult recipients following non-myeloablative therapy and HLA-mismatched donor bone marrow transplantation. Lancet 353:1755-1759, 1999.

4. Ohdan H, Yang Y-G, Shimizu A, Swenson KG and Sykes M. Mixed chimerism induced without lethal conditioning prevents T cell- and anti-Gala1,3Gal-mediated graft rejection. J. Clin. Invest. 104:281-290, 1999.

5. Wekerle T, Kurtz J, Ito H, Ronquillo JV, Dong V, Zhao G, Shaffer J, Sayegh MH, Sykes M. Allogeneic bone marrow transplantation with costimulatory blockade induces macrochimerism and tolerance without cytoreductive host treatment. Nature Med. 6:464-469, 2000.

6. Leykin I, Nikolic B, Sykes M. Mixed bone marrow chimerism as a treatment for autiommune diabetes. Transplant. Proc. 33: 120, 2001.

7. Ito H, Kurtz J, Shaffer J, Sykes M. CD4 T cell-mediated alloresistance to fully MHC-mismatched allogeneic bone marrow engraftment is dependant on CD40-CD40 ligand interactions, and lasting T cell tolerance is induced by bone marrow transplantation with initial blockade of this pathway. J. Immunol. 166: 2970-2981, 2001.

8. Wekerle T, Kurtz J, Sayegh MH, Ito H, Wells A, Bensigner S, Shaffer J, Turka LA, Sykes M. Peripheral deletion after bone marrow transplantation with costimulatory blockade has features of both activated-induced cell death and passive cell death. J. Immunol. 166: 2311-2316, 2001.

9. Mapara MY, Kim Y-M, Wang S-P, Bronson R, Sachs DH, Sykes M. Donor lymphocyte infusions (DLI) mediate superior graft-versus-leukemia (GvL) effects in mixed compared to fully allogeneic chimeras: A critical role for host antigen-presenting cells. Blood 100:1903-1909, 2002.

10. Kim Y-M, Sachs T, Asavaroengchai W, Bronson R, Sykes M. Graft-versus-host disease (GvHD) can be separated from graft-versus-lymphoma effects by controlling lymphocyte trafficking with FTY720. J. Clin. Invest. 111:659-669, 2003.

11. Kurtz J, Lie A, Griffith M, Eysaman S, Shaffer J, Anosova N, Turka L, Benichou G, Sykes M. Lack of role for CsA-sensitive or Fas pathways in the tolerization of CD4 T cells via BMT and anti-CD40L. Am J Transplant. 2003; 3:804-816.

12. Zhao Y, Ohdan H, Manilay JO, Sykes M. NK cell tolerance in mixed allogeneic chimeras. J. Immunol. 2003; 170:5398-5405.

13. Takeuchi Y, Ito H, Kurtz J, Wekerle T, Ho L, Sykes M. Earlier low dose TBI or DST overcomes CD8⁺ T cell-mediated alloresistance to allogeneic marrow in recipients of anti-CD40L. Am J Transplantation, 2004; 4:31-40.

14. Nikolic B, Takeuchi Y, Leykin I, Smith RN, Sykes M. Mixed hematopoietic chimerism allows cure of autoimmune diabetes through allogeneic tolerance and reversal of autoimmunity. Diabetes, 2004; 53 (2) 376-383.

15. Kurtz J, Shaffer J, Lie A, Anosova N, Benichou G, Sykes M. Mechanisms of early peripheral CD4 cell tolerance induction by anti-CD154 monoclonal antibody and allogeneic bone marrow transplantation: Evidence for anergy and deletion, but not regulatory cells. Blood, In press.


