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Interpretive Handbook

Test 62751 :
Thiopurine Methyltransferase (TPMT) Genotyping, Saliva

Clinical Information Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test

The thiopurine drugs are purine antimetabolites that are useful in the treatment of acute lymphoblastic leukemia, autoimmune disorders (eg, Crohn disease and rheumatoid arthritis), and organ transplant recipients. The thiopurine drugs, 6-mercaptopurine (6-MP), 6-thioguanine (6-TG), and azathioprine (AZA), are prodrugs that require intracellular activation to 6-thioguanine nucleotides (6-TGN). This activation is catalyzed by multiple enzymes. The cytotoxic effects of thiopurine drugs are achieved mainly through incorporation of 6-TGNs into DNA and RNA. The pathway that leads to synthesis of active cytotoxic 6-TGNs is in competition with inactivation pathways catalyzed by thiopurine methyltransferase (TPMT). Evaluation of this pathway is important because the levels of 6-TGNs measured in red blood cells have been correlated with both thiopurine therapeutic efficacy and toxicity such as myelosuppression.

 

TPMT activity is inherited as a monogenic co-dominant trait and variable TPMT activity is associated with TPMT genetic variants. The distribution of TPMT activity in red blood cells is trimodal in Caucasian population, with approximately 0.3% of people having deficient (undetectable) TPMT activity, 11% low (intermediate) activity, and 89% normal TPMT activity. The allele for normal TPMT activity (wild-type) has been designated TPMT*1. Four TPMT alleles, comprised of a combination of 3 different Single nucleotide polymorphisms (SNPs), account for the majority of inactivating alleles in some ethnicities, including Caucasians: TPMT*2, TPMT*3A, TPMT*3B, and TPMT*3C. Additional less frequent TPMT alleles TPMT*4, TPMT*5, TPMT*8, and TPMT*12 also have been implicated as deficiency alleles. There is substantial evidence linking TPMT genotype to phenotypic variability and, therefore, genotyping tests have a high likelihood of being informative in these populations.

 

TPMT Allele

cDNA Nucleotide Change

Effect on Enzyme Metabolism**

*1

None (wild type)

Extensive (normal) metabolizer

*2

c.238G->C

Nonfunctional, no activity

*3A

c.460G->A and c.719A->G

Nonfunctional, no activity

*3B

c.460G->A

Nonfunctional, no activity

*3C

c.719A->G

Nonfunctional, no activity

*4

c.626-1G->A

Nonfunctional, no activity

*5

c.146T->C

Nonfunctional, no activity

*8

c.644G->A

Reduced-function/decreased activity

*12

c.374C->T

Reduced-function/decreased activity

 

The US Food and Drug Administration, the Clinical Pharmacogenetics Implementation Consortium, and certain professional societies recommend consideration of TPMT genotype or TPMT erythrocyte testing prior to the initiation of therapy with thiopurine drugs. Dose adjustments based upon TPMT genotype have reduced thiopurine-induced adverse effects without compromising desired antitumor and immunosuppressive therapeutic effects in several clinical settings. Complementary clinical tests are available to measure TPMT enzymatic activity in erythrocytes. Genotyping is not impacted by other medications known to inhibit TPMT activity. This testing can be complimented by the TPMT erythrocyte phenotype testing if the clinician wants to check for lower TPMT enzyme activity regardless of cause.

Useful For Suggests clinical disorders or settings where the test may be helpful

Predicting potential for toxicity to thiopurine drugs (6-mercaptopurine, 6-thioguanine, and azathioprine)

 

Genotyping patients who prefer not to have their blood drawn

Interpretation Provides information to assist in interpretation of the test results

An interpretive report will be provided that includes assay information, genotype, and an interpretation indicating whether results are predictive of a poor, intermediate, or extensive metabolizer phenotype.

 

The genotype, with associated star alleles, is assigned using standard allelic nomenclature as published by the TPMT Nomenclature Committee.(1)

Cautions Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances

Saliva samples may contain donor DNA if obtained from patients who received heterologous blood transfusions or allogeneic blood or marrow transplantation. Results from samples obtained under these circumstances may not accurately reflect the recipient's genotype. For individuals who have received blood transfusions, the genotype usually reverts to that of the recipient within 6 weeks. For individuals who have received allogeneic blood or marrow transplantation, a pretransplant DNA specimen is recommended for testing. 

 

Rare variants exist that could lead to false-negative or false-positive results. If genotype results obtained do not match the clinical findings, additional testing should be considered for thiopurine methyltransferase (TPMT).

 

The results do not rule out the possibility that a patient harbors another variation that can impact drug response or drug side effects. These genotyping procedures will not distinguish between heterozygous *3A from the rare *3B/*3C, which has a frequency of 1:120,890. This rare genotype is associated with low enzyme activity. Enzyme activity evaluation is necessary to definitively identify this rare genotype TPMT.

 

This PCR-based assay tests for the presence of TPMT*2 (c.238G->C), TPMT*3A (c.460G->A, c.719A->G), TPMT*3B (c.460G->A), TPMT*3C (c.719A->G), TPMT*4 (c. 626-1G->A), TPMT*5 (c.146T->C), TPMT*8 (c.644G->A), and TPMT*12 (c.374C->T). This test will not detect all TPMT genetic variants. A negative result does not rule out the possibility of toxicity if thiopurines are used, since multiple factors (eg, other genetic factors, drug-drug interactions) are known to play a role. Coprescription of allopurinol might inhibit TPMT activity. Drugs that have been shown to inhibit TPMT activity include: naproxen, ibuprofen, ketoprofen, furosemide, sulfasalazine, mesalamine, olsalazine, mefenamic acid, thiazide diuretics, and benzoic acid inhibitors.

Reference Values Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.

An interpretive report is provided.

Clinical References Provides recommendations for further in-depth reading of a clinical nature

1. TPMT nomenclature committee. Available at: http://www.imh.liu.se/tpmtalleles

2. Appell ML, Berg J, Duley J et al: Nomenclature for alleles of the thiopurine methyltransferase gene. Pharmacogenet Genomics 2013;23(4):242-248

3. Nguyen CM, Mendes MA, Ma JD: Thiopurine methyltransferase (TPMT) genotyping to predict myelosuppression risk. PLoS Curr 2011;3:RRN1236

4. 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(3):387-391

5. Weinshilboum R: Thiopurine pharmacogenetics clinical and molecular studies of thiopurine methyltransferase. Drug Metab Dispos 2001 Apr;29(4 Pt 2):601-605

6. Zaza G, Cheok M, Krynetskaia N, et al: Thiopurine pathway. Pharmacogenet Genomics 2010 Sep;20(9):573-574


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