Disease relevance of TPMT

• The autosomal recessive trait of thiopurine S-methytransferase (TPMT) deficiency is associated with severe hematopoietic toxicity when patients are treated with standard doses of mercaptopurine, azathioprine, or thioguanine [1].
• RESULTS: 27 patients completed the study per protocol, while 33 were withdrawn (early protocol violation (n = 5), TPMT deficiency (n = 1), thiopurine related adverse events (n = 27)); 67% of patients with adverse events tolerated long term treatment on a lower dose (median 1.32 mg azathioprine/kg body weight) [2].
• The ability of plasma from individual uremic patients to inhibit TPMT also correlated with its ability to inhibit two other drug metabolizing methyltransferases in the RBC, catechol-O-methyltransferase and phenol-O-methyltransferase, RBC TMT activity is not altered in patients with uremia [3].
• There are large individual variations in inhibition of RBC TPMT by plasma from patients with renal failure [3].
• METHODS: Seventy-two azathioprine treated paediatric inflammatory bowel disease patients, 47% girls, mean age 12.5 years (range 6.5-17.5), were assessed for TPMT and ITPA polymorphisms and for adverse events [4].
• Fifty patients with inflammatory bowel disease with normal TPMT activity were all homozygous for six GCC repeats [(GCC)6] [5].

Psychiatry related information on TPMT

• A genetic polymorphism regulating TPMT activity in human tissue is an important factor responsible for individual differences in the toxicity and therapeutic efficacy of these drugs [6].
• CONCLUSION: No impact of child development on TPMT activity could be evidenced, suggesting that TPMT activity is already mature at birth [7].
• Prospective, randomized, controlled trials are needed to determine whether prior TPMT phenotype testing can be used to adjust the dose of AZA effectively to improve clinical response time and rate [8].

High impact information on TPMT

• Leukemia cells that had acquired an additional chromosome containing a wild-type TPMT or GGH allele had significantly lower accumulation of thioguanine nucleotides or methotrexate polyglutamates, respectively [9].
• Drugs (and enzymes) for which there is a reasonable evidence base supporting genotyping or phenotyping include suxamethonium/mivacurium (butyrylcholinesterase), and azathioprine/6-mercaptopurine (thiopurine methyltransferase) [10].
• Phenotyping for the thiopurine methyltransferase genetic polymorphism represents one of the first examples in which testing for a pharmacogenetic variant has entered standard clinical practice [11].
• We studied the metabolism, dose requirements, and tolerance of 6-mercaptopurine among patients with different TPMT phenotypes [12].
• CONCLUSION: We conclude that genetic polymorphism in TPMT is an important determinant of mercaptopurine toxicity, even among patients who are heterozygous for this trait [12].

Chemical compound and disease context of TPMT

• To determine the molecular genetic basis for this polymorphism, we cloned the TPMT cDNA from a TPMT-deficient patient who had developed severe hematopoietic toxicity during mercaptopurine therapy [13].
• Deficiency of purine enzymes, including TPMT and 5NT, may be a cause of AZA-related bone marrow toxicity in patients with RA [14].
• The drug-metabolizing enzyme thiopurine S-methyltransferase (TPMT) catalyzes the S-methylation of thiopurines such as 6-mercaptopurine, 6-thioguanine, and azathiopurine, which are used as immunosuppressants and in the treatment of acute lymphoblastic leukemia and rheumatoid arthritis [15].
• Polymorphisms at three loci in the thiopurine methyltransferase (TPMT) gene are known to be responsible for azathioprine and 6-mercaptopurine (6MP) toxicity [16].
• Using a bacterial orthologue of TPMT from Pseudomonas syringae, we have used NMR spectroscopy to describe the consequences of binding sinefungin, a SAM analogue, on the structure and dynamics of the TPMT protein backbone [17].
• Advanced fibrosis but not TPMT genotype or activity predicts azathioprine toxicity in AIH [18].

Biological context of TPMT

• In addition, genotype was determined in 6 TPMT-deficient patients [19].
• 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 [19].
• TPMT genetic polymorphisms represent a striking example of the potential clinical value of pharmacogenetics [20].
• Furthermore, there was a correlation among TPMT half-life values in rabbit reticulocyte lysate, aggresome formation in COS-1 cells, and protein aggregation in vitro for the three variant allozymes encoded by alleles that include the two TPMT*3A single-nucleotide polymorphisms [20].
• Sequencing of the mutant TPMT cDNA revealed a single point mutation (G238-->C), leading to an amino acid substitution at codon 80 (Ala80-->Pro) [13].

