The immune response and immunosuppressant drugs - Medical Pharmacology and Therapeutics, 4e

Medical Pharmacology and Therapeutics, 4e

38. The immune response and immunosuppressant drugs

Biological basis of the immune response

Innate immunity

Adaptive immunity

Unwanted immune reactions

Immunosuppressant drugs

Calcineurin inhibitors

Mammalian target of rapamycin inhibitors

Antiproliferative agents

Folic acid antagonist

Interleukin-2 receptor antibodies

Selective co-stimulation blocker

Immunosuppression in organ transplantation

Immunosuppression in other disorders

Biological basis of the immune response

The immune system is composed of innate (natural) and adaptive components, which protect the host against a wide variety of pathogens and also tumour cells. The adaptive system is further divided into humoral and cell-mediated immunity. Overall the immune system has the ability to distinguish between self and non-self proteins and to protect the host (self) against non-self infectious and other pathogenic agents.

Innate Immunity

The term innate immunity (sometimes called natural immunity) was used because it is an inherited system and is part of our genetic make-up. It is made up of several generally non-specific protective mechanisms, some of which do not involve the immune system. All foreign pathogens are recognised approximately equally by pattern-recognition receptors on cells involved in the innate immune system. Pattern-recognition receptors identify pathogenic ‘groups’ such as bacterial lipopolysaccharides and bacterial DNA rather than responding to individual antigens. The innate immune system has an important role in processing foreign pathogens and triggering the highly selective adaptive system described below.

The innate immune system incorporates the following processes:

physicochemical barriers, e.g. intact skin and mucous membrane, low stomach pH, antibacterial agents (lysozyme) in skin and tear secretions,

macrophages and dendritic cells, particularly in lungs, liver, lymph nodes and spleen, phagocytose pathogenic material and produce antigen fragments (short peptides of approximately 8–25 amino acids) from the pathogenic material, which they then display on their surfaces. The cells are then described as antigen-presenting cells (APCs) and are necessary for the presentation of the antigen to T-lymphocytes and the triggering of the adaptive immune system (see below),

attraction of immune cells to sites of inflammation by substances released from cells such as cytokines,

phagocytosis of bacteria and parasites by granulocytes, including neutrophils, monocytes and macrophages,

actions of natural killer cells (large granular lymphocytes),

binding of antigens to IgE antibody on mast cells and basophils and the subsequent release of inflammatory mediators from the cell,

fever,

complement activation.

The innate immune system may be an adequate defence to deal with many pathogens but, unlike adaptive immunity, long-term specific immune protection following initial exposure to a pathogen does not occur.

Adaptive Immunity

Adaptive immunity is superimposed upon the innate mechanisms. It differs from innate immunity in that it is slower to respond, offers long-term specific protection and has exquisite specificity in recognising individual non-self molecules. Adaptive immunity has two basic complementary and interacting mechanisms: cell-mediated immunity and humoral immunity (Figs 38.1 and 38.2).

FIG. 38.1 Aspects of cell-mediated immunity.

This shows in simplified form some steps in T-cell activation following antigen presentation to the T-cell receptor (TCR). Pathogenic antigens are presented by antigen-presenting cells to the uncommitted CD4⁺ lymphocyte which carries the specific receptor to the antigen, in association with major histocompatibility complex (MHC) class I and co-stimulatory molecules. Under the influence of interleukin-2 (IL-2), Th1 cells undergo clonal proliferation and play a variety of roles in cell-mediated immunity, including activation of macrophages and other cells. Antigens can also be presented to CD8⁺ lymphocytes, which mature into cytotoxic T-cells. Drugs used as immunosuppressants (red arrows) act at the sites shown. Corticosteroids act at many sites (see also Fig. 38.2 and Ch. 44). Ag, antigen; IL-2R, interleukin-2 receptor; Th, T-helper cell.

FIG. 38.2 Aspects of humoral immunity.

Adaptive immunity can result in production of antibodies (humoral response) or a cell-mediated response (Fig. 38.1). Antigens on bacteria or bacterial toxins bind to immunoglobulins on B cells. Before proliferation and antibody production can occur, the B-cell also has to be stimulated by activated T-cell cytokine production, usually of the Th2 type. Antigen fragments can be presented with major histocompatibility complex (MHC) class II molecules to T-cells via a T-cell receptor (TCR) that recognises the antigen. T-cells undergo clonal proliferation and produce cytokines that stimulate B-cells to produce humoral antibodies (IgG, IgM, IgA, IgD and IgE). In atopic individuals, the T-cells are tipped towards the Th2 type and produce interleukins IL-4, IL-5, IL-10 and transforming growth factor β (TGFβ), which induce the B-cells to produce IgE. Ag, antigen; APC, antigen-presenting cell; CR, cytokine receptor; P, plasma cell; Th, T-helper cell.

