Molecular diagnosis of thiopurine S-methyltransferase deficiency: Genetic basis for azathioprine and mercaptopurine intolerance

Charles Yates, Eugene Y. Krynetski, Thrina Loennechen, Michael Y. Fessing, Hung Liang Tai, Ching Hon Pui, Mary V. Relling, William E. Evans

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Abstract

Background: Thiopurine S-methyltransferase (TPMT) catalyzes the S- methylation (that is, inactivation) of mercaptopurine, azathioprine, and thioguanine and exhibits genetic polymorphism. About 10% of patients have intermediate TPMT activity because of heterozygosity, and about 1 in 300 inherit TPMT deficiency as an autosomal recessive trait. If they receive standard doses of thiopurine medications (for example, 75 mg/m2 body surface area per day), TPMT-deficient patients accumulate excessive thioguanine nucleotides in hematopoietic tissues, which leads to severe and possibly fatal myelosuppression. Objective: To elucidate the genetic basis and develop molecular methods for the diagnosis of TPMT deficiency and heterozygosity. Design: Diagnostic test evaluation. Setting: Research hospital. Patients: The TPMT phenotype was determined in 282 unrelated white persons, and TPMT genotype was determined in all persons who had intermediate TPMT activity (heterozygotes) and a randomly selected, equal number of persons who had high activity. In addition, genotype was determined in 6 TPMT-deficient patients. Measurements: Polymerase chain reaction (PCR) assays were developed to detect the G238C transversion in TPMT*2 and the G460A and A719G transitions in TPMT*3 alleles. Radiochemical assay was used to measure TPMT activity. Mutations of TPMT were identified in genomic DNA, and the concordance of TPMT genotype and phenotype was determined. Results: 21 patients who had a heterozygous phenotype were identified (7.4% of sample [95% CI, 4.7% to 11.2%]). TPMT*3A was the most prevalent mutant allele (18 of 21 mutant alleles in heterozygotes; 85%); TPMT*2 and TPMT*3C were more rare (about 5% each). All 6 patients who had TPMT deficiency had two mutant alleles, 20 of 21 patients (95% [CI, 76% to 99.9%]) who had intermediate TPMT activity had one mutant allele, and 21 of 21 patients (100% [CI, 83% to 100%]) who had high activity had no known TPMT mutation. Detection of TPMT mutations in genomic DNA by PCR coincided perfectly with genotypes detected by complementary DNA sequencing. Conclusions: The major inactivating mutations at the human TPMT locus have been identified and can be reliably detected by PCR-based methods, which show an excellent concordance between genotype and phenotype. The detection of TPMT mutations provides a molecular diagnostic method for prospectively identifying TPMT-deficient and heterozygous patients.

Original languageEnglish (US)
Pages (from-to)608-614
Number of pages7
JournalAnnals of Internal Medicine
Volume126
Issue number8
DOIs
StatePublished - Apr 15 1997

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thiopurine methyltransferase
6-Mercaptopurine
Azathioprine
Alleles
Genotype
Mutation
Thiopurine S methyltranferase deficiency
Phenotype
Thioguanine

All Science Journal Classification (ASJC) codes

  • Internal Medicine

Cite this

Molecular diagnosis of thiopurine S-methyltransferase deficiency : Genetic basis for azathioprine and mercaptopurine intolerance. / Yates, Charles; Krynetski, Eugene Y.; Loennechen, Thrina; Fessing, Michael Y.; Tai, Hung Liang; Pui, Ching Hon; Relling, Mary V.; Evans, William E.

In: Annals of Internal Medicine, Vol. 126, No. 8, 15.04.1997, p. 608-614.

