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Thiopurine Test Selection Guide

Thiopurine Test Selection Guide

Test Guide

Thiopurine Test Selection Guide

  

Thiopurines (azathioprine, 6-mercaptopurine, and thioguanine) are used to treat patients with inflammatory bowel disease (IBD), leukemia, rheumatic disease, or solid organ transplantation. These drugs are converted, through a multistep process (Figure), to the active metabolite 6-thioguanine nucleotide (6-TGN). Though 6-TGN is responsible for the therapeutic effects of thiopurines, excess 6-TGN can lead to toxicity. In contrast, low levels of 6-TGN can lead to an inadequate response to treatment. Laboratory testing can help assess the risk of toxicity and identify the cause of inadequate response.

Figure. Thiopurine Metabolism

Levels of thiopurine metabolites are partially dependent on thiopurine methyltransferase (TPMT). The TPMT enzyme decreases 6-TGN production by converting thiopurine metabolites to inactive forms, including 6-methylmercaptopurine (6-MMP). Patients who have low TPMT activity and receive standard thiopurine doses usually have high levels of 6-TGN and may be susceptible to hematologic toxicity (eg, myelosuppression).2,3 Measurement of TPMT activity or TPMT genotype can help identify these patients. In contrast, a subset of patients preferentially convert thiopurine to 6-MMP instead of 6-TGN. These “shunters” may have high levels of 6-MMP and be susceptible to hepatotoxicity.4 Measurement of thiopurine metabolites (6-TGN and 6-MMP) can help identify these patients. Furthermore, metabolite levels can help identify causes of inadequate response to thiopurine.

Thiopurine package inserts recommend considering testing for TPMT activity or genotype.5,6 Although the inserts do not provide specific dosing advice in response to test results, the Clinical Pharmacogenetics Implementation Consortium (CPIC) published guidelines in 2011 (and affirmed them in 2013) that do.1,7 The American Gastroenterological Association (AGA) also suggests TPMT testing (activity or genotype) for adults with IBD who are starting thiopurine treatment; the suggestion is based on reducing risk of adverse events in patients with abnormal TPMT activity.8 The AGA also suggests testing thiopurine metabolites for adults with active IBD or those who experience adverse side effects that suggest thiopurine toxicity.8

This Guide provides an overview of laboratory tests that, in the context of the patient's clinical and laboratory history, may assist with decision-making related to thiopurine treatment. A list of relevant tests is provided in Table 1. The Table is provided for informational purposes only and is not intended as medical advice. A physician's test selection and interpretation, diagnosis, and patient-management decisions should be based on his/her education, clinical expertise, and assessment of the patient.

Table 1. Available Tests Related to Thiopurine Treatment

Test Code

Test Name

Clinical Use8

18831a

TPMT Activity

Identify patients at risk for toxicity from thiopurine drugs; help determine need to adjust drug dosage or select alternative treatment

37742a

Thiopurine S-Methyltransferase (TPMT) Genotype

Identify patients at risk for toxicity from thiopurine drugs; help determine need to adjust drug dosage or select alternative treatment

91745a

Thiopurine Metabolites
Includes 6-TGN and 6-methylmercaptopurine (6-MMP)

Help guide thiopurine treatment in patients with active IBD or experiencing adverse effects that suggest thiopurine toxicity

6-MMP indicates 6-methylmercaptopurine; 6-TGN, 6-thioguanine nucleotide; IBD, inflammatory bowel disease; TPMT, thiopurine methyltransferase.

a

This test was developed and its analytical performance characteristics have been determined by Quest Diagnostics. It has not been cleared or approved by the FDA. This assay has been validated pursuant to the CLIA regulations and is used for clinical purposes.

TPMT Activity and TPMT Genotype Tests
Package inserts5,6 and guidelines8 often group testing for TPMT activity and TPMT genotype together because the tests are related: alterations in the TPMT gene can cause lower TPMT activity. Individuals with 2 variant alleles have low or no TPMT activity (0.3% of individuals), while those with 1 variant allele have intermediate activity (~10% of individuals). On the other hand, wild-type homozygotes have normal enzyme activity (~90% of individuals).9

Though the tests are related, they have different limitations. Understanding the limitations of each assay may help a physician decide which one to use.

  • TPMT activity assays have the limitation that test results may be falsely elevated by recent blood transfusions.4-6  In addition, results can be affected by medications (eg, aminosalicylates, aspirin, furosemide) that inhibit TPMT activity.6 Thus, TPMT genotype testing may be appropriate for patients who have received such medications or recent transfusions.
  • TPMT genotype assays are limited by the specific TPMT alleles that are tested; the assays also do not identify variant alleles in other enzymes involved in thiopurine metabolism. Though 4 alleles account for most instances of lower TPMT activity, toxicity can occur when the tested variant alleles are absent. Thus, measuring TPMT activity may be appropriate when no allelic variants are identified in patients who experience toxicity.

