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Therapeutic Drug Monitoring Dec 2004This study examined thiopurine methyltransferase (TPMT) and the relationship to thioguanine nucleotides (TGN) and methylthioinosine monophosphate (meTIMP) in a large... (Comparative Study)
Comparative Study
This study examined thiopurine methyltransferase (TPMT) and the relationship to thioguanine nucleotides (TGN) and methylthioinosine monophosphate (meTIMP) in a large Swedish patient population. The current hypothesis is that the cytotoxic effects of thiopurine drugs are mediated by the incorporation of TGN into DNA. The authors assayed the TPMT activity in red blood cells from 1151 subjects and the concentrations of TGN (n = 602) and meTIMP (n = 593) from patients treated with thiopurine drugs. The TPMT frequency distribution in both adults and children showed some differences from what had been found in unselected general populations. Children had lower median TPMT activity than adults (12.0 versus 12.9 U/mL RBC; P < 0.001). Relative differences in both TGN formation [medians: normal TPMT, 1.3; intermediate TPMT, 3.3; low TPMT, 47.9 pmol/8 x 10(8) RBC per mg azathioprine (AZA); P < 0.001] and meTIMP formation (medians: normal TPMT, 13; intermediate TPMT, 7.3; low TPMT, 0 pmol/8 x 10(8) RBC per mg AZA; P = 0.001) per 1 mg administered drug were noted among the 3 TPMT activity groups. Women formed higher concentrations of both TGN (1.5 versus 1.3 pmol/8 x 10(8) RBC per mg AZA; P = 0.01) and meTIMP (14.4 versus 10.7 pmol/8 x 10(8) RBC per mg AZA; P = 0.01) than men did. There was a significant correlation between the AZA dose and the meTIMP concentrations (r = 0.45; P < 0.001). Furthermore, dose alterations made in subjects with normal TPMT (n = 84) and intermediate TPMT (n = 22) activity resulted in more pronounced increases in TGN concentrations (170 versus 30 pmol/8 x 10(8) RBC; P < 0.001) in intermediate TPMT activity, whereas in normal TPMT activity changes in meTIMP concentrations were more pronounced (1.3 versus 0 nmol/8 x 10(8) RBC; P < 0.001). In normal TPMT activity both metabolites increased in a dose-dependent fashion, whereas in intermediate TPMT activity only TGN concentrations increased. The results of this study demonstrate the dynamic nature of thiopurine metabolism and its importance for thiopurine dosing.
Topics: Adolescent; Adult; Aged; Aged, 80 and over; Azathioprine; Child; Child, Preschool; Databases, Factual; Dose-Response Relationship, Drug; Drug Monitoring; Female; Humans; Infant; Male; Methyltransferases; Middle Aged; Patient Selection; Sweden
PubMed: 15570193
DOI: 10.1097/00007691-200412000-00014 -
Oncology Research 2004The aim of this study was to examine the expression of methylthioadenosine phosphorylase (MTAP) in 21 fresh tumor samples from patients with soft tissue sarcomas (STS)... (Comparative Study)
Comparative Study
The aim of this study was to examine the expression of methylthioadenosine phosphorylase (MTAP) in 21 fresh tumor samples from patients with soft tissue sarcomas (STS) and 11 human soft tissue sarcoma cell lines, and to determine if loss of expression of this enzyme was correlated with increased sensitivity to L-alanosine and/or 6-methylmercaptopurine. We used a polyclonal antibody to measure the expression of MTAP in soft tissue sarcoma cell lines and in fresh tumor samples. Transfection of the HT-1080 cell line with a plasmid containing the cDNA for the MTAP gene was also performed to generate cell lines for in vitro and in vivo comparative sensitivity studies. MTAP was not expressed in 8 of 21 fresh STS tumors. The expression of MTAP was also not detectable in 3 of the 11 soft tissue sarcoma cell lines (HT-1080, HS42, and M-9 110). These three cell lines were more sensitive to L-alanosine, a potent inhibitor of de novo AMP synthesis, and to an inhibitor of de novo purine nucleotide synthesis, 6-methylmercaptopurine riboside (MMPR). The IC50 values for L-alanosine and MMPR were >20-fold lower in MTAP-deficient cells than in MTAP-positive cells. Restoration of MTAP into HT-1080 MTAP-deficient cells also led to decreased sensitivity to L-alanosine and MMPR. An in vivo study using HT-1080 cell tumors with and without MTAP expression confirmed that tumors lacking MTAP were more sensitive to L-alanosine than tumors expressing MTAP. These results provide the basis for selective therapy using inhibitors of de novo purine nucleotide synthesis such as L-alanosine or MMPR to treat patients with STS lacking this enzyme.
