Organ Transplantation - Immunosuppressive Drugs

Diagnosis

Indications for Testing

  • Immunosuppressant dose optimization
  • Failure to respond to immunosuppressants
  • Signs or symptoms consistent with inadequate or excessive immunosuppression
  • Changes to concomitant medications or other variables that affect pharmacokinetics

Monitoring

  • Therapeutic drug monitoring is required for patients on therapy

Pharmacogenetics and Therapeutic Drug Monitoring

  • Thiopurine drugs
    • Thiopurine prodrugs are metabolized via thiopurine methyltransferase (TPMT) enzymatic activity
    • Deficiency of TPMT predicts hematopoietic toxicity after thiopurine treatment
    • Testing to determine activity level may be helpful in dosing thiopurine drugs and help avert bone marrow suppression
      • For deficient activity, dose reduction of 80-90% may be required
      • For intermediate activity, dose reduction of 20-50% may be required
    • Genotype for TPMT cannot be inferred from TPMT activity (phenotype)
    •  TPMT phenotype testing does not replace need for clinical monitoring of patients treated with thiopurine drugs
      • Phenotype testing should not be requested for patients currently treated with thiopurine drugs; results will be falsely low
      • Current TPMT phenotype may not reflect future TPMT phenotype, particularly in patients who received blood transfusions within 30-60 days of testing

Clinical Background

Organ transplantation is the strategy of choice for end-stage organ disease. Immunosuppressive therapies have allowed patients to extend the life of the organ but require careful monitoring to prevent toxicity and rejection. Therapeutic ranges and toxic thresholds should be carefully considered based on the clinical setting. Important factors include the organ transplanted, time posttransplant, age and clinical status of the patient, and concomitant medications.

Epidemiology

  • Prevalence – in 2005, >60,000 patients in the U.S. were living with functioning organ transplants
  • Survival rate 1 year post graft – 90%

Immunosuppressive Drug Regimens for Organ Transplants

Available Immunosuppressives (immunosuppressive regimen depends on organ transplanted; most common combination is steroid plus calcineurin)

Mechanism of Action

Toxicity

Therapeutic Ranges

Antithymocyte globulin (ATG, Atgam, Thymoglobulin)

Depletes lymphocytes

Increases risk of infection

Aim for 0.1-0.3 lymphocytes/mL

Corticosteroids

Proapoptotic effect on lymphoid cells; suppresses eicosanoid production; increases TGF expression

Increases risk of infection and malignancy (nonmelanoma skin cancers [NMSC])

 

IL-2 antibodies

  • Daclizumab (Zenapax)
  • Basiliximab (Simulect)

Selectively blocks IL-2 receptors on T helper cells, preventing T-cell proliferation

Increases risk of malignancy (NMSC and lymphomas)

 

Muromonab-CD3

(Orthoclone OKT3)

Depletes lymphocyte T-cells 

Increases risk of infection

500-1,500 ng/mL during steady state treatment

Alemtuzumab (Campath-IH)Targets CD52 on T-cells, B-cells, and NK-cells, causing depletionIncreases risk of infection 
Rituximab (Rituxan, MabThera)Binds CD20 and B-cells mediating lysisIncreases risk of infection 
Belatacept (LEA 29Y)Binds to T-cells to prevent CD28 signallingIncreases risk of infection 

Immunosuppressants

  • Azathioprine (Imuran)
  • 6-mercaptopurine (Puinethol)

Inhibits purine nucleotide synthesis, which interferes with DNA synthesis

Increases risk of infection and malignancy (acute myeloid leukemia and myelodysplastic syndromes)

 
  • Mycophenolic acid (CellCept, Myfortic)

Selectively inhibits inosine monophosphate dehydrogenase – interferes with DNA purine synthesis

Increases risk of infection

Toxic ranges are not well established, except for mycophenolic acid

Mycophenolic acid – >25 μg/mL

Suggested therapeutic range (for 2 g/day dosing

  • Mycophenolic acid – 1.0-3.5 μg/mL
    • 3 g/day dose may have up to 5.0 μg/mL concentration
      • 2-4 μg/mL range is suggested for maximal efficacy with minimal toxicity
    • Mycophenolic acid glucuronide – 35-100 μg/mL
    • Mycophenolic acid acyl-glucuronide – not well established
  • Sirolimus (rapamycin, Rapamune)
  • Everolimus (Zortress)

Blocks B- and T-cell proliferation by blocking pathway between IL-2 receptor and nucleus; does not block calcineurin pathway; synergistic with cyclosporine

Increases risk of hyperlipidemia and infection

Measured as trough level

Sirolimus therapeutic ranges for kidney transplant (in combination with cyclosporine)

  • 4-12 ng/mL

Liver transplant

  • 12-20 ng/mL (suggested range)

Everolimus – 3-8 ng/mL

  • Cyclosporine A (Sandimmune, Neoral, Gengraf) 

Inhibits calcineurin phosphatase and reduces IL-2 expression

Increases risk of malignancy, nephrotoxicity, cardiotoxicity, hyperlipidemia

Toxic level – >700 ng/mL

General therapeutic range – 100-400 ng/mL

Kidney transplant (in combination with Everolimus)

  • 1 month posttransplant – 100-200 ng/mL
  • 2-3 months posttransplant – 75-150 ng/mL
  • 4-5 months posttransplant – 50-100 ng/mL
  • 6-12 months posttransplant – 25-50 ng/mL