The Quest Diagnostics TPMT Activity test quantifies 6-MMP levels; the test uses liquid chromatography-tandem mass spectrometry (LC-MS/MS). The TPMT Genotype test identifies 4 variant alleles that together account for 80% to 95% of reduced TPMT activity10: TPMT*2 (238G>C), TPMT*3A (460G>A and 719A>G), TPMT*3B (460G>A), and TPMT*3C (719A>G); the test uses a polymerase chain reaction (PCR) method followed by single-nucleotide primer extension. A “normal” result for either assay does not rule out the occurrence of adverse events; viral infection, concurrent use of some medications, and disruption of other components of thiopurine metabolism can contribute to adverse events.

Thiopurine Metabolites
Measuring thiopurine metabolites can help identify the cause of adverse effects related to thiopurine treatment. High levels of 6-TGN increase the risk of hematologic toxicity. High levels of 6-MMP increase the risk of hepatotoxicity.4

Levels of 6-TGN and 6-MMP can also help determine the cause of inadequate response to treatment (Table 2). Low 6-TGN levels could indicate a patient is receiving inadequate dose (underdosing), not taking the prescribed medication (nonadherence), or has an enzyme variant that preferentially converts thiopurine to 6-MMP instead of 6-TGN (shunting). Levels of 6-MMP can help differentiate these causes. Therapeutic or high 6-TGN levels in the presence of inadequate treatment response may indicate that a patient's condition is refractory to thiopurine.4

Table 2. Thiopurine Metabolite Levels and Possible Causes of Inadequate Response4  

6-TGN Levela

6-MMP Level

Possible Cause

Low

Normal

Underdosing

Low

Low

Nonadherence

Low

Highb

Shunting

Therapeutic

Normal or high

Treatment refractory, appropriately dosed

High

Normal or high

Treatment refractory, overdosed

6-MMP indicates 6-methylmercaptopurine; 6-TGN, 6-thioguanine nucleotide; RBC, red blood cell.

a

6-TGN units are pmol/8x108 RBCs: Low, <230; therapeutic, 230 to 450; high, >450.

b

>5700 pmol/8x108 RBCs, with 6-MMP/6-TGN ratio >11.

The Quest Diagnostics Thiopurine Metabolites test quantifies 6-TGN and 6-MMP levels using LC-MS/MS.

References

  1. Relling MV, Gardner EE, Sandborn WJ, et al. Clinical Pharmacogenetics Implementation Consortium guidelines for thiopurine methyltransferase genotype and thiopurine dosing. Clin Pharmacol Ther. 2011;89:387-391.

  2. Colombel JF, Ferrari N, Debuysere H, et al. Genotypic analysis of thiopurine S-methyltransferase in patients with Crohn's disease and severe myelosuppression during azathioprine therapy. Gastroenterology. 2000;118:1025-1030.

  3. Evans WE. Pharmacogenetics of thiopurine S-methyltransferase and thiopurine therapy. Ther Drug Monit. 2004;26:186-191.

  4. Vande Casteele N, Herfarth H, Katz J, et al. American Gastroenterological Association institute technical review on the role of therapeutic drug monitoring in the management of inflammatory bowel diseases. Gastroenterology. 2017;153:835-857.e836.

  5. Imuran [package insert]. San Diego, CA: Prometheus Laboratories, Inc; 2011.

  6. Purinethol [package insert]. Sellersville, PA: Gate Pharmaceuticals; 2011.

  7. Relling MV, Gardner EE, Sandborn WJ, et al. Clinical Pharmacogenetics Implementation Consortium guidelines for thiopurine methyltransferase genotype and thiopurine dosing: 2013 update. Clin Pharmacol Ther. 2013;93:324-325.

  8. Feuerstein JD, Nguyen GC, Kupfer SS, et al. American Gastroenterological Association institute guideline on therapeutic drug monitoring in inflammatory bowel disease. Gastroenterology. 2017;153:827-834.

  9. McLeod HL, Siva C. The thiopurine S-methyltransferase gene locus -- implications for clinical pharmacogenomics. Pharmacogenomics. 2002;3:89-98.

  10. Booth RA, Ansari MT, Loit E, et al. Assessment of thiopurine S-methyltransferase activity in patients prescribed thiopurines: a systematic review. Ann Intern Med. 2011;154:814-823, w-295-29.
     

Content reviewed 01/2018

 

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* The tests listed by specialist are a select group of tests offered. For a complete list of Quest Diagnostics tests, please refer to our Directory of Services.