Immune Modulation to Prevent Rejection PDF

Immune modulation to prevent rejection through alloreaction - immunosuppression TU867 4 Immunosuppression A majority of patients receive organs mismatched at one or more HLA loci Administration of these drugs, which is greatest pre- and post- transplantation renders patie...

Immune modulation to prevent rejection through alloreaction - immunosuppression TU867 4 Immunosuppression A majority of patients receive organs mismatched at one or more HLA loci Administration of these drugs, which is greatest pre- and post- transplantation renders patients highly susceptible to infection The dose of drugs is gradually reduced to ‘maintenance levels’ that prevent acute rejection while sustaining active defences against infection – the likelihood of chronic rejection increases There are side-effects, they vary for different drugs. The drugs are used in combinations that increase their immunosuppressive effects but not their toxic effects There is a higher risk (x 3) of certain types of malignant disease for patients on these drugs for long periods – e.g. carcinomas of the skin and genital tract, lymphoma and Kaposi’s sarcoma Immunosuppressive drugs 3 classes Those that non-specifically deplete the majority of lymphocytes and monocytes and inhibit the responsiveness of those that remain (Abs and corticosteroids – given prior to transplantation) Those that specifically interfere with the 3 signals needed for T cell activation The signal from the TCR (TCR/CD3) – NFKB and NFAT The co-stimulatory signal (B7/CD28) The signal generated by IL-2 binding to it’s receptor Those that have their effect after activated naïve T cells have begun to proliferate (cytotoxic drugs) Class 1 of immunosuppressive drugs Abs that react broadly with WBCs are given prior to and post transplantation to deplete these cells One example is rabbit antithymocyte globulin (rATG) – a polyclonal mixture of high affinity Abs that binds T, B, NK, dendritic and endothelial cells – fix complement and act as opsonins Another example is humanised rat mIgG that is specific for CD52 (alemtuzumab) – CD52 is expressed on almost all lymphocytes, monocytes and macrophages – results in a profound, long-lasting lymphopenia CD52 – function unknown The cell surface complex of CD52 and anti-CD52 is unusually efficient at fixing complement CD52 is a tiny protein of 49aas (8kDa) that is tethered to cell membranes by a glycophosphatidylinositol tail Abs that bind CD52 are very close to the cell membrane increasing the likelihood that C3b will bind covalently to the leucocyte surface Corticosteroids – most effective when administered prior to transplantation Hydrocortisone (cortisol), a steroid made by the adrenal cortex, has been used clinically for more than 50 years to reduce inflammation Prednisone, a synthetic derivative of hydrocortisone, is a pro-drug, and when enzymatically converted into prednisolone is about 4 times more potent in reducing inflammation Alone it is insufficiently immunosuppressive to prevent graft rejection but works well in combination with a cytotoxic drug  Corticosteroids do not act on cell-surface receptors but diffuse across the plasma membrane and bind cytoplasmic receptors A key immunosuppressive effect of hydrocortisone and prednisolone is to prevent the action of NFkB – they increase the production of IkBa (inhibitor)  DAG activates PKCq which leads to the assembly of a membrane-bound complex that includes - CARMA1 Adaptor proteins - BCL-10 - MALT1 This complex activates a multiprotein enzyme complex, inhibitor of kB kinase (IKK) IKK phosphorylates the inhibitor of kB (IkB) IkB normally binds NF-kB, keeping it in the cytosol, but after the activity of IKK, NF-kB is released and migrates to the nucleus resulting in gene transcription – e.g. IL-2 (early gene)  The transcription of approx. 1% a cell’s genes can be influenced by corticosteroids There is a decrease in inflammatory cytokine production, e.g., IL-1 Prednisolone alters lymphocyte homing, lymphocytes are barred from entering SLT, instead they congregate in the BM Adverse side-effects include fluid retention, weight gain, diabetes, loss of bone mineral and thinning of the skin – prolonged use avoided Class 2 of immunosuppressive drugs Blocking signals from the TCR – NFAT activation by calcineurin Introduced at the end of the 1970s – marked impact on clinical transplantation in the ‘80s and ‘90s – this period became known as the ‘cyclosporin era’ as cyclosporin was the first of these drugs to be introduced Cyclosporin A (cyclosporine) - a cyclic decapeptide derived from the soil fungus Tolypocladium inflatum – inhibits T cell activation by disrupting transduction signals from the TCR Tacrolimus (FK506) isolated from Streptomyces tsukabaensis (soil actinomycete) – a macrolide – structurally distinct from cyclosporin A but works through a similar mechanism  Cyclophilins (bind Cyclosporin A) and FK- binding proteins (bind FK506/tacrolimus) are distinct peptidyl-prolyl isomerases and are known collectively as immunophilins. The complexes created bind calcineurin, preventing it from activating NFAT – IL-2 is never made  As these drugs do not target proliferating cells reduced haematopoiesis and intestinal damage do not occur. Nephrotoxicity may occur – patients become sensitised to the drug, can no longer tolerate it Antibodies specific for CD3 were used to prevent and control acute rejection In 1986 the CD3-specific mouse Lymphocytes mAb OKT3 