S-methyltransferases in human intestine: differential distribution of the microsomal thiol methyltransferase and cytosolic thiopurine methyltransferase along the human bowel

Pacifici, Gian Maria; Romiti, P.; Santerini, Sabrina; Giuliani, L.

doi:10.3109/00498259309059404

Cited by 25 publications

(9 citation statements)

References 11 publications

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“…Of interest, the open reading frame of cDNA clones obtained from total liver RNA was also identical to the published wild-type colon carcinoma TPMT cDNA, which was not unexpected since TPMT activities in colon, leukocytes, and liver are correlated (25). These results confirm the authenticity of the fragment amplified and provide the initial characterization of the human liver TPMT cDNA.…”

Section: Discussionsupporting

confidence: 69%

A single point mutation leading to loss of catalytic activity in human thiopurine S-methyltransferase.

Krynetski

Schuetz

Galpin

et al. 1995

Proc. Natl. Acad. Sci. U.S.A.

294

183

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Thiopurine S-methyltransferase (TPMT; Sadenosyl-L-methionine:thiopurine S-methyltransferase, EC 2.1.1.67) activity exhibits genetic polymorphism, with -0.33% of Caucasians and African-Americans inheriting TPMT deficiency as an autosomal recessive trait. 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. 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. Sequencing of the mutant TPMT cDNA revealed a single point mutation (G238 C), leading to an amino acid substitution at codon 80 (Ala80 Pro). When assessed in a yeast heterologous expression system, this mutation led to a 100-fold reduction in TPMT catalytic activity relative to the wild-type cDNA, despite a comparable level of mRNA expression. A mutation-specific PCR amplification method was developed and used to detect the G238 -> C mutation in genomic DNA of the propositus and her mother. This inactivating mutation in the human TPMT gene provides insights into the genetic basis for this inherited polymorphism in drug metabolism.Thiopurine S-methyltransferase (TPMT; EC 2.1.1.67) is a cytoplasmic enzyme that preferentially catalyzes the Smethylation of aromatic and heterocyclic sulfhydryl compounds, including the anticancer agents 6-mercaptopurine and 6-thioguanine. TPMT activity exhibits genetic polymorphism, with -89% of Caucasians and African-Americans having high TPMT activity, -11% intermediate activity (presumed heterozygotes), and -1 in 300 inheriting TPMT deficiency as an autosomal recessive trait (1, 2). TPMT activity is typically measured in erythrocytes, as the level of TPMT activity in human liver, kidney, and normal lymphocytes has been shown to correlate with that in erythrocytes (3, 4).As part of multiagent chemotherapy for the treatment of acute lymphoblastic leukemia, mercaptopurine typically comprises a large percentage of therapy and is often administered daily throughout 2.5 years of continuation treatment. Mercaptopurine is a prodrug with no intrinsic anticancer activity, requiring intracellular conversion to thioguanine nucleotides, with subsequent incorporation into DNA, as one mechanism of its antiproliferative effects (5). Alternatively, mercaptopurine is metabolized to 6-methylmercaptopurine by TPMT or to 6-thiouric acid by xanthine oxidase. 6-Thiouric acid is an inactive metabolite, whereas 6-methylmercaptopurine nucleotide inhibits phosphoribosyl diphosphate amidotransferase, an enzyme catalyzing the first step in de novo purine synthesis (6, 7). It is not clear whether mercaptopurine's principal mechanism of cytotoxicity is via incorporation of thioguanine nucleotides into DNA and RNA, or via 6-methylmercaptopurine nucleotide inhibition of de novo purine synthesis, or a combination of these effects.Clinical studies in children with acute lymphoblastic l...

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Section: Discussionsupporting

confidence: 69%

A single point mutation leading to loss of catalytic activity in human thiopurine S-methyltransferase.

Krynetski

Schuetz

Galpin

et al. 1995

Proc. Natl. Acad. Sci. U.S.A.

