Immunosuppression in Transplantation

Physiology of Immunorecognition

The immune system evolved to discriminate self from non-self, and this response against non-self consists of an array of receptor-mediated sensing and effector mechanisms broadly described as innate and adaptive. Innate immunity is primitive, does not require priming, and is of relatively low affinity but broadly reactive. Adaptive immunity is antigen-specific, depends on antigen exposure or priming, and can be of very high affinity. The major effectors of innate immunity are complement, granulocytes, monocytes/macrophages, natural killer (NK) cells, mast cells, and basophils. The major effectors of adaptive immunity are B and T lymphocytes.

A transplant between genetically distinct individuals of the same species is called an allogeneic graft, or an allograft . The immune response to an allograft requires three elements: recognition of foreign antigens, activation of antigen-specific lymphocytes, and the effector phase of graft rejection. The recognition of antigens as peptide fragments bound to major histocompatibility complex (MHC) molecules, known as human leukocyte antigens (HLAs), is the central event in the initiation of an alloresponse. HLA molecules ( Fig. 61.1 ) are highly polymorphic, follow Mendelian codominant inheritance, and constitute the principal antigenic barrier to transplantation. The degree of HLA matching between the donor and the recipient has been long recognized to play an important role in graft survival, and HLA matching has been incorporated into kidney allocation. More recently, mismatched “eplets,” or patches of polymorphic residues within a radius of 3.0–3.5 Å, which form essential components of HLA epitopes recognized by antibodies, have been found to be a significant risk factor for the development of de novo donor-specific antibody, chronic humoral rejection, and graft loss. In addition, non-HLA molecules, such as MHC class I–related chain A (MICA), are recognized as playing a significant role in rejection, particularly in recipients of well–HLA-matched kidneys.

There are two types of HLA molecules: class I and class II. Class I HLA molecules are expressed on all nucleated cells, whereas class II molecules are usually expressed only on antigen-presenting cells (APCs), which include dendritic cells, B lymphocytes, and macrophages. Cytokines such as interferon-γ (IFN-γ) induce, upregulate, and broaden HLA expression, so that all cells in a graft can become potential targets of the immune response. Ischemia-reperfusion injury in the graft leads to the production of inflammatory cytokines and recruitment of macrophages, and acute rejection episodes are more common in grafts with prolonged ischemia times. Recipient T cells may respond directly to peptides/HLA complexes presented by donor APCs in the graft or to donor HLA peptides presented on the recipient’s own APCs ( Fig. 61.2 ). Acute rejection of an allograft is believed to be primarily dependent on direct allorecognition, whereas the indirect pathway may play a larger role in chronic rejection.

T cells are critically important in the rejection of allogeneic grafts. CD4 T cells (helper T cells) are thought to mediate the initial recognition of an allograft as well as amplify and coordinate the subsequent immune response, including providing help to CD8 (effector) T cells. T cell recognition of the alloantigen occurs via binding of the T cell receptor (TCR)/CD3 complex on the T cell’s surface to the peptide/MHC complex on APCs. This is referred to as signal 1 and leads to phosphorylation of TCR-associated proteins and downstream activation of several pathways, including calcineurin, protein kinase C, and mitogen-activated protein (MAP) kinase pathways. The calcineurin pathway is the most well characterized, and it involves the activation of calcineurin (a phosphatase) by an increase in cytosolic calcium. Calcineurin dephosphorylates nuclear factor of activated T cells (NFAT), allowing NFAT to translocate from the cytoplasm to the nucleus. The NFAT binds to regulatory sequences and increases gene transcription of several cytokines, including interleukin (IL)-2, a T cell growth factor, as well as IL-4, IFN-γ, and tumor necrosis factor-α (TNF-α).