was the first mAb surrounding an licensed for clinical use arteriole in a kidney undergoing rejection After transplantation patients Lymphocytes are maintained on a surrounding the renal combination of tubules of the same immunosuppressive drugs that, kidney because of their cytotoxicity and the immunodeficiency they cause, are gradually reduced in Staining of CD3 in the dosage. Sometimes early same section symptoms of rejection appear  Treatment involved a 5 – 15 day course of daily injections of mouse anti-CD3 Ab combined with prednisolone Anti-CD3 caused the TCRs to be internalised and therefore they were unable to recognise Ag Attempts to convert therapeutic mouse anti-CD3 Abs into either chimeric or humanised forms were unsuccessful (see next slide) OKT3 was taken off the market in 2008 – anti-CD52 and rATG became the preferred reagents Immune complexes are deposited in small blood vessels – activate complement and Skin phagocytes inducing fever haemorrhage and E.g. mouse Serum sickness, an monoclonal – extreme form of 1st exposure – type III primary IR hypersensitivity reaction Urticarial rash Symptoms abate once Ag cleared Blocking the co-stimulatory Mutations here increase signal avidity for B7 Belatacept – generic name for drug approved for use in transplantation in 2011 Mutations here eliminate Fc- mediated effector A soluble, synthetic chimeric protein that function combines the extracellular domain of CTLA-4 with the hinge and Fc domains of the IgG1 heavy chain Acts to prevent alloreactive T cells receiving their co-stimulatory signal from activated DCs expressing B7 and presenting alloAg  Works as well as the best of the older, established drugs and is better at preserving kidney function It is associated with an increased incidence of episodes of acute rejection Blocking the IL-2 signal IL-2 acts in both an autocrine and paracrine fashion by binding to the IL-2 receptor expressed by T cells Naïve alloreactive T cells express a low-affinity receptor for IL-2 (consists of b and g chains – next slide) On recognition of alloAg the a chain (CD25) is synthesised - high affinity receptor Chimeric basiliximab and humanised daclizumab are IgG mAbs specific for CD25 These Abs are first given just before the transplant is performed. Subsequent infusions are given during the first 2 months after transplantation Benefit is that this treatment targets T cells embarking on activation - does not induce significant immunodeficiency Blocking the IL-2 signal IL-2 acts in both an autocrine and paracrine fashion by binding to the IL-2 receptor expressed by T cells Naïve alloreactive T cells express a low-affinity receptor for IL-2 (consists of b and g chains – next slide) On recognition of alloAg the a chain (CD25) is synthesised - high affinity receptor Chimeric basiliximab and humanised daclizumab are IgG mAbs specific for CD25 These Abs are first given just before the transplant is performed. Subsequent infusions are given during the first 2 months after transplantation Benefit is that this treatment targets T cells embarking on activation - do not induce significant immunodeficiency  Rapamycin (sirolimus) – Streptomyces hygroscpicus a soil bacterium found on Easter Island (Rapa ui) – binds FK-binding proteins (like Tacrolimus) but prevents T cell activation by inhibiting signal transduction from the IL-2R - the complex physically interacts with the protein “mammalian target of rapamycin” (mTOR). Also induces Treg development. Impedes progression through the G1/S transition of the proliferation cycle Inhibits the p70 S6 kinase (p70s6k) – decreases protein synthesis inhibits the enzymatic activity of the cyclin-dependent kinase cdk2-cyclin E complex, which functions as a crucial regulator of G1/S transition mTOR acts upstream of p70s6k and cdk2-cyclin E More toxic than cyclosporin A or tacrolimus but useful in combination therapy Class 3 of immunosuppressive drugs Cytotoxic drugs kill alloantigen-activated proliferating T cells – usually administered after transplantation Azathioprine (much used in kidney transplantation) – a pro-drug – eventually converted to 6-thioinosinic acid – inhibits the production of inosinic acid, an intermediate in the production of adenine and guanine – inhibits DNA replication. Like all cytotoxic drugs it damages all tissues normally active in cell division e.g BM, intestinal epithelium, hair follicles – anaemia, leukopenia, thrombocytopenia, intestinal damage and hair loss. Toxic to the liver. It is administered after transplantation Mycophenalate mofetil – product of Penicillium stoloniferum - metabolised in the liver to mycophenolic acid – inhibits inosine monophosphate dehydrogenase, necessary for guanine synthesis  Cyclophosphamide – N2 mustard compound developed as a chemical weapon, used heavily during WWI. A pro-drug converted to phosphoramide mustard – alkylates and cross-links DNA molecules – affects normal cell division and transcription – equally immunosuppressive pre- and post – Ag stimulation. In addition to side- effects shared with other cytotoxic drugs it specifically damages the bladder (cancer or haemorrhagic cystitis). Not particularly toxic to the liver – useful alternative – used in short courses Methotrexate – effective in cancer treatment – prevents DNA replication by inhibiting dihydrofolate reductase – essential for the synthesis of thymidine – drug of choice for inhibiting GVHD in BM transplant recipients The chemical structures of cytotoxic drugs Rapamycin