History of immunosuppressants by cGYCwrJ

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									    History of
immunosuppressants

        Dr.
1st successful kidney transplant

Dec 23, 1954
– Peter Bent Brigham
  Hospital, Boston MA
– Dr. Joseph Murray
23 yr old identical twin
brothers
– No immunosuppression
  used
– d. 1962 due to disease
  recurrence
  (glomerulonephritis)



                           N Engl J Med 2004;351:2761-6
History of immunosuppression in
         transplantation
Goals of immunosuppression (IMS)
 To prevent allograft rejection
 To prolong allograft functional life
 To optimise allograft function

 Prolong patient survival
 Improve patient quality of life
 To minimize toxicity of IMS agents
 – immunodeficiency complications
 – non-immune toxicities
Transplantation and the immune
            system
Recognition of allograft as non-self
Donor and recipient antigen presenting
cells
(APC’s = dendritic cells, macrophages)
– Present donor antigen to host T-lymphocyte



         Key event:= T-lymphocyte activation
         Immunosuppression:
          3 ways to achieve
Depletion of lymphocytes
– Polyclonal antibodies (horse, rabbit anti-thymocyte
  globulin)
– Mouse monoclonal anti-CD3 antibody (OKT3)
– Humanized monoclonal anti-CD52 (alemtuzumab)
– B-lymphocyte monoclonal anti-CD20 (rituximab)


Diversion of lymphocyte traffic
Blocking of lymphocyte response

                                       NEJM 2004;351:2715-29.
                           Calcineurin inhibitors Anti-IL2 receptor Antibodies
                                     Ciclosporin                   Basiliximab
    Corticosteroids                  Tacrolimus                    Daclizumab



                                     A
                                     C
                                     T
 Antigen                Helper       I              Activated
presenting
   cell
                          T
                      Lymphocyte
                                     V
                                     A
                                                        T
                                                   Lymphocyte
                                                                                      IL-2
                                                                                       IL-2
                                     T
                                     I
                                     O                          IL-
                                                                IL-2 R (High affinity)
                                     N
        Antigen /T-cell
        receptor/ MHC II


                                                                                 Everolimus
                                                                                  Sirolimus
       P
       R         T
       O     Lymphocyte
       L
       I
       F
       E                                                                    DNA
       R                                                                  synthesis
       A
       T         T
       I
       O     Lymphocyte
       N
                                   Mycophenolic acid
                                     Azathioprine
         Immune system:
3 signal model of T-cell activation
Signal 1: Antigen-specific signal
 Donor antigen is presented on APC
 (antigen presenting cell)
 T-cell recognized antigen as “non-self”
 Complex formed between MHC-Antigen-
 Tcell receptor (TCR)
 Immune signal is transduced through CD3
 complex


                         N Engl J Med 2004;351:2715-29.
 Signal 2: Non-antigen-specific
     co-stimulatory signal
Binding of co-receptors between APC and
T-cell
– CD80/86 (aka B7) on APC with CD28 on T-
  cell
“reinforces” and strengthens immune
signal transduced through CD3 complex
Combined action of signals 1 & 2 activate
important intracellular pathways

                          N Engl J Med 2004;351:2715-29.
Combined action of signals 1 & 2 activate
important intracellular pathways
– Calcium-calcineurin pathway
– RAS-MAP kinase pathway
– Nulcear factor-κB pathway
Leads to
– production of IL-2 and other growth promoting
  cytokines
– Expression of IL-2 receptor (CD25)

                             N Engl J Med 2004;351:2715-29.
 Signal 3: Mammalian target of
 rapamycin (mTOR) activation
IL-2 produced binds to newly activated IL-
2 receptor (CD25)
Binding activates mTOR
mTOR activation triggers cell cycle
– Lymphocyte proliferation
– Results in large numbers of effector T-cells




                             N Engl J Med 2004;351:2715-29.
   Result of effector T-cells
Effector T-cells targeted at donor antigen
infiltrate graft
– Interstitial and/or perivascular infiltration
– Cascade of activation of macrophages, B-
  cells, plasma cells
– Resultant cell lysis
– Severe – edema, hemorrhage, vasculitis
              Three Signal Model of T cell Activation
             [adapted from Sing-Leung 2001]

              Antigen Presenting Cell



             Signal 1          Signal 2                       Signal 3


                   MHC II            B7
                         Antigen              Interleukin-2

CD45   CD4              CD3         CD28                          Interleukin-2 receptor


        TCR
         Calcineurin                                     TOR
          Pathway                                       Pathway