Essential components of the adaptive system are the two populations of lymphocytes:

T-lymphocytes are produced in the bone marrow and migrate to the thymus, where they mature, express receptors for antigens and interact with immunogenic self peptides. T-cells are selected in the thymus for low or high avidity for self peptides, and those showing high avidity are destroyed. The surviving T-cells retain the potential to cross-react with multiple foreign non-self antigens but not self molecules.

B-lymphocytes make up about 10% of the lymphocyte population and mature in the bone marrow.

T-cells and B-cells are coated with vast numbers of proteins which act as receptors or as ligands for other receptors. These proteins can be defined by antibody typing (immunophenotyping) and are given cluster of differentiation (CD) numbers such as CD4, CD8, etc. When T-cells leave the thymus they are considered naïve or uncommitted since they have not yet been exposed to the non-self antigens. At this stage, naïve T-cells consist of two major populations, known as helper (Th; CD4⁺) and cytotoxic or killer (Tc; CD8⁺) T-cells (Fig. 38.1). Immunogenic peptides are presented to T-cells within the cleft of major histocompatibility complex (MHC) class II molecules on the surface of APCs from the innate immune system. The CD4⁺ (Th) cells have surface receptors with a high affinity for class II MHC which binds antigenic peptides on APCs. The CD8⁺ (Tc) cells have an affinity for class I MHC, which displays specific antigens on the surface of tumour cells or infected host cells.

The CD4⁺ Th-cell is activated if its receptors recognise and bind avidly to an antigen, but only if the antigen is processed and presented on an APC. This triggers a series of complex activation pathways which prepare the Th-cell for its immune role. Acivated Th-cells secrete the T-cell growth factor interleukin-2 (IL-2), which acts in an autocrine fashion on the Th-cells and causes them to proliferate. The Th-cells are then committed to become a type 1 Th-cell (Th1) or a type 2 Th-cell (Th2). The pattern of differentiation may be determined by the type of antigen or possibly the concentration of antigen presented to the Th-cell. Differentiated Th-cells orchestrate the immune response by secretion of specific cytokines (Figs 38.1 and 38.2). Th1-cells interact with macrophages to enhance their phagocytic activity and stimulate the proliferation of cytotoxic Tc-cells. Th2-cells stimulate B-cells to grow and divide, and activate humoral immune responses. The details of these responses are not well understood, and the existence of other Th-cell populations (such as Th3 and Th17) adds to the complexity of this cell family. The activities of Th-cells and APCs are modulated by regulatory T-cells (Treg).

Co-stimulatory signals involving cell surface CD proteins are important processes in immune cell function. For example, CD40 protein on APCs interacts with Th-cell proteins and has a co-stimulatory role in APC activation. The CD80 and CD86 proteins on APCs interact with CD28 proteins on Th- and Tc-cells to co-activate them together with Th-cell cytokines. The expression of CD20 protein on B-cells enables an optimum immune response against T-cell-independent antigens (antigens that elicit a full humoral immune response without participation of T-cell cytokines).

Immature B-cells can bind antigen with the cooperation of T-cells, but on subsequent exposure antigen binds directly to immunoglobulins on the B-cell (Fig. 38.2)

Cell-mediated immunity

Cell-mediated immunity is largely T-cell-driven, utilising Th1 (CD4⁺) and cytotoxic T-cell (CD8⁺) subtypes, and is involved in responses to viral infection, graft rejection, chronic inflammation and tumour immunity. Figure 38.1 shows schematically the basic processes occurring in cell-mediated immunity. T-cells that possess the receptor to the antigen of the invasive pathogen that is presented on APCs are stimulated to express IL-2 and the IL-2 receptor. For clarity, the many co-stimulatory processes that are described in the text above are not shown in the figure.

Stimulation of the IL-2 receptor induces the Th-cell to:

activate macrophages to phagocytose the pathogen,

stimulate cytotoxic T-cells to proliferate,

stimulate B-cells to proliferate,

attract macrophages and neutrophils to the site,

produce memory B-cells that respond rapidly to the pathogen on future exposure.

Cytotoxic T-cells that recognise the foreign antigen presented on MHC type I on APCs are activated to proliferate and attack pathogens expressing the antigen. When cytotoxic cells bind to an antigen on a pathogenic cell they release a variety of proteases or lysins to destroy the cell.

Humoral immunity

Figure 38.2 illustrates the basic processes in humoral immunity. The foreign antigen is recognised by immunoglobulin (Ig) molecules or specific receptors to that antigen on the surface of a specific clone of B-cells. The presence of nearby Th2-cells that have been activated to secrete IL-5 and IL-10 is required for initial antigen recognition by B-cells. The secreted interleukins cause B-cell clonal proliferation, and convert B-cells into active plasma cells that can secrete antibodies of the IgG, IgM, IgA, IgD and IgE classes which bind to and destroy pathogenic antigens.