Anatomical context of TPMT

• In the present study, we established that erythrocyte TPMT activity was significantly related to the amount of TPMT protein on Western blots of erythrocytes from patients with TPMT activities of 0.4-23 units/ml pRBC (rs = 0.99; P < 0.001) [21].
• Northern blot analysis of RNA isolated from leukocytes of the deficient patient demonstrated the presence of TPMT mRNAs of comparable size to that in subjects with high TPMT activity [13].
• Two patients with AZA-related bone marrow toxicity were found to have a TPMT deficiency, 1 partial and 1 total [14].
• The TPMT cDNA ORF was then used to screen a human lymphocytes genomic DNA library in an attempt to clone the TPMT gene(s) in humans [22].
• Therefore, it was important to clone a TPMT cDNA from a human drug-metabolizing organ such as the liver to determine whether its sequence matched that of the cDNA cloned from the T84 cell line [22].

Associations of TPMT with chemical compounds

• 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 [19].
• TPMT is a cytosolic enzyme that catalyzes the S-methylation of aromatic and heterocyclic sulfhydryl compounds, including medications such as mercaptopurine and thioguanine [21].
• The plasma of uremic patients reversibly inhibits RBC TPMT activity to a greater extent than normal plasma does and contains higher concentrations of endogenous methyl acceptors than normal plasma [3].
• The first pharmacogenetic trait identified was monogenic, and concerned the prototypic example of thiopurine methyltransferase (TPMT) implicated in azathioprine metabolism [23].
• Geldanamycin treatment of COS-1 cells transfected with FLAG-tagged wild-type also resulted in a time and geldanamycin concentration-dependent decrease in TPMT activity and protein, which was compatible with results obtained in the RRL [24].
• Leukocyte genetic TPMT testing revealed that the patient had homozygous polymorphisms associated with the absence of TPMT activity and severe azathioprine-induced myelotoxicity [25].

Regulatory relationships of TPMT

• Genetic variation in the MTHFR gene influences thiopurine methyltransferase activity [26].
• Both mercaptopurine and its prodrug azathioprine are metabolised to active compounds (6-thioguanine nucleotides; 6-TGN) by hypoxanthine-guanine phosphoribosyltransferase and to inactive metabolites by the polymorphically expressed thiopurine S-methyltransferase (TPMT) and xanthine oxidase [27].

Other interactions of TPMT

• Candidate gene approaches have led to clinical tests for toxicity avoidance (eg, TPMT, UGT1A1) and efficacy prediction (eg, epidermal growth factor receptor-activating mutations) [28].
• Reduced thiopurine methyltransferase activity and development of side effects of azathioprine treatment in patients with rheumatoid arthritis [29].
• Do ITPA and TPMT genotypes predict the development of side effects to AZA [30]?
• In conclusion, our findings suggest that TPMT, ITPA, and MTHFR genotypes do not predict adverse drug reactions, including bone marrow suppression, in liver transplant patients [31].
• Only few examples, such as TPMT and CYP2C9 genetic polymorphisms, can illustrate this rare situation [32].

Analytical, diagnostic and therapeutic context of TPMT

• Detection of TPMT mutations in genomic DNA by PCR coincided perfectly with genotypes detected by complementary DNA sequencing [19].
• TPMT genotype was determined in all individuals with TPMT activity indicative of a heterozygous genotype (</=10.1 U/ml pRBC, n = 23African-Americans, n = 21 Caucasians) and a control group with TPMT activity indicative of a homozygous wild-type genotype (>10.2 U/ml pRBC, n = 23 African-Americans, n = 21 Caucasians) [33].
• RBC TPMT activity is not affected by hemodialysis [3].
• Characterization of variant alleles for low TPMT enzyme activity will help make it possible to assess the potential clinical utility of deoxyribonucleic acid-based diagnostic tests for determining TPMT genotype [34].
• Analysis of recent data suggests that by optimizing the AZA dose on the basis of TPMT status testing (with a substantial reduction in dose for patients homozygous for mutant TPMT alleles), a reduction in drug-induced morbidity and cost savings can be made by avoiding hospitalization and rescue therapy for leucopenic events [35].

References