Test Catalog

Test ID: TPNUV    
Thiopurine Methyltransferase (TPMT) and Nudix Hydrolase (NUDT15) Genotyping, Varies

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)

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, 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-TGN into DNA and RNA. The pathway that leads to synthesis of active cytotoxic 6-TGN is in competition with inactivation pathways catalyzed by thiopurine methyltransferase (TPMT). Evaluation of this pathway is important because the level of 6-TGN 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 codominant trait, and variable TPMT activity is associated with TPMT genetic variants. The distribution of TPMT activity in red blood cells is trimodal in Caucasians, 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 substitutions (SNP), account for the majority of inactivating alleles in some ethnicities, including Caucasians: TPMT*2, TPMT*3A, TPMT*3B, and TPMT*3C. Less frequently occurring TPMT alleles TPMT*4, TPMT*5, TPMT*8, and TPMT*12 also have been implicated as deficiency alleles. If no TPMT variant alleles are detected by this assay, the most likely genotype is that of TPMT*1/*1 although the presence of other rarer alleles cannot be excluded.

 

Nudix hydrolase (NUDT15) is thought to dephosphorylate the active metabolites of thiopurines, TGTP and TdGTP, which prevents their incorporation into DNA and decreases their cytotoxic effects. Genetic variants in NUDT15 that decrease this activity are strongly associated with thiopurine-related myelosuppression. NUDT deficiency is most common among East Asians (22.6%), followed by South Asians (13.6%), and Native American populations (12.5%-21.2%). Studies in other populations are ongoing. This test evaluates variants associated with NUDT15*2, NUDT15*3, NUDT15*4, and NUDT15 *5. If no NUDT15 variant alleles are detected by this assay, the most likely genotype is that of NUDT15*1/*1 although the presence of other rarer alleles cannot be excluded. Individuals with variants in both TPMT and NUDT15 have been identified and were significantly more sensitive to mercaptopurine than individuals with variants in only 1 gene. Integration of both TPMT and NUDT15 testing may allow for more accurate prediction of thiopurine-related toxicity risk to guide dosing, particularly among patients from diverse populations.

TPMT Allele

cDNA Nucleotide Change

Amino Acid Change

Effect on Enzyme Metabolism

*1

None (wild type)

None (wild type)

Normal function

*2

c.238G>C

p.Ala80Pro (p.A80P)

No activity

*3A

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

p.Ala154Thr (p.A154T) and p.Tyr240Cys (p.Y240C)

No activity

*3B

c.460G>A

p.Ala154Thr (p.A154T)

No activity

*3C

c.719A>G

p.Tyr240Cys (p.Y240C)

No activity

*4

c.626-1G>A

Not applicable, splice site

No activity

*5

c.146T>C

p.Leu49Ser (p.L49S)

No activity

*8

c.644G>A

p.Arg215His (p.R215H)

Reduced activity

*12

c.374C>T

p.Ser125Leu (p.S125L)

Reduced activity

 

The US Food and Drug Administration, the Clinical Pharmacogenetics Implementation Consortium, and some professional societies recommend consideration of TPMT genotype or TPMT erythrocyte testing prior to the initiation of therapy with thiopurine drugs. There is substantial evidence linking TPMT genotype to phenotypic variability. Dose adjustments based upon TPMT genotype have reduced thiopurine-induced adverse effects without compromising desired antitumor and immunosuppressive therapeutic effects in several clinical settings.

 

Genotyping is not impacted by other medications known to inhibit TPMT activity. Complementary clinical testing is available to measure TPMT enzymatic activity in erythrocytes (TPMT3 / Thiopurine Methyltransferase (TPMT) Activity Profile, Erythrocytes) if the clinician wants to check for lower TPMT enzyme activity, regardless of cause. Although there currently aren't guidelines or professional society recommendations related to NUDT15 genotyping to guide thiopurine use, this practice is substantially supported by the literature. Testing for TPMT enzyme activity is not impacted by variants in NUDT15.

NUDT15 Allele

cDNA Nucleotide Change

Amino Acid Change

Effect on Enzyme Metabolism

*1

None (wild type)

None (wild type)

Normal activity

*2 or *3

c.415C>T

p.Arg139Cys (p.R139C)

No activity

*4

c.416G>A

p.Arg139His (p.R139H)

No activity

*5

c.52G>A

p.Val18Ile (p.V18I)

No activity

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 will be provided.

Interpretation Provides information to assist in interpretation of the test results

An interpretive report will be provided.

 

The TPMT genotype, with associated star alleles, is assigned using standard allelic nomenclature as published by the TPMT Nomenclature Committee.(1) NUDT15 genotype and associated star alleles are as described by Moriyama et al.(2)

 

For additional information regarding pharmacogenomic genes and their associated drugs, see the Pharmacogenomics Associations Tables in Special Instructions. This resource also includes information regarding enzyme inhibitors and inducers, as well as potential alternate drug choices.

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

Rare variants may be present that could lead to false-negative or false-positive results. If no TPMT variant alleles are detected by this assay the most likely genotype is that of TPMT*1/*1 although the presence of other rarer alleles cannot be excluded. In addition, if no NUDT15 variant alleles are detected by this assay the most likely genotype is that of NUDT15*1/*1 although the presence of other rarer alleles cannot be excluded.

 

If genotype results obtained do not match the clinical findings, additional testing should be considered for thiopurine methyltransferase enzyme activity (TPMT3 / Thiopurine Methyltransferase [TPMT] Activity Profile, Erythrocytes). A corresponding activity assay for NUDT15 is not currently available.

 

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.

 

The results do not rule out the possibility that a patient harbors another variant in TPMT, NUDT15, or another gene that can impact drug response or drug side effects. These genotyping procedures will not distinguish between heterozygous TPMT*3A from the rare TPMT*3B/*3C, which has an estimated 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 (TPMT3 / Thiopurine Methyltransferase [TPMT] Activity Profile, Erythrocytes).

 

This test will not detect all TPMT or NUDT15 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.

Clinical Reference Recommendations for in-depth reading of a clinical nature

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

2. Moriyama T, Nishii R, Perez-Andreu V, et al: NUDT15 polymorphisms alter thiopurine metabolism and hematopoietic toxicity. Nat Genet 2016;48:367-373

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

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

5. 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

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

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

Special Instructions Library of PDFs including pertinent information and forms related to the test