                  Cytokine gene                                            Purine
                                                       Cell Cycle         Synthesis
                     nucleus
                 T Lymphocyte                                 Target lymphocyte
   Sites of Action of Immunosuppressive Medications
  [adapted from Sing-Leung 2001]

              Antigen Presenting Cell
                                                        Steroids


             Signal 1           Signal 2                                Signal 3


 ATG                MHC II           B7
                                                        Interleukin-2
                                                                                     Anti-IL-2R
                     OKT3
                                           Belatacept

CD45    CD4                CD3       CD28                                  Interleukin-2 receptor

       TCR                                                                         Sirolimus
             Calcineurin         Cyclosporine                       TOR
                                 & Tacrolimus                      Pathway          MMF
              Pathway
Steroids
                  Cytokine gene
                                                                                    Purine
                                                                   Cell Cycle
                      nucleus                                                      Synthesis

                  T Lymphocyte                                       Target lymphocyte
Individual Immunosuppressive Drugs and Sites of
Action in the Three-Signal Model




                              N Engl J Med 2004;351:2715-2729
Target Antigens
  Immunosuppressant drugs
                         Anti-metabolites
                         – Azathioprine
Calcineurin inhibitors   – Mycophenolates
                         – Leflunomide?
– Ciclosporin A
– Tacrolimus
                         m-TOR inhibitors
                         – Sirolimus
                         – Everolimus
     Newer antibody drugs
Alemtuzumab
Rituximab



IL-2 receptor ‘mabs’
– Basiliximab
– Daclizumab
             CNI’s:
  Tacrolimus and Cyclosporine
Major breakthrough in modern
transplantation
Lead to significant improvements in
outcome

“Backbone” of most IMS protocols
Similar mechanisms of action
– Differ in structure and receptor interactions
       Transplant Proc 2004;36:25S-32S. Am J Kidney Dis 2006;47:S3-21.
  Calcineurin inhibitors (CNIs) :
      Mechanism of action
Forms complex with cytosolic proteins (aka
immunophilins)
– CSA = cyclophilin
– Tacrolimus = FKBP-12 (FK-binding protein)

↑ affinity and binding of immunophilin-drug
complex to calcineurin

Inhibits of transcription of IL-2, other cytokines
↓ T-cell activation & proliferation

         Transplant Proc 2004;36:25S-32S. Circulation 2004;110:3858-65.
 Calcineurin inhibitors (CNIs)
Ciclosporin A (CyA) binds to intracellular
protein, cyclophilin
    only active once bound


Tacrolimus (FK506) binds to intracellular
protein, FKBP-12
    only active once bound
The fungus Tolypocladium
        inflatum
             Fungal source of CyA

             Hydrophobic cyclic
             endecapeptide
Scanning electron micrograph
of Streptomyces tsukubaensis
               Bacterial source of
               tacrolimus

               Sirolimus from
               Streptomyces
               hygroscopicus & very
               similar structure
       Basic mode of action
(Drug + immunophilin) inhibits calcineurin

– Prevents dephosphorylation (activation) of NF-Act T-
  cell
– factors which stimulate cytokine (i.e. IL-2/IFN-) gene
  transcription
– Net result: impaired IL-2 production

‘reversible inhibition of T-cell activation,
proliferation & clonal expansion’
– Stops cell cycle at G0-G1 stage
Ciclosporin A & tacrolimus share a
mechanism of action and metabolic route

Never given together (additive
nephrotoxicity)
    Tac interacting with CyA?
    12-24hr gap only if converting


Differences include ADR’s & efficacy
               Ciclosporin
Oral absorption
– Large inter & intra-patient variability
– Food & juices
– IV dose & administration
Distribution – extensive (not dialysed)
Metabolism
– CYP450 3A4 with adult half-life 5-18 hours
    Shorter in children
    Steady state 2-3 days after change of dose
               Tacrolimus
Absorption
– Fatty food decreases rate & extent
>80% distributes into erythrocytes
– Whole blood trough levels
– Remainder 99% protein bound
CYP450 3A4 metabolism
– Adult half-life approx 9-12 hours
    Clearance faster in <6yrs
   Metabolic interactions on CNI & m-TOR inhibitor levels