Research output: Contribution to journalArticle

Yates, Charles ; Krynetski, Eugene Y. ; Loennechen, Thrina ; Fessing, Michael Y. ; Tai, Hung Liang ; Pui, Ching Hon ; Relling, Mary V. ; Evans, William E. / Molecular diagnosis of thiopurine S-methyltransferase deficiency : Genetic basis for azathioprine and mercaptopurine intolerance. In: Annals of Internal Medicine. 1997 ; Vol. 126, No. 8. pp. 608-614.
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title = "Molecular diagnosis of thiopurine S-methyltransferase deficiency: Genetic basis for azathioprine and mercaptopurine intolerance",
abstract = "Background: Thiopurine S-methyltransferase (TPMT) catalyzes the S- methylation (that is, inactivation) of mercaptopurine, azathioprine, and thioguanine and exhibits genetic polymorphism. About 10{\%} of patients have intermediate TPMT activity because of heterozygosity, and about 1 in 300 inherit TPMT deficiency as an autosomal recessive trait. If they receive standard doses of thiopurine medications (for example, 75 mg/m2 body surface area per day), TPMT-deficient patients accumulate excessive thioguanine nucleotides in hematopoietic tissues, which leads to severe and possibly fatal myelosuppression. Objective: To elucidate the genetic basis and develop molecular methods for the diagnosis of TPMT deficiency and heterozygosity. Design: Diagnostic test evaluation. Setting: Research hospital. Patients: The TPMT phenotype was determined in 282 unrelated white persons, and TPMT genotype was determined in all persons who had intermediate TPMT activity (heterozygotes) and a randomly selected, equal number of persons who had high activity. In addition, genotype was determined in 6 TPMT-deficient patients. Measurements: Polymerase chain reaction (PCR) assays were developed to detect the G238C transversion in TPMT*2 and the G460A and A719G transitions in TPMT*3 alleles. Radiochemical assay was used to measure TPMT activity. Mutations of TPMT were identified in genomic DNA, and the concordance of TPMT genotype and phenotype was determined. Results: 21 patients who had a heterozygous phenotype were identified (7.4{\%} of sample [95{\%} CI, 4.7{\%} to 11.2{\%}]). TPMT*3A was the most prevalent mutant allele (18 of 21 mutant alleles in heterozygotes; 85{\%}); TPMT*2 and TPMT*3C were more rare (about 5{\%} each). All 6 patients who had TPMT deficiency had two mutant alleles, 20 of 21 patients (95{\%} [CI, 76{\%} to 99.9{\%}]) who had intermediate TPMT activity had one mutant allele, and 21 of 21 patients (100{\%} [CI, 83{\%} to 100{\%}]) who had high activity had no known TPMT mutation. Detection of TPMT mutations in genomic DNA by PCR coincided perfectly with genotypes detected by complementary DNA sequencing. Conclusions: The major inactivating mutations at the human TPMT locus have been identified and can be reliably detected by PCR-based methods, which show an excellent concordance between genotype and phenotype. The detection of TPMT mutations provides a molecular diagnostic method for prospectively identifying TPMT-deficient and heterozygous patients.",
author = "Charles Yates and Krynetski, {Eugene Y.} and Thrina Loennechen and Fessing, {Michael Y.} and Tai, {Hung Liang} and Pui, {Ching Hon} and Relling, {Mary V.} and Evans, {William E.}",
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T1 - Molecular diagnosis of thiopurine S-methyltransferase deficiency

T2 - Genetic basis for azathioprine and mercaptopurine intolerance

AU - Yates, Charles

AU - Krynetski, Eugene Y.

AU - Loennechen, Thrina

AU - Fessing, Michael Y.

AU - Tai, Hung Liang

AU - Pui, Ching Hon

AU - Relling, Mary V.

AU - Evans, William E.

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N2 - Background: Thiopurine S-methyltransferase (TPMT) catalyzes the S- methylation (that is, inactivation) of mercaptopurine, azathioprine, and thioguanine and exhibits genetic polymorphism. About 10% of patients have intermediate TPMT activity because of heterozygosity, and about 1 in 300 inherit TPMT deficiency as an autosomal recessive trait. If they receive standard doses of thiopurine medications (for example, 75 mg/m2 body surface area per day), TPMT-deficient patients accumulate excessive thioguanine nucleotides in hematopoietic tissues, which leads to severe and possibly fatal myelosuppression. Objective: To elucidate the genetic basis and develop molecular methods for the diagnosis of TPMT deficiency and heterozygosity. Design: Diagnostic test evaluation. Setting: Research hospital. Patients: The TPMT phenotype was determined in 282 unrelated white persons, and TPMT genotype was determined in all persons who had intermediate TPMT activity (heterozygotes) and a randomly selected, equal number of persons who had high activity. In addition, genotype was determined in 6 TPMT-deficient patients. Measurements: Polymerase chain reaction (PCR) assays were developed to detect the G238C transversion in TPMT*2 and the G460A and A719G transitions in TPMT*3 alleles. Radiochemical assay was used to measure TPMT activity. Mutations of TPMT were identified in genomic DNA, and the concordance of TPMT genotype and phenotype was determined. Results: 21 patients who had a heterozygous phenotype were identified (7.4% of sample [95% CI, 4.7% to 11.2%]). TPMT*3A was the most prevalent mutant allele (18 of 21 mutant alleles in heterozygotes; 85%); TPMT*2 and TPMT*3C were more rare (about 5% each). All 6 patients who had TPMT deficiency had two mutant alleles, 20 of 21 patients (95% [CI, 76% to 99.9%]) who had intermediate TPMT activity had one mutant allele, and 21 of 21 patients (100% [CI, 83% to 100%]) who had high activity had no known TPMT mutation. Detection of TPMT mutations in genomic DNA by PCR coincided perfectly with genotypes detected by complementary DNA sequencing. Conclusions: The major inactivating mutations at the human TPMT locus have been identified and can be reliably detected by PCR-based methods, which show an excellent concordance between genotype and phenotype. The detection of TPMT mutations provides a molecular diagnostic method for prospectively identifying TPMT-deficient and heterozygous patients.

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