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Transplantation Immunology

Transplant Immunology

1. Phases of Immunosuppression

A. Early rejection prophylaxis
B. Maintenance rejection prophylaxis
C. Treatment of established rejection

2. Mechanism of Action of Immunosuppressive Agents

A. Inhibitors of Interleukin -2
B. Production
1) Cyclosporine A
2) Tacrolimus
C. Action
1) Rapamycin (Sirolimus)
2) SDZ RAD
3) Interleukin-2 Receptor Blockers
D. Daclizumab
E. Basiliximab
F. Inhibitors of purine or pyrimidine biosynthesis
G. Purine
1) Azathioprine
2) Methotrexate
3) Mycophenolate mofetil
4) Mizoribine (bredinin)
H. Pyrimidine
1) Brequinar sodium
2) Leflunomide
I. Both purine and pyrimidine
1) Cyclophosphamide
J. Opsonization of lymphocytes
1) Murine monoclonal anti-CD-3 antibody (OKT3)
2) Polyclonal antibodies (horse, rabbit)
I. Multiple mechanisms or not clearly defined mechanisms
1) Adrenocorticosteroids
2) 15-Deoxyspergualin

3. Murine Monoclonal CD-3 Antibody (OKT3)

A. Identification: IgG2a Murine Immunoglobulin
B. Mechanism: Inhibits signal transduction of antigen recognition, opsonizes CD-3 lymphocytes
C. Dose/route: 5-10 mg/day, IV
D. Side effects: First dose reactions, HAMA formation
E. Interactions: None
F. Use: Early rejection prophylaxis, treatment of rejection
G. Monitoring: CD-3 Counts, OKT3 levels

4. Polyclonal Antibodies

A. Identification: Horse (ATGAM) or rabbit (Thymoglobulin) immunoglobulin
B. Mechanism: RES-mediated removal of opsonized cells
C. Dose/route: ATGAM 10-20 mg/kg/day IV; Thymoglobulin 1.5mg/kg IV
D. Side effects: Leukopenia, thrombocytopenia, fever, arthralgias, serum sickness
E. Interactions: None
F. Use: Early rejection prophylaxis, treatment of rejection
G. Monitoring: CD-2 counts

5. Cyclosporine

A. Identification: Metabolite of tolypocladium inflatum gams
B. Mechanism: Inhibits m-RNA transcription of interleukin-2
C. Dose/route: 3-6 mg/kg/day orally; IV:Oral = 1:3
D. Side effects: Nephrotoxicity, hypertension, tremor, headache/paresthesias, hirsutism, gingival hyperplasia
E. Interactions: Increase clearance of cyclosporine
1) Rifampin
2) Isoniazid
3) Phenytoin
4) Phenobarbital
F. Decrease clearance of cyclosporine
1) Erythromycin
2) Ketoconazole
3) Diltiazem
4) Verapamil
5) Nicardipine
6) Cimetidine
7) Use: Maintenance immunosuppression
8) Monitoring: Blood or serum level determination
Cyclosporine Formulations
1. Sandimmune LiquidLiquid & Capsules
2. Neoral (microemulsion) LiquidLiquid & Capsules
3. Sang CYA (microemulsion) LiquidLiquid

6. Tacrolimus (FK-506)

A. Identification: Fermentation product of Streptomyces tsukubaenis
B. Mechanism: Inhibits mRNA transcription of interleukin-2
C. Dose/route: 0.05 - 0.075 mg/kg orally q 12 hours 0.03 mg/kg intravenously q 24 hours
D. Side-effects:
1) Nephrotoxicity
2) Hyperglycemia
3) Neurotoxicity
4) Hypertension
E. Interactions: Believed similar to cyclosporine
F. Use: Maintenance immunosuppression
G. Monitoring: Blood level determination

7. Azathioprine

A. Identification: Precursor to 6 mercaptopurine
B. Mechanism: Disrupts normal purine incorporation into ribonucleic acids
C. Dose/route: 1 - 4 mg/kg/day; IV:Oral = 1:1
D. Side effects: Hematologic, pancreatitis, cholestatic jaundice, hepatitis, interstitial pneumonitis
E. Interactions: Increased levels with allopurinol
F. Use: Maintenance immunosuppression
G. Monitoring: White blood cell count