294

183

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“…TMT activity in the colonic mucosa has been reported as low21 or high,29 30 with more recent studies28 intermediary to these values. The method we used to assay TMT activity has previously been validated with regard to reproducibility and coefficient of variance 28.…”

Section: Discussionmentioning

confidence: 94%

“…The method we used to assay TMT activity has previously been validated with regard to reproducibility and coefficient of variance 28. As the activity of TMT may vary along the length of the intestinal tract29 30 a fixed region for biopsy was chosen in which colitis usually manifests. Also of concern was whether biopsies, a composite tissue of immune and epithelial cells, would yield TMT activities reflective of isolated epithelial cells 28.…”

Section: Discussionmentioning

confidence: 99%

Thiol methyltransferase activity in inflammatory bowel disease

Roediger

2000

Gut

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Background-Luminal anionic sulphide may contribute to epithelial damage in ulcerative colitis. Thiol methyltransferase (TMT) governs sulphide detoxification by the colonic mucosa and circulating erythrocytes. Aims-To measure levels of TMT activity in erythrocytes of surgically treated cases of colitis or in rectal biopsies of defined groups of colitis. Patients-Venepuncture blood was obtained from 37 blood donors and 27 subjects who had previously undergone a proctocolectomy for colitis: 18 for ulcerative colitis and nine for Crohn's colitis. Rectal biopsies from 122 cases were obtained: 47 without mucosal disease, 33 post-colon resection for cancer, 14 with moderate to severe ulcerative colitis, 15 with quiescent ulcerative colitis, seven with acute Crohn's colitis, and six with radiation proctitis. Methods-TMT activity was measured by high performance liquid chromatography with radioactive detection to measure 14 C methylmercaptoethanol formation, the reaction product of cell extracts incubated with mercaptoethanol and 14 C S-adenosylmethionine. Results-Erythrocyte TMT activity of surgically treated cases of colitis was significantly elevated (p<0.001) compared with control cases. TMT activity of rectal biopsies was significantly decreased (p<0.02) in acute but not quiescent ulcerative colitis, Crohn's colitis, or radiation colitis. Conclusions-Erythrocyte TMT activity was persistently elevated after proctocolectomy for Crohn's disease and ulcerative colitis. No primary defect of TMT activity was found in any case of unoperated colitis but mucosal activity was diminished with disease progression of ulcerative colitis. Studies of genetic control of TMT activity of erythrocytes in inflammatory bowel disease appear worthwhile.

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“…The activity of TMT was measured using a method based on that of Pacifici et al [15] and Weinshilboum et al [16] using 2-mercaptoethanol as substrate. Rectal biopsy homogenates or erythrocyte lysates were incubated with 100 mM 2-mercaptoethanol, 100 µM 3 H-S-adenosyl methionine (500 mCi/mmol; Amersham, UK) in 0.1 M potassium phosphate buffer with 1 mM EDTA and 0.5% Triton X-100 (pH 7.9) at 37 • C for 10 min in a 25-µl final volume.…”

Section: Tmt Assaymentioning

confidence: 99%

Impaired Detoxication of Hydrogen Sulfide in Ulcerative Colitis?

et al. 2007

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Impaired butyrate oxidation and raised counts of sulfate-reducing bacteria in the colon of patients with ulcerative colitis (UC) indicate that the disease may be induced or aggravated by hydrogen sulfide toxicity. We aimed to examine enzymatic removal of H(2)S in erythrocytes and colonic mucosa from controls and patients with UC and Crohn's disease (CD). Rhodanese (RHOD) and thiol methyltransferase (TMT) activities were measured in rectal mucosa and erythrocytes, and plasma thiocyanate was determined. Four groups were analyzed: patients with UC, patients with CD, hospital controls (patients with dyspepsia or IBS), and a group of healthy volunteers. RHOD and TMT activity in rectal biopsies did not differ significantly between controls and patients with UC or CD (n=56). Control levels of RHOD were significantly higher in men than in women (212+/-25 and 132+/-14 nmol/mg/min, respectively; P<0.01). In erythrocytes (n=128) RHOD activity was significantly higher in UC patients than in hospital or volunteer controls (1.15+/-0.12 compared with 0.88+/-0.12 and 0.66+/-0.02 nmol/mg/min; P<0.05 and P<0.02, respectively). TMT activity was also significantly higher in erythrocytes from UC patients and hospital controls than volunteer controls (2.02+/-0.13 pmol/mg/min [P<0.001], 1.51+/-0.21 pmol/mg/min [P<0.05], and 1.17+/-0.18 pmol/mg/min, respectively). We found no evidence of defective enzymic detoxication of sulfide by RHOD or TMT in patients with UC or CD.

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S-methyltransferases in human intestine: differential distribution of the microsomal thiol methyltransferase and cytosolic thiopurine methyltransferase along the human bowel

Cited by 25 publications

References 11 publications

A single point mutation leading to loss of catalytic activity in human thiopurine S-methyltransferase.

A single point mutation leading to loss of catalytic activity in human thiopurine S-methyltransferase.

Thiol methyltransferase activity in inflammatory bowel disease

Impaired Detoxication of Hydrogen Sulfide in Ulcerative Colitis?

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