Understanding Graft Rejection and Immunosuppression

1. Transplantation • David Straus, Ph.D. • Objectives • Understand the following: • 1. the types of graft rejection • 2. what are alloantigens • 3. basis for alloreactivity • 4. the classes of immunosuppressive agents and how they work • 5. why a bone marrow transplant might be done, and the importance of HLA matching in bone marrow transplants • 6. basis for graft vs. host disease • 7. problems associated with xenotransplantation • Parham, P. Chapter 12 p. 391 - 412, 417 - 420

2. Types of graft rejection Hyperacute - extremely rapid rejection (hours) initiated by pre-existing antibodies Acute - days to weeks after transplant adaptive immune response Chronic -months to years after transplant graft vascular disease

3. Hyperacute rejection occurs within hours of transplantation. Pre-existing antibodies bind the vasculature of the graft, inducing clotting and occlusion of the vessels

4. Acute graft rejection occurs 1-2 weeks following transplantation as a result of an adaptive immune response

5. “Chronic” rejection occurs months or years following transplantation, and is typified by graft vascular disease associated with inflammatory injury Concentric fibrosis in cardiac allograft

6. Graft rejection occurs as a result of a host immune response to antigens on donor tissue - an “alloresponse” to “alloantigens” Alloantigens - structural differences which are recognized by the recipient’s immune system. -Blood group antigens, -HLA, -minor histocompatibility antigens (not HLA)

7. Erythrocytes express the A, B, O blood group antigens. These alloantigens are glycolipids with distinct carbohydrate structures

8. HLA (MHC) molecules present peptide antigens

9. CD8+ T cells bind MHC I - peptide and CD4+ T cells bind MHC II-peptide

10. HLA genes are highly polymorphic The HLA locus encodes multiple class I and class II genes Numbers of different HLA gene alleles found in the human population

11. HLA polymorphisms can effect peptide binding and T cell receptor binding

12. HLA alloreactivity mechanisms Transplant recipient T cells have not been restricted during their development by the donor HLA: -Presentation of a distinct self-peptide spectrum on donor HLA may result in activation of some recipient T cells. -Recipient T cells may respond directly to allogeneic HLA on donor APCs. Donor HLA may be also be processed and presented to recipient T cells like any pathogen antigen (or other alloantigens).

13. Alloreactivity can be determined by peptide determinants, or by HLA determinants

14. Mixed Lymphocyte Response (MLR) demonstrates direct alloreactivity

15. Non-HLA antigens Many fewer T cells are able to respond to allogenic differences outside of HLA genes However, minor histocompatability differences may be sufficient to mediate rejection in HLA-identical transplants Minor histocompatibility antigens

16. What APCs are responsible for stimulating acute graft rejection?

17. Both donor and recipient APCs can present alloantigens to recipient T cells Allogeneic HLA may be recognized directly on donor APCs, or processed and presented on recipient APCs. Additionally, “novel” peptides may be presented by allogeneic HLA on donor APCs.

18. Improving engraftment HLA matching Immunosuppression non-specific: corticosteriods, cytotoxic drugs specific: cyclosporin A, FK506, rapamycin anti-T cell antibodies Tolerance induction

19. Corticosteriods Non-specific immunosuppressant Used acutely Modulates gene expression; blocks production of inflammatory cytokines • Prednisone – hydrocortisone derivative, is a pro-drug; • converted to its active form, prednisolone, in vivo

20. Cytotoxic drugs: kill dividing cells by inhibiting DNA replication Azathioprine: blocks purine metabolism Cyclophosphamide: alkylates DNA Methotrexate: inhibits thymidine synthesis

21. Calcineurin phosphatase controls NFAT activity

22. Cyclosporin A and tacrolimus block T cell activation by inhibiting calcineurin and NFAT function CsA and tacrolimus bind target proteins in the cytosol The drug- protein complex associates with calcineurin preventing the dephosphorylation of NFAT NFAT is unable to translocate into the nucleus to activate IL-2 gene expression

23. Antigen recognition in the absence of co-stimulation leads to tolerance

24. Blocking CD28 co-stimulation signals with CTLA4Ig can and suppress immune responses and enhance graft survival

25. Bone Marrow Transplantation Reconstitution of hematopoietic system through transfer of stem cells. Restoration of immune function requires shared HLA alleles so that T cell repertoire selected during development can function with bone marrow derived APCs. Transfer of T cells in bone marrow presents reciprocal problem of graft rejection: graft vs. host disease.

26. Bone marrow transplantation can reconstitute the hematopoietic system in cases of inherent or induced deficiency

27. Reconstitution of a functional immune system requires sharing of some HLA haplotypes: T cell selection in the host thymus determines recognition of donor APCs

28. Mature T cells in bone marrow can mediate graft vs. host disease: systemic “autoimmune” disease

29. Graft-versus-Host Disease is associated with pre-treatment tissue damage 1. Pretreatment to ablate host immune system, or reduce malignancy, causes tissue damage. 2. APCs recognize damage and activate allo-responsive donor T cells. 3. Activated T cells and other immune cells induce tissue damage.

30. Depletion of T cells from bone marrow prior to transplant reduces graft-versus-host disease, but also graft vs. leukemia effect

31. You are waiting for a kidney to replace your own, but time is running out. Out of desperation you accept a kidney from a donor with a substantially different HLA type. Initially the graft is accepted, but as you are weaned off immunosuppressive drugs, a T cell response develops and the kidney is rejected. A). How can the difference in HLA type lead to a T cell response? B). If APCs could be depleted from graft, how would it change the success of the engraftment? If APCs were depleted in you (the recipient) instead, how would this change the T cell response? C). If the same donor also provided bone marrow for a transplant at the same time as the kidney, would this improve kidney graft survival?

32. Xenotransplantation • Why pigs? • non-primates with organs of similar size to humans. • can make transgenic pigs to address some cross-species problems • Problems • Hyperacute rejection by pre-existing xenoantibodies. Complement inhibitory proteins are unable to function across • a large species gap which increases hyperacute rejection of xenogenic tissues. • A potential risk of transferring cross-species pathogens under immunosuppressive conditions