	Megan Sykes, M.D. Mixed Chimerism to Achieve Transplant Tolerance Bone marrow transplantation (BMT) from diabetes-resistant strains has already been shown to prevent insulitis and diabetes in experimental mouse models of type I diabetes. However, the routine use of BMT for the treatment of autoimmune diabetes has been prohibited so far, largely because of the unacceptable toxicity associated with the conditioning needed to achieve engraftment, and because of the complications associated with allo-BMT. We have recently described a method of inducing tolerance that uses allo-BMT and costimulatory blockade to induce high levels of allogeneic stem cell engraftment and permanent deletional tolerance, without requiring host T cell depletion or myeloablative conditioning. We have now shown that mixed chimerism, induced by a non-myeloablative, non-T cell depleting conditioning regimen is beneficial in pre-diabetic and diabetic NOD mice by 1) reversing the autoimmune T cell response and thus preventing the development and recurrence of disease; and 2) inducing tolerance toward transplanted allogeneic donor islets. We will further evaluate the potential of mixed bone marrow chimerism induced with non-toxic protocols as a treatment for diabetes in the nonobese diabetic (NOD) mouse model, which closely resembles human diabetes. Mechanistic studies will provide insight into this means of curing diabetes. References: 1. Wekerle T, Sayegh MH, Hill J, Zhao Y, Chandraker A, Swenson KG, Zhao G and Sykes M. Extrathymic T cell deletion and allogeneic stem cell engraftment induced with costimulatory blockade is followed by central T cell tolerance. Journal of Experimental Medicine. 1998; 187:2037-2044. 2. Yang Y-G, Dey BR, Sergio JJ, Pearson DA and Sykes M. Donor-derived Interferon g is required for inhibition of acute graft-versus-host disease by Interleukin 12. J.Clin. Invest. 1998; 102:2126-2135. 3. Sykes M, Preffer F, McAfee S, Saidman SL, Weymouth D, Andrews D, Colby C, Sackstein R, Sachs DH and Spitzer TR. Mixed lymphohematopoietic chimerism and graft-vs-lymphoma effects are achievable in adult recipients following non-myeloablative therapy and HLA-mismatched donor bone marrow transplantation. Lancet 353:1755-1759, 1999. 4. Ohdan H, Yang Y-G, Shimizu A, Swenson KG and Sykes M. Mixed chimerism induced without lethal conditioning prevents T cell- and anti-Gala1,3Gal-mediated graft rejection. J. Clin. Invest. 104:281-290, 1999. 5. Wekerle T, Kurtz J, Ito H, Ronquillo JV, Dong V, Zhao G, Shaffer J, Sayegh MH, Sykes M. Allogeneic bone marrow transplantation with costimulatory blockade induces macrochimerism and tolerance without cytoreductive host treatment. Nature Med. 6:464-469, 2000. 6. Leykin I, Nikolic B, Sykes M. Mixed bone marrow chimerism as a treatment for autiommune diabetes. Transplant. Proc. 33: 120, 2001. 7. Ito H, Kurtz J, Shaffer J, Sykes M. CD4 T cell-mediated alloresistance to fully MHC-mismatched allogeneic bone marrow engraftment is dependant on CD40-CD40 ligand interactions, and lasting T cell tolerance is induced by bone marrow transplantation with initial blockade of this pathway. J. Immunol. 166: 2970-2981, 2001. 8. Wekerle T, Kurtz J, Sayegh MH, Ito H, Wells A, Bensigner S, Shaffer J, Turka LA, Sykes M. Peripheral deletion after bone marrow transplantation with costimulatory blockade has features of both activated-induced cell death and passive cell death. J. Immunol. 166: 2311-2316, 2001. 9. Mapara MY, Kim Y-M, Wang S-P, Bronson R, Sachs DH, Sykes M. Donor lymphocyte infusions (DLI) mediate superior graft-versus-leukemia (GvL) effects in mixed compared to fully allogeneic chimeras: A critical role for host antigen-presenting cells. Blood 100:1903-1909, 2002. 10. Kim Y-M, Sachs T, Asavaroengchai W, Bronson R, Sykes M. Graft-versus-host disease (GvHD) can be separated from graft-versus-lymphoma effects by controlling lymphocyte trafficking with FTY720. J. Clin. Invest. 111:659-669, 2003. 11. Kurtz J, Lie A, Griffith M, Eysaman S, Shaffer J, Anosova N, Turka L, Benichou G, Sykes M. Lack of role for CsA-sensitive or Fas pathways in the tolerization of CD4 T cells via BMT and anti-CD40L. Am J Transplant. 2003; 3:804-816. 12. Zhao Y, Ohdan H, Manilay JO, Sykes M. NK cell tolerance in mixed allogeneic chimeras. J. Immunol. 2003; 170:5398-5405. 13. Takeuchi Y, Ito H, Kurtz J, Wekerle T, Ho L, Sykes M. Earlier low dose TBI or DST overcomes CD8⁺ T cell-mediated alloresistance to allogeneic marrow in recipients of anti-CD40L. Am J Transplantation, 2004; 4:31-40. 14. Nikolic B, Takeuchi Y, Leykin I, Smith RN, Sykes M. Mixed hematopoietic chimerism allows cure of autoimmune diabetes through allogeneic tolerance and reversal of autoimmunity. Diabetes, 2004; 53 (2) 376-383. 15. Kurtz J, Shaffer J, Lie A, Anosova N, Benichou G, Sykes M. Mechanisms of early peripheral CD4 cell tolerance induction by anti-CD154 monoclonal antibody and allogeneic bone marrow transplantation: Evidence for anergy and deletion, but not regulatory cells. Blood, In press.