8. Mycophenolate Mofetil (RS-61443)

A. Identification: Morpholinoethylester of mycophenolic acid, a fermentation product ofPenicillium species
B. Mechanism: Inhibits inosine monophostate dehydrogenase in the de novo pathway of guanine nucleotide biosynthesis
C. Dose/route: 1,000 - 1,500 mg orally q 12 hours
D. Side-effects: Leukopenia, Nausea, vomiting, diarrhea
E. Interactions: Probably with acyclovir
F. Use: Maintenance immunosuppression
G. Monitoring: None

9. Corticosteroids (Prednisone, hydrocortisone, methylprednisolone)

A. Mechanism:
1) Inhibit transcription of IL-1 and IL-6 encoding m-RNA in macrophages
2) Block antigen recognition, decrease IL-1 AND IL-6 driven effects
3) Redistribution of lymphocytes
B. Dose/route: Prednisone 1 mg = hydrocortisone 4 mg = methylprednisolone 0.8 mg
C. Side effects:
1) Cushing's syndrome, osteoporosis, myopathy, cataracts, peptic ulcers
2) Glucose intolerance, hypercholesterolemia, skin fragility, adrenal suppression
D. Interactions: None clinically significant
E. Use: Maintenance immunosuppression, rejection treatment

10. Immunosuppression: Early Rejection Prophylaxis

A. Standard Triple therapy
B. Preoperative
1) Cyclosporine: 2-6 mg/kg po based on renal function
2) Azathioprine: 4 mg/kg IV
C. Intraoperative
1) Methylprednisolone: 500 mg
D. Postoperative
1) Cyclosporine: 2-6 mg/kg po bid based on trough levels and renal function
2) Azathioprine: 2 mg/kg/day
3) Methylprednisolone: 125 mg IV every 8 hours for 3-4 doses, followed by prednisone
4) Prednisone: (beginning after Methylprednisolone)1 mg/kg/day tapering over 1 week to 0.5 mg/kg/day, followed by further tapering over 2-3 months to 0.2-0.3 mg/kg/day
E. Quadruple Therapy- OKT3 *
F. Preoperative
1) Cyclosporine: None
2) Azathioprine: 4 mg/kg IV
G. Intraoperative
1) Methylprednisolone: 500 mg
2) OKT3: 5-10 mg (or administer first dose of OKT3, 5 mg IV 24-48 hours postoperatively)
H. Post operative
1) OKT3: 5 mg/day IV for 7-10 days post operative
2) Cyclosporine: Beginning on the fourth post operative day, 2-6 mg/kg po bid based on trough levels and renal function
3) Azathioprine: 2 mg/kg/day
4) Methylprednisolone: 25 mg IV every 8 hours for 3-4 doses, followed by prednisone
5) Prednisone: (beginning after Methylprednisolone)0.25 mg/kg/day during the time of OKT3 administration. After OKT3 course completed, increase to 1 mg/kg/day for 7 days, then taper either completely off over 4 weeks or to 0.2-0.3 mg/kg/day by 1-3 months.
I. * OKT3 should be premedicated daily for three days with diphenhydramine 50 mg IV, acetaminophen 650 mg po or per rectum, and ranitidine 100 mg IV. OKT3 should be post-medicated every 6, 12, and 18 hours after the first 3 doses with diphenhydramine 25 mg IV, acetaminophen 650 mg po or per rectum, and ranitidine 50 mg IV.
J. Quadruple Therapy - ATG/ALG/ALS**
K. Preoperative
1) Cyclosporine: None
2) Azathioprine: 4 mg/kg IV
L. Intraoperative
1) Methylprednisolone: 500 mg
M. Post operative
1) ATG/ALG/ALS: Daily dosing for 7-10 days, Dose depends on preparation
2) Cyclosporine: Beginning on the second or third post-operative day, 2 - 6 mg/kg po bid based on trough levels and renal function
3) Azathioprine: 2 mg/kg/day
4) Methylprednisolone: 125 mg IV every 8 hours for 3-4 doses, followed by prednisone
5) Prednisone: (beginning after Methylprednisolone) 0.25mg/kg/day during the time of ATG/ALG/ALS, followed by 1mg/kg/day for 7 days, then taper either completely off over 4 weeks or to 0.2-0.3 mg/kg/day by 1-3 months.
N. ** ATG/ALG/ALS should be pre-medicated daily with diphenhydramine 25-50 mg IV and acetaminophen 650 mg po or per rectum

