Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.1.1.67 (thiopurine methyltransferase)
 551 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The current article describes a new assay to measure thiopurine methyltransferase (TPMT) activity from red blood cells. This method is based on the measurement of the reaction product 6-methylmercaptopurine (6-MMP) by high-performance liquid chromatography (HPLC). 6-MMP is extracted by ethyl acetate with recoveries of 85%, 80%, 80%, and 92% for 50, 250, 500, and 1,000 ng/100 microL packed red blood cells, respectively. 6-MMP was identified and measured by a Zorbax CN column installed in an HPLC system. The chromatograms were resolved using a mobile phase consisting of 40 mmol/L sodium phosphate buffer (pH 3) and methanol in a gradient from 1% to 20% of methanol. Under these conditions 6-MMP is well resolved from substrates (6-mercaptopurine and S-adenosyl-L-methionine) and endogenous peaks. When the TPMT activity from 20 patients was measured by the HPLC-linked assay and the classic radiochemical method, a linear correlation was obtained between both procedures ( y = 0.99x + 0.33; x-axis, radiochemical assay; y-axis, HPLC-linked assay; r = 0.98). In conclusion, the current report describes a new, reliable, safe, and nonradioactive method to measure TPMT activity that is shorter and simpler than the previously described ones.
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PMID:Determination of thiopurine methyltransferase activity in human erythrocytes by high-performance liquid chromatography: comparison with the radiochemical method. 1159

Genetic polymorphism of the S-methylation pathway catalyzed by thiopurine methyltransferase (TPMT) is responsible for variation in the metabolism, toxicity, and therapeutic efficacy of thiopurine drugs. This paper describe a new simple, nonradioactive HPLC method for determination of TPMT activity in isolated erythrocytes (Ery), based on the conversion of 6-mercaptopurine (pH 7.5, 37 degrees C) to 6-methylmercaptopurine (6-MMP) using S-adenosyl-l-methionine as methyl donor. The incubation step was stopped by a mixture of trichloroacetic acid/acetonitrile containing the internal standard 4-aminoacetophenone. 6-MMP was quantified by absorbance at 290 nm after chromatographic separation on a Zorbax SB-Phenyl column (5 microm, 4.6 x 250 mm) using mobile phases (flow rate 1.1 mL/min) consisting of acetonitrile, phosphate buffer pH 3.0, triethylamine, and dithiothreitol. The assay was linear up to 50 nmol/(mL Ery. h), and the detection limit was 0.3 nmol/(mL Ery. h). The extraction efficiency of 6-MMP was 95-103% (n = 3), and its analytic recovery ranged between 98.3% and 101.8% (n = 12). The within-day imprecision using pooled human erythrocytes (n = 12) was 4.4% at a TPMT activity of 14.3 nmol/(mL Ery.h) and 4.9% at 6.5 nmol/(mL Ery.h). The between-day imprecision (n = 12) was 6.8% and 7.5% nmol/(mL Ery.h), respectively. A very good agreement was found between TPMT activity determined with this method (y) and a widely used radiochemical procedure (x) (r = 0.94; n = 130; y = 0.502 + 0.946x; P < 0.05). Genotype analysis of all individuals with TPMT activity under 12.5 nmol/(mL Ery.h) revealed a genotype/phenotype concordance of 86%. The new HPLC method for determination of TPMT activity in Ery is a simple, rapid, and reliable nonradioactive procedure that can be successfully used for both research and routine clinical analysis.
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PMID:Determination of thiopurine methyltransferase phenotype in isolated human erythrocytes using a new simple nonradioactive HPLC method. 1450 88

Iatrogenic malignancies represent a devastating and often fatal long-term effect of therapy administered for a prior condition, usually a primary cancer. Earlier diagnosis and the development of more effective cancer treatments over the last 30 years have considerably improved the long-term survival of patients. However, the burgeoning number of cancer survivors has led to a parallel increase in the number of cases of iatrogenic malignancy. Consequently, understanding host susceptibility factors, such that high-risk patients can be identified, has become a priority. However, this task is made difficult by the heterogeneity of iatrogenic malignancies. Nevertheless, the identification of polymorphic loci and pathways predicted to modify dose (e.g., glutathione S-transferases, nicotinamide adenine dinucleotide phosphate: quinone oxidoreductase, cytochrome P450, and thiopurine S-methyltransferase) or determine cellular outcome (e.g., nucleotide excision DNA repair, base excision DNA repair, DNA mismatch repair, and cell death signaling) after therapy has provided insight into how host genetics may impact on the risk of developing iatrogenic malignancy.
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PMID:Genetic susceptibility to iatrogenic malignancy. 1614 1