On encountering an antigen, the primary immune response consists of IgM, replaced later by IgG. B-cells that are primed to produce specific antibodies survive as memory B-cells. On a further encounter with the antigen, the secondary immune response occurs more rapidly and consists of large amounts of IgG produced by plasma cells derived from reactivation of memory B-cells.

Unwanted Immune Reactions

The processes of inflammation and immunity described above are essential to protect the host against pathogens and other damage, but excessive, inappropriately prolonged or misdirected immune responses can cause disease, including hypersensitivity reactions, graft rejection and autoimmune diseases.

It is not always easy to decide whether predominantly Th1- or Th2-mediated immune responses are involved in a particular disease; in part this is due to the fact that Th2 cytokines can inhibit Th1-cell functions. Th1-mediated immune responses are significantly involved in rheumatoid arthritis (Ch. 30) and in the formation of atheroma (Ch. 48). Th2-mediated responses are important in mild to moderate asthma but with increasing participation of Th1-mediated responses in severe asthma (Ch. 12).

Hypersensitivity reactions

Hypersensitivity reactions were classified by Gell and Coombs in the late 1950s.

Type 1 (Acute, Immediate): This category includes hay fever and acute asthma. IgE molecules on the surface of mast cells and basophils are crosslinked by harmless antigens (allergens such as pollens or house dust mites), leading to the synthesis and/or release of inflammatory mediators. These include cysteinyl-leukotrienes, prostaglandins, histamine, platelet-activating factor, proteases and cytokines.

Type 2 (Cytotoxic): Cell surface antigens, including microbial proteins and drug molecules haptenised onto cell surfaces, are recognised and bound by IgG and IgM antibodies (opsonisation), leading to activation of complement (classic pathway) and cytolysis of the target cell. Examples include destruction of red cells after incompatible blood transfusion, and haemolytic anaemia caused by binding of some drugs to host cells (see Ch. 53).

Type 3 (Complex-Mediated): Soluble antigens react with excess circulating antibodies to form complexes that precipitate in small blood vessels, causing vasculitis and organ damage. The various forms of extrinsic allergic alveolitis, caused by exposure to animal or vegetable dusts, are systemic type 3 reactions, while the Arthus reaction is a local response to an injected antigen (e.g. non-human insulins).

Type 4 (Cell-Mediated, Delayed-Type Hypersensitivity): Inappropriate regulation of cell-mediated immunity may cause damaging chronic inflammation, leading to fibrosis and granuloma formation. Cell-mediated immunity misdirected against harmless foreign proteins (allergens) can lead to chronic allergic inflammation (such as occurs in eczema), or cause contact sensitivity in the skin to haptenising metals and chemicals. In allergy, Th2-cells secrete cytokines, including IL-4, IL-5 and IL-13, which promote eosinophilic inflammation and overproduction of IgE by B-cells.

Transplant rejection

In blood transfusion, rejection usually occurs because non-self antigens on the transfused red blood cells (ABO system) trigger a type 2 hypersensitivity reaction in the recipient. In immunodeficient people, transfused T-cells react against recipient antigens (graft-versus-host reactions).

For organ transplants, hyperacute rejection can occur if there is ABO incompatibility, or host-versus-graft reactions can arise later with foreign MHC molecules (human leucocyte antigens, HLA). The latter can be reduced by HLA tissue typing to increase the chance of selecting a graft that is compatible with the host tissues. This will reduce the rate of tissue destruction but not prevent chronic rejection. The immune response and its place in the rejection of a transplanted organ are complex. The antigens on the graft are recognised as foreign, and the cascaded responses outlined in Figures 38.1 and 38.2 occur, with increased production of B-cells, cytotoxic T-cells and monocyte/macrophages. Graft destruction occurs from antibody production against the graft, lysis of graft cells and delayed hypersensitivity responses. Rejection can be immediate (days), acute (weeks) or chronic (years).

Autoimmunity

Normally the immune system is tolerant of self antigens. T-cells in the thymus that express receptors with high avidity for self peptides are normally destroyed in a process known as negative selection (see above). If this self-tolerance breaks down, autoimmune disorders result. Numerous mechanisms can trigger autoimmune diseases, including viral infection of host cells, binding of drug molecules to host cells (e.g. penicillin), sharing of antigens between host cells and microbes and sequestration of antigens liberated by cell damage. Examples of autoimmune disease include haemolytic anaemia, type 1 diabetes mellitus, Addison's disease, rheumatoid arthritis, myasthenia gravis, systemic lupus erythematosus and Graves' disease.