Topics: Alanine; Antibiotics, Antineoplastic; DNA, Complementary; Drug Screening Assays, Antitumor; Gene Expression Regulation, Neoplastic; Humans; Methylthioinosine; Purine-Nucleoside Phosphorylase; Sarcoma; Transfection; Tumor Cells, Cultured
PubMed: 15301428
DOI: 10.3727/0965040041292332 -
The Journal of Biological Chemistry Apr 2004Purine nucleoside phosphorylase from Plasmodium falciparum (PfPNP) is an anti-malarial target based on the activity of Immucillins. The crystal structure of... (Comparative Study)
Comparative Study
Purine nucleoside phosphorylase from Plasmodium falciparum (PfPNP) is an anti-malarial target based on the activity of Immucillins. The crystal structure of PfPNP.Immucillin-H (ImmH).SO(4) reveals a homohexamer with ImmH and SO(4) bound at each catalytic site. A solvent-filled cavity close to the 5'-hydroxyl group of ImmH suggested that PfPNP can accept additional functional groups at the 5'-carbon. Assays established 5'-methylthioinosine (MTI) as a substrate for PfPNP. MTI is not found in human metabolism. These properties of PfPNP suggest unusual purine pathways in P. falciparum and provide structural and mechanistic foundations for the design of malaria-specific transition state analogue inhibitors. 5'-Methylthio-Immucillin-H (MT-ImmH) was designed to resemble the transition state of PfPNP and binds to PfPNP and human-PNP with K(d) values of 2.7 and 303 nm, respectively, to give a discrimination factor of 112. MT-ImmH is the first inhibitor that favors PfPNP inhibition. The structure of PfPNP.MT-ImmH.SO(4) shows that the hydrophobic methylthio group inserts into a hydrophobic region adjacent to the more hydrophilic 5'-hydroxyl binding site of ImmH. The catalytic features of PfPNP indicate a dual cellular function in purine salvage and polyamine metabolism. Combined metabolic functions in a single enzyme strengthen the rationale for targeting PfPNP in anti-malarial action.
Topics: Animals; Catalysis; Catalytic Domain; Crystallography, X-Ray; Enzyme Inhibitors; Humans; Hydrophobic and Hydrophilic Interactions; Methylthioinosine; Molecular Structure; Plasmodium falciparum; Purine Nucleosides; Purine-Nucleoside Phosphorylase; Pyrimidinones; Pyrroles
PubMed: 14982926
DOI: 10.1074/jbc.C400068200 -
Inflammatory Bowel Diseases Nov 2003
Topics: Azathioprine; Child; Drug Monitoring; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Mercaptopurine; Methylthioinosine; Methyltransferases; Polymorphism, Genetic; Thionucleotides
PubMed: 14671489
DOI: 10.1097/00054725-200311000-00008 -
Inflammatory Bowel Diseases Nov 2003
Topics: Azathioprine; Child; Drug Monitoring; Guanine Nucleotides; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Mercaptopurine; Methylthioinosine; Methyltransferases; Polymorphism, Genetic; Thionucleotides
PubMed: 14671488
DOI: 10.1097/00054725-200311000-00007 -
Nucleic Acids Research Aug 2003The N6-alkyladenosines and 2-methylthio-N6-alkyladenosines are the most common modified adenosine nucleosides and transfer ribonucleic acids (tRNA) are particularly rich...