Although the specificity of the immune response is determined by signal 1 , a costimulatory signal, signal 2, which occurs though accessory molecules, is essential for T cell activation. The most potent of these signals regulating T cell clonal expansion and differentiation is provided by the B7/CD28 family of molecules ( Fig. 61.3 ). B7-1 (CD80) and B7-2 (CD86) are ligands on APCs that bind to CD28, expressed on most T cells. Engagement of CD28 increases the production of IL-2 and other cytokines, resulting in T cell proliferation. CD80 and CD86 also regulate T cells by binding another antigen on T cells called cytotoxic T-lymphocyte antigen-4 (CTLA-4) , which inhibits T cell proliferation. A costimulatory interaction between CD40 on APCs and CD40 ligand (CD154, CD40L) on T cells is also critical for activation of APCs and upregulation of B7 expression on T cells. One way to induce T cell anergy in vitro is to provide the T cell with an antigen-specific signal through the TCR (signal 1) in the absence of CD28 engagement (signal 2). However, in most in vivo models of B7 blockade, anergy has been difficult to demonstrate, which is possibly due to the complexity of costimulation that involves multiple stimulatory and inhibitory signals.

Antigen-specific activation of T cells, particularly CD4 T cells, leads to the production of cytokines, the recruitment of monocytes, and the proliferation of CD8 T cells, NK cells, and B cells. CD8 T cells cause cell death in the graft through the release of soluble cytotoxic factors (granzymes and perforin) as well as upregulated Fas ligand on T cells that bind to Fas (CD95) on target cells and trigger apoptosis.

In addition to T cells, B cells and the humoral arm of the immune system play a major role in acute and chronic graft injury. Antibodies produced by the differentiation of B cells into plasma cells cause cell injury through complement fixation or antibody-dependent cellular cytotoxicity. Hyperacute rejection occurs when preformed recipient antibodies to donor HLA antigens or ABO blood group antigens result in complement activation, intravascular coagulation, and graft necrosis within 24 hours of transplantation. Although cross matching and ABO blood typing have virtually eliminated hyperacute rejection, B cells and plasma cells continue to play an important role in subsequent antibody-mediated rejection (AMR) and may be important mediators of chronic graft injury and late graft loss.

Strategies for Immunosuppression

The first attempts at immunosuppression used total-body irradiation. Azathioprine was introduced in the early 1960s, and soon thereafter was routinely accompanied by prednisolone in an immunosuppressive regimen. The polyclonal antilymphocyte antibody preparations became available in the mid-1970s. The introduction of cyclosporine in the early 1980s dramatically improved 1-year graft survival rates from 50% to over 80%, and in 1985, OKT3, a monoclonal antibody to CD3, was introduced for the treatment of acute rejection. In the 1990s, tacrolimus and mycophenolate mofetil (MMF) emerged as alternatives to cyclosporine and azathioprine, anti–IL-2 receptor antibodies were approved for induction, and sirolimus became available. Everolimus was approved in 2010 for use in kidney transplant recipients in combination with calcineurin inhibitors. In 2011, belatacept was approved as the first biologic agent for use in maintenance immunotherapy. Commonly used immunosuppressants and their mechanisms of action are listed in Table 61.1 .

Transplant immunosuppression is guided by three key principles: (1) multiple agents directed at different molecular targets within the alloimmune response are used simultaneously to maximize synergy and efficacy while minimizing toxicity; (2) greater immunosuppression (induction) is needed for early engraftment or to treat established rejection rather than for long-term graft maintenance; and (3) continuous vigilance is essential to identify rejection, drug toxicity, and infection so that the immunosuppressive regimen can be modified appropriately.

Mechanisms of Action of Immunosuppressive Drugs

T cells have historically been the major target of immunosuppression. The three-signal model of T cell activation and subsequent cellular proliferation provides a useful guide to the sites of action of the major immunosuppressive agents ( Fig. 61.4 ). Signal 1 is the antigen-specific signal provided by the interaction of the MHC/peptide complex on APCs with the TCR/CD3 complex. Signal 2 is a non-antigen-specific costimulatory signal provided by the engagement of B7 on APCs with CD28 on the T cell. These two signals activate intracellular pathways, leading to the production of IL-2 and other cytokines. Stimulation of the IL-2 receptor (CD25) leads to activation of mammalian target of rapamycin (mTOR), a protein kinase, and provides signal 3, which triggers cell proliferation. Therapies targeting antibody-mediated injury are directed against B cells, plasma cells, and complement activation. In general, all drugs in current clinical use have been more effective at suppressing primary rather than memory immune responses.