Immunosuppressant drugs

The immune system presents a large number of potential molecular targets for therapeutic intervention. Drugs currently used to suppress the immune response tend to be non-specific immunosuppressants with a range of unwanted effects. Immunosuppressant drugs are widely used in many diseases; examples include rheumatoid arthritis, psoriasis and inflammatory bowel disease. Their benefit is achieved both through modulation of the immune system and, in some cases, through their anti-inflammatory properties. Drug such as methotrexate and cyclophosphamide which are used for their cytotoxic actions in cancer chemotherapy (Ch. 52), are also used for their immunosuppressant properties in various disease states. Methotrexate and cyclophosphamide have immunosuppressant properties at doses much lower than those required to treat malignancy (Ch. 52). Corticosteroids (e.g. dexamethasone and prednisone; Ch. 44) are highly effective anti-inflammatory drugs that can be used systemically to suppress type 4 hypersensitivity reactions, autoimmune diseases and graft rejection. They are also used topically for inflammatory skin disease (Ch. 49), inflammatory bowel disease (Ch. 34) and allergic rhinitis (Ch. 39), and by inhalation for asthma (e.g. beclometasone, fluticasone; Ch. 12).

Co-stimulatory molecules on immune cells have recently been targeted in the development of drugs for the treatment of autoimmune disease: belatacept is an antibody that binds to CD80 and CD86 proteins on APCs and rituximab is an antibody that binds to CD20 on B-cells, thus modulating the destructive immune responsiveness against self molecules in disease (Ch. 30, Fig. 30.1).

As an alternative to immunosuppression, inflammatory mediators released during immune reactions can be blocked by antagonists at their receptors on target cells, or by inhibiting their synthesis. This is a useful strategy for management of allergic disorders and some inflammatory conditions. Anti-mediator drugs include histamine H₁ receptor antagonists (antihistamines), cysteinyl-leukotriene receptor antagonists (LTRAs) and cyclo-oxygenase inhibitors (non-steroidal anti-inflammatory drugs, NSAIDs). Antihistamines (e.g. loratadine and cetirizine) are used in the control of hay fever and other allergic disorders (Ch. 39), while oral LTRAs (e.g. montelukast and zafirlukast) are used in asthma (Ch. 12). Oral NSAIDs block the synthesis of prostaglandins and are used in inflammatory soft-tissue disorders and arthritis (Chs 29 and 30).

Calcineurin Inhibitors

Examples

ciclosporin, tacrolimus

Ciclosporin

Mechanism Of Action: Ciclosporin is a fungal cyclic peptide which inhibits T-cell division. It binds in the cell cytoplasm to the protein cyclophilin to form a complex that inhibits calcineurin. Calcineurin is a calmodulin-/Ca²⁺-dependent phosphatase which is a key component in T-cell activation (Fig. 38.1). Activated calcineurin is produced in response to an antigenic signal at T-cell receptors and dephosphorylates nuclear factor of activation in T-cells (NFAT). NFAT then enters the cell nucleus and binds to a promoter region of the IL-2 gene. IL-2 stimulates T-cell division. By inhibiting calcineurin, ciclosporin prevents dephosphorylation of NFAT, which remains in the cytoplasm. This inhibits IL-2 production, and the T-cell division cycle is arrested between G₀ and G₁.

Ciclosporin also inhibits various cellular mitogen-activated protein kinases (such as c-Jun N-terminal kinase and p38 kinases) which are triggered by inflammatory cytokines such as tumour necrosis factor α (TNFα) and IL-1. These protein kinases phosphorylate transcription factors involved in upregulation of c-Fos-mediated gene transcription, a process that is involved in activation of many cell types in response to inflammation.

Ciclosporin also stimulates the production of transforming growth factor β (TGFβ), possibly by releasing an inhibitory effect of calcineurin on TGFβ gene transcription. This may be responsible for some of the nephrotoxicity that occurs with the drug.

Pharmacokinetics: Oral absorption of ciclosporin is variable and incomplete, requiring initial dispersion by bile salts. To overcome this, a microemulsion formulation is used which disperses when it comes into contact with water in the gut so that absorption is independent of bile production. However, different formulations have varying absorption characteristics and switching between formulations should be avoided. Ciclosporin can also be given by intravenous infusion. Ciclosporin selectively concentrates in some tissues, including liver, kidney, several endocrine glands, lymph nodes, spleen and bone marrow. It is extensively metabolised in the liver by the CYP3A4 isoenzyme and has a long half-life (27 h). Monitoring of trough plasma drug concentration has traditionally been used to guide dosage for optimal effectiveness and to minimise toxicity. However, recent evidence suggests that a blood concentration 2 h post-dose may be a better guide to kidney graft survival and minimising toxicity.