11. Maintenance Immunosuppression Goal

A. Lowest overall level of immunosuppression to prevent rejection
B. Cyclosporine levels
1) Low therapeutic after 1-2 years
C. Azathioprine
1) 1-2 mg/kg/day after 1-2 years
D. Prednisone
1) 0 - 0.1 mg/kg/day after 1 year

12. Treatment of Rejection - Considerations

A. Histologic grade of biopsy
B. Allograft function
C. Time after transplantation
D. Past rejection history
E. Concomitant immunosuppression
F. Optimize cyclosporine/azathioprine

13. TREATMENT OF REJECTION

GRADEMild
ModerateNone or oral corticosteroid augmentation
ModerateOral corticosteroid augmentation or IV corticosteroids
Severe TREATMENTIV corticosteroids and ATG/ALG OR OKT3

14. Other options

A. Alteration of maintenance regimen
1) Change from cyclosporine to Tacrolimus
2) Change from azathioprine to mycophenolate mofetil
3) Change from azathioprine to cyclophosphamide (vascular rejection)
B. Methotrexate course (2.5 - 7.5 mg. Q 12 hrs x 3 doses/week for 8-12 weeks)
C. Plasmapheresis (vascular rejection)
D. Total lymphoid irradiation
E. Photophoresis
F. Re-transplantation

EXTENDED OUTLINE

A. Major Histocompatability Complex (MHC)-prime physiologic role is to recognize “self” from “nonself”; in humans, this is known as the HLA system
B. HLA: class I—HLA-A, B, C; expressed on all cells of an organism. Class I molecules present antigenic peptides to activated T lymphocytes expressing CD8 phenotype
C. class II—DP, DQ, DR; expressed on antigen presenting cells, e.g., B cells, T cells, macrophages, dendritic cells, and endothelium. Present to T lymphocytes expressing the CD4 phenotype.
D. Pivotal cells moderating rejection are the T cells expressing the CD4 complex. These T cells recognize foreign Class II antigens on antigen presenting cells (APCs)—these cells not only present, but also provide signals (lymphokines/adhesion molecules) for T cell activation (second signal). There are two pathways for this to occur—direct and indirect routes of sensitization
E. Activated CD4 cells are divided into Th1and Th2 populations: Th1 subpopulation produces: IL-2 (CD8 differentiation), INF (MHC class II differentiation), TNF (NO radicals/O2/Prostaglandins) Th2: IL-4,5,10—augments B cell mediated responses
G. Effectors of Graft Rejection:
1) CD8 activation is thought to involve recognition of class I antigen (first signal) in a setting of increased levels of IL-2 (second signal) secreted by activated CD4 cells. Graft destruction ensues.
2) Hyperacute rejection is secondary to pre-exisiting blood group antibodies, anti-MHC antibodies, or natural antibodies which react with the endothelial antigens—complement, coagulation, and kallikrein/bradykinin cascades activated. Leads to graft edema, hemorrhage, and vascular thrombosis.
3) Accelerated rejection from IgM/IgG antibodies formed in response to the donor graft. Biopsy shows vascular destruction with a paucity of cellular infiltrate.
H. Hallmark of cellular rejection is graft infiltration:
1) leukocyte attachment to the endothelium
a) mediated by cell adhesion molecules: selectins (rolling effect), integrins (bind the attached molecules), immunoglobin superfamily-related molecules. This is followed by diapedesis—ICAM-1 and LFA-1 interaction
2) transmigration through the vessel wall
3) migration within the graft
4) selective retention of activated cells in the graft
5) local proliferation of cells