                 Interacting drug             Effect on blood level

Erythromycin & clarithromycin              Increased

Diltiazem, nicardipine, verapamil          Increased

Fluconazole, itraconazole, ketoconazole,   Increased
   voriconazole

Rifampicin                                 Decreased

Carbamazepine                              Decreased

Phenobarbital                              Decreased

Phenytoin                                  Decreased

St John’s Wort                             Decreased
    Typical CNI doses target levels
Calcineurin    Common            Typical target blood
 inhibitor     initial oral             levels
                 doses        0-6 months Greater 6
                                            months
Ciclosporin   5mg/Kg bd       150 to        75 to
                              300ng/mL 150ng/mL
Tacrolimus    0.15mg/Kg       10 to         5 to
              bd              15ng/mL       10ng/mL

              Once daily?
         Anti-metabolites
Only purine anti-metabolites routinely used

1) Azathioprine

2) Two mycophenolate presentations
– mofetil
– sodium EC
            Azathioprine
Guanine anti-metabolite
– rapidly converted  6-mercaptopurine
   thioguanine nucleotides
– Disrupts synthesis of developing cellular DNA
  & RNA strands & so prevent mitosis
‘Prevents early stages of activated T & B-
cell proliferation’
One metabolic route involves thiopurine
methyltransferase (TPMT).
– Absent in 1 in 300
    Toxicity
Final metabolism to inactive 6-thiouric acid
– Via xanthine oxidase
    Quarter dose in combination with allopurinol


Usual starting dose 1-2mg/Kg
– Can use 50% dose if giving iv
Maintenance immunosuppression over first year after transplantation (1994-2003).
                    (2005 OPTN/SRTR Annual report)
Mycophenolic acid precursors
Isolated in 1896 from Penicillium spp.

mofetil ester rapidly cleaved  active
mycophenolic acid (MPA)
1 enzyme is rate limiting step in lymphocyte de
novo purine biosynthesis
– Inosine Mono-Phosphate DeHydrogenase


Mycophenolic acid non-competitively inhibits
IMPDH
– Prevents production of Guanosine triphosphate (GTP)
– Impairs production of DNA limiting lymphocyte
  proliferation


T & B-lymphocytes preferentially use this
pathway for purine production.
– Other cell lines have salvage pathways.
Molar equivalent doses:
– 1gram MMF = 720mg MPS EC
    Paed MMF dose with ciclosporin 600mg/M2 bd
    MMF dose with tacrolimus = less (50-25%)
     – MPA glucuronide undergoes entero-hepatic recirculation
     – Second peak maintains levels
     – ciclosporin inhibits this recirculation

MMF injection available if necessary
– Same dose iv as oral
– Requires aseptic manipulation
             Interactions
Magnesium & aluminium (antacids)
– Reduced absorption
Cholestyramine
– Reduced entero-hepatic recirculation
(Val) aciclovir
– Competes for tubular secretion with MPAG
– Modest effect
    +12% MPAG AUC & +20% aciclovir AUC
Pros & cons of mycophenolate
Reduced incidence of ACR c.f. placebo or
azathioprine with Sandimmune & steroid
Lower incidence CAN c.f. azathioprine
– Reduced interstitial fibrosis?
Much more expensive than azathioprine
Higher incidence of GI ADR - strategies
     diarrhoea 12.7 – 31.5%
     abdominal pain 11.5 – 26%
     nausea 4.2 – 14% and vomiting 2.4 – 12%.
Higher incidence of anaemia (15%)
                      Sirolimus
Another macrocyclic antibiotic
– Immunophilin binder (like CyA & FK506)
      also bind ubiquitous protein, FKBP-12
Does not inhibit cytokine (i.e. IL-2) gene transcription

Complex inhibits m-TOR
– Inhibits multiple biological steps
      e.g. activation of p70S6 kinase
– Prevents IL-2-mediated signal transduction to cell
  nucleus
      Cell cycle arrest at G1-S phase

Also inhibits tumour cell & smooth muscle cell
proliferation (CAN)
Absorption is poor
– Food also affects rate & extent
    Consistency


Widely distributed into tissues

CYP450 3A4 & P-glycoprotein metabolism
– Adult half-life long (57-63 hours)
    Clearance slightly higher in <11yrs cf 12-18yrs


Levels 4-12ng/mL, no more frequently
than every fortnight.
Adverse effect profile dissimilar to CNIs
– Not nephrotoxic


– Wound healing delay & fascial dehiscence
      lymphocele formation
–   Mouth ulceration – transient? Topical steroid
–   Acne and pilosebaceous rash - doxycycline
–   Peripheral & angioedema >10% - withdrawal
–   Dose related mixed hyperlipidaema - statins
–   Anaemia & thrombocytopaenia
–   Arthralgias – dose reduction or withdrawal
–   Interstitial lung disease - withdrawal
Adverse events leading to discontinuation