Unwanted Effects:

Nephrotoxicity almost always occurs, with a dose-dependent increase in serum creatinine in the first few weeks. The acute effect is due to intrarenal vasoconstriction that may persist and contribute to the less common long-term sequelae, which include interstitial fibrosis and tubular atrophy. Induction of TGFβ may be a contributory factor in the nephrotoxic effects. The decline in renal glomerular function is usually reversible, but permanent renal impairment can result.

Hypertension, often associated with fluid retention, occurs in up to 50% of people, and especially after heart transplantation. It usually responds to standard antihypertensive drug treatment.

Hepatic dysfunction.

Tremor, headache, paraesthesia, fatigue, myalgia.

Hypertrichosis (excessive hair growth) and gum hypertrophy are common.

Gastrointestinal disturbances, including anorexia, nausea and vomiting, abdominal pain.

Hyperlipidaemia, hyperuricaemia, hypomagnesaemia, hypokalaemia.

Drug interactions can be dangerous and caution should be taken when ciclosporin is used with other nephrotoxic drugs, such as aminoglycoside antimicrobials and amphotericin (Ch. 51) or NSAIDs (Ch. 29). Drugs that induce hepatic CYP3A4, such as phenytoin and carbamazepine, can reduce the plasma concentrations of ciclosporin to sub-therapeutic levels. Drugs that inhibit cytochrome P450, such as erythromycin and ketoconazole (Ch. 51), can increase ciclosporin plasma concentration and provoke toxicity.

Tacrolimus

Mechanism Of Action And Effects: Tacrolimus inhibits calcineurin, and therefore T-cell proliferation, by arresting the cell cycle between G₀ and G₁ in a similar manner to ciclosporin. After binding to a receptor protein called FK-binding protein-12, the complex binds to calcineurin and inhibits Ca²⁺-dependent calcineurin activation. Tacrolimus also inhibits c-Jun N-terminal kinase and p38 kinases. Unlike ciclosporin, tacrolimus does not stimulate production of TGFβ.

Pharmacokinetics: Tacrolimus is more water soluble than ciclosporin and undergoes more predictable, though poor, absorption from the gut. It is metabolised by the liver and has a highly variable half-life (4–41 h). Monitoring of the trough blood concentration of tacrolimus is essential for appropriate dose adjustment, especially early in treatment.

Unwanted Effects: These are similar to those of ciclosporin except that tacrolimus causes less hypertension, hirsutism and gum hyperplasia. Effects that are more common with tacrolimus include:

pleural and pericardial effusions,

cardiomyopathy in children, who should be monitored by echocardiography.

Mammalian Target Of Rapamycin Inhibitors

Example

sirolimus

Mechanism of action and effects

Sirolimus (previously known as rapamycin) is a natural fungal fermentation product that inhibits T-cell proliferation by arresting the cell between the G₁ and S phases. It binds to intracellular FK-binding protein-12, and the complex inhibits the action of mammalian target of rapamycin (mTOR), a cytoplasmic kinase. mTOR is a key step in a series of intracellular Ca²⁺-independent events that transduce signals from the cell surface IL-2 receptor and other growth factor receptors to cell-cycle regulators that promote DNA and protein synthesis and mitogenesis. The action of sirolimus therefore differs from that of tacrolimus, despite binding to the same intracellular receptor.

Pharmacokinetics

Sirolimus is rapidly absorbed from the gut and the absorption is modulated by P-glycoproteins. It is metabolised by intestinal and hepatic cytochrome P450 and has a very long half-life (60 h).

Unwanted effects

Oedema, ascites, tachycardia, hypertension.

Abdominal pain, nausea, diarrhoea, stomatitis.

Anaemia, thrombocytopenia, neutropenia.

Hyperlipidaemia, hypokalaemia, hypophosphataemia.

Arthralgia, osteonecrosis.

Lymphocele (a complication of renal transplantation that is more common if sirolimus is used; it can cause ureteric compression).

Rash.

Drug interactions: rifampicin reduces plasma sirolimus concentrations by induction of CYP3A4; the antifungal agents itraconazole and ketoconazole increase plasma concentrations of sirolimus by enzyme inhibition.

Antiproliferative Agents

Examples

azathioprine, cyclophosphamide, mycophenolate mofetil

Azathioprine

Mechanism Of Action: Azathioprine is widely used for immunosuppression. Most of the effects result from cleavage to 6-mercaptopurine in the intestine and liver and then to the active derivative thioinosinic acid, a purine analogue (Ch. 52). The antimetabolite action interferes with purine biosynthesis, thus impairing DNA synthesis in the S-phase of the cell cycle (Figs 52.1 and 52.2), and particularly affects fast-growing cells such as lymphocytes. The drug also blocks CD28 co-stimulation of T-cells. Both cell- and antibody-mediated immune reactions are suppressed (Figs 38.1 and 38.2), with impaired synthesis of immunoglobulins by B-cells, and inhibition of the infiltration and survival of mononuclear cells in inflamed tissue.