The N6-alkyladenosines and 2-methylthio-N6-alkyladenosines are the most common modified adenosine nucleosides and transfer ribonucleic acids (tRNA) are particularly rich in these modified nucleosides. They are present at position 37 of the anticodon arm and the contribution of these hypermodified nucleosides to codon-anticodon interactions, as well as translation, are significant, although not fully understood. Herein we described a new chemical synthesis method of the oligoribonucleotides containing N6-alkyladenosines and 2-methylthio-N6-alkyladenosines via post-synthetic modifications of precursor oligoribonucleotides. To obtain oligoribonucleotides containing N6-alkyladenosines, the precursor oligoribonucleotide carrying 6-methylthiopurine riboside residue was used, whereas for the synthesis of oligoribonucleotides containing 2-methylthio-N6-alkyladenosines the precursor oligoribonucleotide carrying the 2-methylthio-6-chloropurine riboside was applied. Among the modified oligoribonucleotides of different length and secondary structures, there were several containing naturally occurring modified nucleosides such as: N6-isopentenyladenosine (i6A), N6-methyladenosine (m6A), 2-methylthio-N6-isopentenyladenosine (ms2i6A), and 2-methylthio-N6-methyladenosine (ms2m6A), as well as several unnaturally modified adenosine derivatives.
Topics: Adenosine; Alkylation; Amines; Methylthioinosine; Oligonucleotides; Oligoribonucleotides; Organophosphorus Compounds; Oxidation-Reduction; Purine Nucleosides; Sulfoxides; Thionucleosides
PubMed: 12888506
DOI: 10.1093/nar/gkg632 -
Molecular Pharmacology Dec 2002Mercaptopurines have been used as anticancer agents for more than 40 years, and most acute lymphoblastic leukemias are treated with 6-mercaptopurine (6MP) or...
Mercaptopurines have been used as anticancer agents for more than 40 years, and most acute lymphoblastic leukemias are treated with 6-mercaptopurine (6MP) or 6-thioguanine (TG). Overexpression of the two related multidrug resistance proteins MRP4 and MRP5 has been shown to confer some resistance against mercaptopurines, which has been attributed to extrusion of mercaptopurine metabolites by these transporters. We have analyzed the mercaptopurine metabolites formed in human embryonic kidney cells and determined which metabolites are extruded by MRP4 and MRP5. Incubation with 6MP led to the formation of thioinosine and thioxanthosine metabolites and we found that thio-IMP was transported by both MRP4 and MRP5; MRP5 showed the highest transport rate. In contrast, only MRP5 transported thioxanthosine monophosphate (tXMP). During incubation with TG, the monophosphorylated form of thioguanosine was transported by both MRP4 and MRP5; the highest transport rate was for MRP4. Similarly, only 6-methyl-thio-IMP was formed during incubation with 6-methyl mercaptopurine riboside. This compound was a substrate for both MRP4 and MRP5; MRP4 showed the highest transport rate. Our results show that all major thiopurine monophosphates important in the efficacy of mercaptopurine treatment are transported by MRP4 and MRP5, although the substrate specificity of the two transporters differs in detail.
Topics: Biological Transport; Cells, Cultured; Chromatography, High Pressure Liquid; Drug Interactions; Humans; Kidney; Kinetics; Mercaptopurine; Methylthioinosine; Multidrug Resistance-Associated Proteins; Ribosomal Proteins; Thioguanine; Transfection
PubMed: 12435799
DOI: 10.1124/mol.62.6.1321 -
Therapeutic Drug Monitoring Jun 2002This study examined the role of thiopurine methyltransferase (TPMT) polymorphism in the metabolism and clinical effects of azathioprine and 6-mercaptopurine in the... (Clinical Trial)
Clinical Trial
Differences between children and adults in thiopurine methyltransferase activity and metabolite formation during thiopurine therapy: possible role of concomitant methotrexate.