         Reason            SRL              CNI
                          (N=85)           (N=53)

      Chronic allograft     0                 1
       nephropathy
     Raised creatinine     1                  1
       Mouth ulcers        6                  0
       Pneumonitis         2                  0
           Rash            10                 0
         Diarrhoea         3                  0
    Other Adverse Event    10                 0
           Total           32                 2


                            Transplantation 2003: 76; 364-370.
         Use of sirolimus
Licensed for use with ciclosporin A for 1st
3 months only in adult renal transplant
– Rarely used in this way
Where to position sirolimus?
– Replace CNI or replace anti-metabolite
Appears to offer better GFR (graft
survival?) than CNI maintenance.
Skin cancers??
              Alemtuzumab
Licensed for chronic lypmphcytic leukaemia

Recombinant humanised mab vs CD52
– CD52 cell surface antigen
     T&B lymphocytes, monocytes, macrophages

Depleting mab
– Binds and leads to immediate cell lysis

Single 30mg dose leads to profound lymphocyte
depletion
– Persists 6-12 months before recovery
– Optimal dose, frequency & timing unknown
Mainly as induction immunosuppression
    Instead of rATG or IL-2R mabs
– Also used to treat acute rejection


Very limited RCT evidence
    Similar 1 year graft/patient survival & ACR
    incidence to basiliximab, dacilzumab & rATG
Infusion-related ADRs very common
– Fever, rigors, hypotension, urticaria,
  hypotension, bronchospasm, ARDS, arrest
– Must be infused over 2 hours
    Preceded by iv MePred, Piriton & paracetamol


Pancytopenia, leukopenia,
thrombocytopenia
                   Rituximab
Chimeric mouse/human mab vs CD20
– CD20 cell surface antigen
    Pre-B & mature B lymphocytes
      – Not stem cells or other cell lines

Depleting mab
– Inhibits B cell proliferation
    Reduces PRA titre
– New, healthy cells regenerate 9-12 months
    Effects still apparent at 15 months
Role of rituximab in transplant unclear
– Licensed for lymphoma/leukaemia &
  autoimmune disorders

Humeral rejection episodes
    Some data but no local experience


Pre-transplant, pre-conditioning regimes
– High donor-relevant HLA antibody titre
    With DFPE, IVIG (?), mycophenolate
    Single doses 375mg/M2
– ABO incompatible transplant
Early infusion-related ADRs in > 50%
– Classic ‘cytokine –release’ syndrome
– Fever, rigors, hypotension, urticaria,
  bronchospasm, angioedema, ARDS, death
– Infusion started slowly and rate adjusted
    Preceded by iv steroid, Piriton & paracetamol
Tumour-lysis syndrome association
Leucopaenia/thrombocytopenia
– Less common than with alemtuzumab
      Anti-CD 25 antibodies
Two IL-2 receptor (IL-2R) ‘mabs’
– Daclizumab
– Basiliximab


Both ‘humanised’ monoclonal antibodies
against the  chain of IL-2R
– aka anti-Tac/CD25 antibodies
Bind to, saturate & block activated IL-2R
– don’t affect resting T-cells

Both manipulated to prevent patient
antibody formation
– unlike OKT3 or ???Thymoglobuline


Used exclusively as induction agents
– Duration
Pooled analysis for anti-CD 25 antibodies


              Biopsy confirmed      Biopsy confirmed
              acute rejection       acute rejection
              absolute difference   odds ratio
              (95% CI)              (95% CI)

Daclizumab    -16%                  0.48               New Engl J Med
                                                       1998: 338; 161-165
versus        (-24% to –8%)         (0.33 to 0.70)     Transplantation
placebo                                                1999: 67; 110-115

Basiliximab                                            Lancet 1997: 350;
              -14%                  0.51               1193-1198
versus        (-20% to –9%)         (0.40 to 0.65)     Transplantation 1999:
placebo                                                67; 276-284
                                                       Transplantation 2001:
                                                       72; 1261-1267
        IL-2R blocking ‘mabs’
Advantages                         Disadvantages
– reduce renal rejection rates     – Cost
  by approx 30% in classic              But cost-effective to add
  triple therapy                        basiliximab to ciclosporin
                                        regimens
                                        Same report states that
– short courses
                                        should not be added to
      iv, in hospital only              tacrolimus based
      Compliance guaranteed             regimens.

– ‘Few’ adverse effects            – Daclizumab is a 5 dose,
      Severe re-exposure             fortnightly regimen &
      hypersensitivity seen with     requires infusion
      both

								
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