Pharmacokinetics: Oral absorption is almost complete. The half-lives of azathioprine and its 6-mercaptopurine metabolite are short (3–5 h). Azathioprine can be given by intravenous injection, but the solution is alkaline and very irritant.

Unwanted Effects:

Dose-dependent bone marrow suppression, especially leucopenia and thrombocytopenia. Regular monitoring of the full blood count (at least every 3 months) is essential.

Hypersensitivity reactions, with malaise, dizziness, vomiting, diarrhoea, fever, myalgia, arthralgia, rash and hypotension. The drug should be stopped immediately if these arise.

Increased susceptibility to infection, often with ‘opportunistic’ organisms.

Alopecia.

There is a small risk of carcinogenicity, especially lymphomas.

Drug interactions: the most important interaction is with allopurinol (Ch. 31). Allopurinol inhibits xanthine oxidase, which is involved in the catabolism of 6-mercaptopurine, and the dose of azathioprine should be reduced by 75% if the drugs are used together.

Cyclophosphamide

Cyclophosphamide is an alkylating drug that is less commonly used as an immunosuppressant. It is discussed in detail in Ch. 52.

Mycophenolate mofetil and mycophenolic acid

Mechanism Of Action And Effects: Mycophenolate mofetil is a prodrug of mycophenolic acid, which reduces purine synthesis by reversible non-competitive inhibition of inosine monophosphate (IMP) dehydrogenase and guanylyl synthase. These enzymes are involved in the conversion of IMP to xanthosine monophosphate, and then to the precursor of guanosine triphosphate that is involved in RNA, DNA and protein synthesis. Inhibition of these enzymes depletes the cell of guanine nucleotides and inhibits cellular DNA synthesis. T-cells, B-cells and monocytes rely on de novo purine nucleotide synthesis, unlike neutrophils and other cells, which can use pre-formed guanine released from the breakdown of pre-formed nucleic acids (the salvage pathway). Mycophenolate therefore is a selective inhibitor of lymphocyte function.

A further action of mycophenolate mofetil is inhibition of smooth muscle proliferation in arterial walls, which may also help to reduce graft rejection that arises from obliterative arteriopathy.

Pharmacokinetics: Mycophenolate mofetil is a prodrug ester which is almost completely absorbed from the gut and hydrolysed rapidly to mycophenolic acid. Elimination of mycophenolic acid is via hepatic metabolism and it has a long half-life (18 h) due to enterohepatic circulation. Mycophenolate mofetil and mycophenolic acid can also be given by intravenous infusion.

Unwanted Effects:

Gastrointestinal upset is very common, including nausea, vomiting, diarrhoea, abdominal cramps and, occasionally, hepatitis or pancreatitis. Tolerance to the gastrointestinal symptoms often occurs.

Hypertension, oedema, tachycardia, chest pain.

Dyspnoea, cough.

Dizziness, insomnia, headache, tremor, seizures.

Bone marrow suppression resulting in leucopenia, thrombocytopenia and anaemia.

Opportunistic infections may be increased, especially with cytomegalovirus, Herpes simplex, Aspergillus and Candida, as well as bacterial urinary tract infection and pneumonia.

Lymphoproliferative disease and skin cancer.

Folic Acid Antagonist

Example

methotrexate

Mechanism of action and uses

Methotrexate inhibits dihydrofolate reductase. This blocks purine and thymidylate synthesis and inhibits the synthesis of DNA, RNA and protein. It is specific for the S-phase of cell division and slows G₁- to S-phase (see Ch. 52). However, in inflammatory disease methotrexate may have a different action by inhibiting enzymes involved in purine metabolism, leading to accumulation of adenosine and its release from cells. Adenosine acts on specific cell surface receptors and suppresses the expression of adhesion molecules on T-cells and neutrophils, inhibiting their accumulation in inflamed tissues, and also reduces cytokine release from a variety of inflammatory cells.

Methotrexate is used by intermittent administration once a week in many conditions such as inflammatory joint disease, Crohn's disease and psoriasis (Chs 30, 34 and 49).

Pharmacokinetics

Methotrexate is well absorbed from the gut but can also be given intravenously, intramuscularly or subcutaneously. It is eliminated by renal excretion, but a small amount may be retained intracellularly for longer periods bound to dihydrofolate reductase and as polyglutamate conjugates.

Unwanted effects

Toxicity to normal rapidly dividing tissues (especially the bone marrow).

Hepatotoxicity can follow chronic therapy (as in psoriasis).