Heart transplant

  • Up to 3 months posttransplant – 350-525 ng/mL
  • ≥4 months posttransplant – 145-350 ng/mL

Liver transplant 

  • 290-525 ng/mL
  • Tacrolimus (FK506, Prograf)

Inhibits calcineurin phosphatase and reduces IL-2 expression

Increases risk of malignancy, nephrotoxicity, cardiotoxicity, hyperlipidemia

Toxic level – ≥25 ng/mL

Measured as trough level

Therapeutic ranges for kidney transplant

  • 0-3 months posttransplant – 7.0-20.0 ng/mL
  • ≥3 months posttransplant – 5.0-15.0 ng/mL

Heart transplant

  • 0-3 months posttransplant – 10.0-20.0 ng/mL
  • ≥3 months posttransplant – 5.0-15.0 ng/mL

Liver transplant

  • 1-12 months posttransplant – 5-20 ng/mL

Indications for Laboratory Testing

  • Tests generally appear in the order most useful for common clinical situations
  • Click on number for test-specific information in the ARUP Laboratory Test Directory
Test Name and Number Recommended Use Limitations Follow Up
Cyclosporine A by Tandem Mass Spectrometry 0070035
Method: Quantitative Liquid Chromatography-Tandem Mass Spectrometry

Optimize dosage; monitor compliance

Results from different methodologies (mass spectrometry versus immunoassay) cannot be used interchangeably

Generally, immunoassay methods have been reported to have a positive bias in results when compared to mass spectrometry due to antibody cross-reactivity

 
Cyclosporine A, 2-Hour Post Dose (C2) by Tandem Mass Spectrometry 0058902
Method: Quantitative Liquid Chromatography-Tandem Mass Spectrometry

Optimize dosage; monitor compliance

   
Everolimus by Tandem Mass Spectrometry 0092118
Method: Quantitative Liquid Chromatography-Tandem Mass Spectrometry

Therapeutic monitoring for individuals taking everolimus

  • Trough concentrations should be assessed ~2 weeks after beginning treatment

Analytical sensitivity – limit of detection is 2.0 ng/mL

  • Interferences from commonly used drugs and associated metabolites have not been observed

Results from different methodologies (mass spectrometry versus immunoassay) cannot be used interchangeably

Generally, immunoassay methods have been reported to have a positive bias in results when compared to mass spectrometry due to antibody cross-reactivity

 
Mercaptopurine and Metabolites Quantitative, Blood 2011006
Method: Quantitative High Performance Liquid Chromatography-Tandem Mass Spectrometry 

Therapeutic monitoring

Measures concentration of parent drug only; risk of toxicity associated with TPMT deficiency should be evaluated by TPMT, RBC

 
Mycophenolic Acid and Metabolites 2010359
Method: Quantitative Liquid Chromatography-Tandem Mass Spectrometry

Therapeutic monitoring

Analytical sensitivity – limit of detection is 0.5 µg/mL

Toxic ranges are not well established except for mycophenolic acid (>25 μg/mL)

 
Sirolimus by Tandem Mass Spectrometry 0098467
Method: Quantitative High Performance Liquid Chromatography-Tandem Mass Spectrometry

Therapeutic monitoring

Results from different methodologies (mass spectrometry versus immunoassay) cannot be used interchangeably

Generally, immunoassay methods have been reported to have a positive bias in results when compared to mass spectrometry due to antibody cross-reactivity

 
Tacrolimus by Tandem Mass Spectrometry 0090612
Method: Quantitative Liquid Chromatography-Tandem Mass Spectrometry

Therapeutic monitoring

Results from different methodologies (mass spectrometry versus immunoassay) cannot be used interchangeably

Generally, immunoassay methods have been reported to have a positive bias in results when compared to mass spectrometry due to antibody cross-reactivity

 
Thiopurine Methyltransferase, RBC 0092066
Method: Enzymatic/Quantitative Liquid Chromatography-Tandem Mass Spectrometry

Detect risk for severe myelosuppression with standard dosing of thiopurine drugs

Individualize dosing of thiopurine drugs

Does not replace clinical monitoring

Genotype cannot be inferred from TPMT activity (phenotype)

TPMT inhibitors may contribute to false-low test results

TPMT activity should be assessed prior to treatment with thiopurine drugs

Blood transfusion within 30 days will reflect donor status

 
Thiopurine Drug Metabolites 2011134
Method: Quantitative Liquid Chromatography/Tandem Mass Spectrometry

Optimize therapy for thiopurine drugs

Identify thiopurine metabolite concentrations that may lead to toxicity

Limit of quantification (LOQ)

  • LOQ – 12.5 pmol/8 x 108 RBC (6-TGN)
  • LOQ – 325 pmol/8 x 108 RBC (6-methyl mercaptopurine nucleotide [6-MMPN])
 
Lymphocyte Transplantation CD3 0095949
Method: Quantitative Flow Cytometry

Monitor immunosuppressive therapy with muromonab-CD3; test verifies CD3 antigen removal

 

For testing of immunocompromised patients, order Lymphocyte Subset Panel 4 – T-Cell Subsets Percent and Absolute 

Lymphocyte Transplantation Profile 0095798
Method: Quantitative Flow Cytometry

Monitor immunosuppressive therapy with anti-lymphocyte drugs, such as muromonab or ATG 

Components include CD2 percent and absolute, CD3 percent and antibodies, CD4 percent, CD8 percent, CD4;CD8 ratio, CD19 percent