1. Rejection Prevention

A. MHC matching
B. Immunosuppression
1) Cyclosporin (CyA) and FK506—inhibit lymphocyte proliferation and lymphokine production by binding to cytosolic intracellular receptors known as immunophilins (CyA-cyclophilins/FK506-FK506 proteins). These complexes inhibit calcineurin an intracellular protein phosphatase which plays a crucial role in the induction of lymphokine genes (IL-2). Side effects: renal dysfunction, GI, CNS, hypertension, and diabetes
2) Corticosteroids—negatively affecting the release of IL-1 and IL-6 from macrophages and thereby inhibiting IL-2 release. Side effects include hypertension, diabetes, cushingoid features, poor wound healing and asceptic bone necrosis
3) Azathioprine works non specifically by virtue of its antimetabolite effects to inhibit lymphocyte proliferation
4) OKT3—mouse monoclonal antibody against T cell receptor CD3 which nonspecifically suppresses all T cell functions. Use is generally in acute rejection episodes. Side effects: cytokine release causing fever, chills, and pulmonary edema; antibody production against the maurine antibody which precludes future courses; dramatic increase in lymphoproliferative disorders.
5) Rapamycin—homolog of FK506, but does not inhibit calcineurin. Mode of action is unclear. Has prevented development of cardiac allograft vasculopathy in rat allografts
6) 15-Deoxyspergualin (DSG)—binds cytoplasmic protein Hsc70 and interferes with antigen presentation and T and B cell development. Good for pancreatic islet cell survival. Causes myelosuppression
7) Mycophenolate mofetiln—inhibits inosine monophosphate dehydrogenase which blocks the de novo pathway for purine synthesis. This pathway is crucial for the proliferative response of T and B cell response. There is a low side effect profile.
8) Brequinar inhibits dihydrooratate dehydrogenase and blocks the de novo synthesis of pyrimidines. The proliferative response is attenuated.
C. Induction therapy
1) its use is associated with a greater cumulative rejection frequency
2) does not delay the onset of first rejection
3) does not reduce the cumulative number of episodes of rejection
D. Tolerance
1) refers to the elimination of the immune response to the antigens of the transplant while the immune response to all other antigens remains intact
2) Anergy—inactivation of cells reactive to the foreign antigen; thought to be the result of T cells binding specific antigen, but not receiving the appropriate second signal from APCs or CD4 cells. IL-2 experimentally has been shown to reverse this
3) Clonal deletion—elimination of cells reactive to the foreign atigen; occurs primarily in the thymus by a process known as negative selection
4) Suppression—suppression of cells responsive to the foreign antigens by another, regulatory immulogic process. Veto cell—inhibits the activity of T cells reactive with antigens on its surface thereby suppressing the activity of the attacking cells

2. Chronic Rejection

A. Cardiac allograft vasculopathy (CAV)
1) is now the leading cause of death or graft failure after the first year.
a) manifested by diffuse and accelerated form of coronary arteriosclerosis—often involves the full length of the artery.
2) virtually all transplant recipients have these findings.
3) rapidly progresses to vessel occlusion and MI
a) pathologic finding is a diffuse intimal thickening and perivascular inflammation extending from large epicardial arteries into medium sized arteries and arterioles
b) the endothelial response to injury theory likely forms the common bond; stimulated endothelial and smoth muscle cells produce cytokines and growth factors causing cell proliferation and smooth muscle and macrophage migration to the intima resulting in concentric lipid-laden calcium-poor plaque. There is evidence to document an inflammatory stage prior to the smooth muscle cell proliferation and also an impairment of endothelial-derived relaxation factor.
c) immune mechanisms are probably at work because the vasculopathy is selective for the allograft which it effects diffusely; the cause of the presumed endothelial injury is unknown
d)Risk factors??—lipid levels, hypertension, smoking, diabetes, and a history of previous atherosclerosis have not correlated with an increased risk of CAV. Only CMV infection has shown a strong association with either death or retransplantation from CAV.
4) use of dobutamine stress echocardiography to follow vs. angiography
a) best addressed by repeat transplantation although this is associated with a 30% or greater lower rate of survival
B. Xenotransplantation
1) widespread preformed antibodies in humans which are reactive for antigens of other species—e.g. pig to human transplant results in hyperacute rejection (discordant) [Concordant rejection is when closely related species reject transplants in a manner similar to allograft rejection]
2) cells and organs from one species may not be able to function in a xenogenic environment
3) cell mediated xenografic rejection may differ from allogeneic rejection and thus require different immunosuppression
4) the future may lie in manipulating the donor organ endothelial system expression of complement inhibitory proteins and therefore mediate hyperacute rejection by preventing complement activation.