Toxicity is increased in the presence of reduced renal excretion and methotrexate should be avoided if there is significant renal impairment. Folic acid is frequently taken after methotrexate to reduce mucositis and myelosuppression. NSAIDs such as aspirin can reduce the renal excretion of methotrexate and increase its toxicity.

Interleukin-2 Receptor Antibodies

Examples

basiliximab

Basiliximab is a chimaeric monoclonal antibody with murine sequences in the hypervariable region. It binds to the IL-2 receptor on activated T-cells and prevents T-cell proliferation. It is used for initial induction therapy prior to transplantation or for treatment of acute transplant rejection.

Pharmacokinetics

Basiliximab is given by intravenous infusion immediately before and again 4 days after surgery. It has a very long half-life of over 1 week.

Unwanted effects

Hypersensitivity reactions occur rarely.

Selective Co-Stimulation Blocker

Examples

belatacept

Belatacept is a fusion protein that binds to CD80 and CD86 molecules on APCs and blocks their co-stimulatory action with CD28 on T-cell activation. It is used to prevent rejection in adults undergoing renal transplantation who are seropositive for the Ebstein–Barr virus.

Pharmacokinetics

Belatacept is given by intravenous infusion and has a very long half-life (8–10 days).

Unwanted effects

Hypersensitivity reactions occur rarely.

lymphoproliferative disorder, especially in those with no prior exposure to Ebstein-Barr virus.

Immunosuppression in organ transplantation

Immunosuppressant drugs block rejection at the steps of T-cell activation, T-cell proliferation and cytokine production (Figs 38.1 and 38.2). A major direction of current research is to find regimens that will induce immune tolerance and allow eventual withdrawal of immunosuppressant drugs.

Effective immunosuppression has improved the early survival of kidney, liver, heart, heart–lung, intestinal and haematopoietic stem cell transplants. However, suppression of acute rejection is more effective than prevention of chronic rejection, which responds poorly to immunosuppressant therapy. Regimens for immunosuppression vary among transplant units and according to the immunogenicity of the transplanted tissue. Combination therapy with a corticosteroid, calcineurin inhibitor and an antiproliferative agent is commonly used.

For kidney transplantation, induction therapy is given at the time of transplantation with either anti-CD3, which is inhibits T-cell activation in the thymus (anti-thymocyte immunoglobulin), or an IL-2 receptor antibody (basiliximab) to reduce initial acute rejection. Immunosuppression is then maintained with oral agents such as the corticosteroid prednisolone (Ch. 44) together with ciclosporin or tacrolimus, or with sirolimus or mycophenolate mofetil if the calcineurin inhibitors are poorly tolerated. Some units add azathioprine to this regimen. With such regimens, 90% of cadaveric kidney grafts will survive beyond 1 year. Only half of those that fail are lost due to rejection, and the rest from thrombosis of the graft blood supply. Progressive graft loss continues after the first year, with only 67% of grafts from ‘brain death donors’ surviving at 10 years. Grafts from living donors have better survival rates of 96% at 1 year and 78% at 10 years. Most of the late graft losses are as a result of chronic vascular rejection. If this occurs, increasing the dosages of the primary immunosuppressant drugs may help; however, there is continuing uncertainty about whether chronic rejection reflects nephrotoxicity from long-term use of calcineurin inhibitors or inadequate immunosuppression. It is possible that drugs such as sirolimus or mycophenolate mofetil may reduce the incidence of chronic rejection, but there are few long-term data on outcome. Late acute rejection is a less common problem. It can sometimes be overcome by high-dose corticosteroid or the use of polyclonal antilymphocytic globulin or monoclonal antilymphocytic antibody (although the use of these is associated with an increased risk of opportunistic infection and long-term malignancy). Tacrolimus or mycophenolate mofetil can also be used successfully as a rescue treatment during late episodes of acute rejection.

In contrast to renal transplants, pancreatic transplants are more immunogenic and quadruple immunosuppressant regimens are widely used. Induction treatment with antilymphocytic globulin is then followed by ciclosporin, azathioprine and a corticosteroid. Mycophenolate mofetil is sometimes substituted for azathioprine, or tacrolimus for ciclosporin. Despite these treatments, 5-year prancreatic graft survival is only about 60% and the risk of post-transplant infection is high.

Triple immunosuppressant therapy is used for heart (50% 10-year survival), heart–lung (30% 10-year survival), liver (70% 10-year survival) and intestinal (40–50% 3-year survival) transplants, often with initial use of an IL-2 receptor antibody (basiliximab). In addition to prednisolone, tacrolimus is often included in these regimens in place of ciclosporin. Mycophenolate mofetil as an alternative may reduce acute rejection rates but there is less evidence for an impact on chronic rejection.

With haematopoietic stem cell transplantation, graft-versus-host disease (GVHD) is the major barrier. This usually begins at least 3 months after the transplant and has three phases. The first phase involves damage to intestinal mucosa and the liver, with activation of host cells and release of inflammatory cytokines. These cytokines upregulate MHC proteins on the host cells, which are then recognised by the donor T-cells. The second phase involves activation and proliferation of donor T-cells, and the third phase includes tissue destruction by monocytes primed by inflammatory cytokines and lipopolysaccharide from T-cells and damaged intestinal mucosa. GVHD can be prevented by inhibition of phase 1, using a calcineurin inhibitor such as ciclosporin or tacrolimus, or possibly mycophenolate mofetil. Acute GVHD can be treated by a corticosteroid with ciclosporin.

Immunosuppression in other disorders

Immunosuppressant therapy is used for several diseases in which autoimmunity may contribute to the pathogenesis. These include many connective tissue diseases such as vasculitis and systemic lupus erythematosus, inflammatory arthritis, polymyalgia rheumatic, certain types of glomerulonephritis, autoimmune hepatitis, psoriasis, inflammatory bowel disease and some haematological disorders. Immunosuppressant drugs may be given alone or in combination. Those most widely used include corticosteroids, azathioprine, methotrexate and cyclophosphamide. Ciclosporin, tacrolimus and mycophenolate mofetil have also been used in disorders such as asthma, inflammatory bowel disease and psoriasis, with some success.

Self-Assessment

True/false questions

1. Immunosuppression requires higher doses of methotrexate than used for cancer chemotherapy.

2. Ciclosporin and tacrolimus reduce interleukin (IL)-2 gene transcription in lymphocytes.

3. Ciclosporin causes bone marrow suppression.

4. Careful assessment of renal function is required with ciclosporin administration.

5. Azathioprine suppresses antibody-mediated immune responses.

6. Azathioprine interacts with allopurinol.

7. Sirolimus and tacrolimus share a common mechanism of action.

8. Corticosteroids have a narrow spectrum of immunosuppressant activity.

9. Mycophenolate is an alternative to azathioprine for preventing acute rejection.

10. Basiliximab is an antibody that blocks the IL-2 receptor.

Case-based questions

A 35-year-old woman was about to receive her second kidney transplant. The previous transplant had lasted 5 years but, despite immunosuppression with prednisolone and ciclosporin, it was eventually rejected.

A How might the chances of acute rejection of the second transplant be reduced?

B What could be the long-term risks of combination chemotherapy with corticosteroids, tacrolimus and azathioprine?

True/false answers

1. False. Immunosuppressant doses of drugs such as methotrexate, cyclophosphamide and azathioprine are lower than those used for cancer chemotherapy.

2. True. These calcineurin inhibitors reduce the transcriptional effects of NFAT on IL-2; the loss of IL-2 suppresses T-cell maturation and proliferation.

3. False. The calcineurin inhibitors are selective suppressors of lymphocyte proliferation.

4. True. Ciclosporin is nephrotoxic and renal monitoring is necessary.

5. True. Azathioprine has a cytotoxic action by inhibiting purine metabolism and the proliferation of lymphocytes and other immunocompetent cells is inhibited.

6. True. Allopurinol inhibits xanthine oxidase, which is involved in deactivating metabolites of azathioprine; doses of azathioprine should be reduced when given with allopurinol.

7. False. Sirolimus (rapamycin) binds to FK-binding protein-12, but the complex inhibits mammalian target of rapamycin (mTOR), a protein kinase involved in IL-2 signalling, unlike the calcineurin inhibitors that reduce IL-2 gene transcription.

8. False. Corticosteroids modulate the transcription of hundreds of immune and inflammatory genes encoding cytokines, mediators, adhesion molecules and apoptotic proteins.

9. True. Mycophenolate may have fewer toxic effects than azathioprine and is increasingly used in preventing acute rejection, but its place in preventing chronic rejection is less clear.

10. True. Basiliximab is a chimaeric monoclonal antibody that blocks the IL-2 receptor.

Case-based answers

A Basiliximab given before and 4 days after renal transplant surgery reduces acute rejection by 35%. This is maintained typically with oral combination therapy of a corticosteroid (usually prednisolone) with a calcineurin or mTOR inhibitor (such as ciclosporin, tacrolimus or sirolimus), and an antiproliferative immunosuppressant (such as azathioprine or mycophenolate). When used in combination, lower doses of the drugs can be administered than when giving the drugs alone. Intensive monitoring of liver and renal functions is important.

B Over-suppression of the immune response brings problems of opportunistic infections. Additional ‘steroid effects’ as described for iatrogenic Cushing-like syndrome may be also apparent (Ch. 44).

Compendium: immunosuppressant drugs

For the immunosuppressant activity of corticosteroids, see Ch. 44. See also the inflammatory arthritides (Ch. 30) and chemotherapy of malignancies (Ch. 52).