This study examined the role of thiopurine methyltransferase (TPMT) polymorphism in the metabolism and clinical effects of azathioprine and 6-mercaptopurine in the treatment of inflammatory bowel disease and childhood leukemia. The current hypothesis is that the cytotoxic effects of thiopurines are caused by the incorporation of thioguanine nucleotides into DNA. In this context, S-methylation catalyzed by TPMT can be regarded as a competing metabolic pathway. The authors assayed the TPMT activity in red blood cells from 122 patients treated with azathioprine or 6-mercaptopurine (83 adults with inflammatory bowel disease and 39 children with acute lymphoblastic leukemia) and in 290 untreated controls (219 adult blood donors and 71 children). The concentrations of thioguanine nucleotides and methylthioinosine monophosphate were also assayed in red blood cells from the patients. The TPMT activity and the concentrations of methylthioinosine monophosphate and thioguanine nucleotides were higher in children than in adults. All children but no adult patient received concomitant methotrexate. Interaction between methotrexate and 6-mercaptopurine has been described, and may explain the results. Low TPMT activity in adult patients with inflammatory bowel disease correlated to an increased incidence of adverse drug reactions. However, there was no correlation between TPMT activity and the red blood cell concentrations of methylthioinosine monophosphate or thioguanine nucleotides, or between the concentrations of these metabolites and the occurrence of adverse effects. The results show that the role of thiopurine metabolism for drug effects is complex.
Topics: Adult; Age Factors; Antimetabolites, Antineoplastic; Azathioprine; Child; Drug Therapy, Combination; Erythrocytes; Female; Humans; Immunosuppressive Agents; Inflammatory Bowel Diseases; Male; Mercaptopurine; Methotrexate; Methyltransferases; Polymorphism, Genetic; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Thioinosine; Thionucleotides
PubMed: 12021625
DOI: 10.1097/00007691-200206000-00005 -
Acta Poloniae Pharmaceutica 2000The kinetics of hydrolysis of ribosilo-6-methylmercaptopurine was studied in aqueous solution at 353 K over a pH range of 0.45-12.13. The decomposition was followed by...
The kinetics of hydrolysis of ribosilo-6-methylmercaptopurine was studied in aqueous solution at 353 K over a pH range of 0.45-12.13. The decomposition was followed by HPLC method. The reaction of ribosilo-6-methylmercaptopurine hydrolysis includes: the reaction catalysed by hydrogen ions, that catalysed by hydroxide ions and spontaneous hydrolysis under the effect of water.
Topics: Chromatography, High Pressure Liquid; Drug Stability; Hydrolysis; Indicators and Reagents; Kinetics; Methylthioinosine; Solutions
PubMed: 11126616
DOI: No ID Found -
Cancer Research May 1999Angiogenesis has been identified as an important target for antineoplastic therapy. The use of purine analogue antimetabolites in combination chemotherapy of solid...
Angiogenesis has been identified as an important target for antineoplastic therapy. The use of purine analogue antimetabolites in combination chemotherapy of solid tumors has been proposed. To assess the possibility that selected purine analogues may affect tumor neovascularization, 6-methylmercaptopurine riboside (6-MMPR), 6-methylmercaptopurine, 2-aminopurine, and adenosine were evaluated for the capacity to inhibit angiogenesis in vitro and in vivo. 6-MMPR inhibited fibroblast growth factor-2 (FGF2)-induced proliferation and delayed the repair of mechanically wounded monolayer in endothelial GM 7373 cell cultures. 6-MMPR also inhibited the formation of solid sprouts within fibrin gel by FGF2-treated murine brain microvascular endothelial cells and the formation of capillary-like structures on Matrigel by murine aortic endothelial cells transfected with FGF2 cDNA. 6-MMPR affected FGF2-induced intracellular signaling in murine aortic endothelial cells by inhibiting the phosphorylation of extracellular signal-regulated kinase-2. The other molecules were ineffective in all of the assays. In vivo, 6-MMPR inhibited vascularization in the chick embryo chorioallantoic membrane and prevented blood vessel formation induced by human endometrial adenocarcinoma specimens grafted onto the chorioallantoic membrane. Also, topical administration of 6-MMPR caused the regression of newly formed blood vessels in the rabbit cornea. Thus, 6-MMPR specifically inhibits both the early and the late phases of the angiogenesis process in vitro and exerts a potent anti-angiogenic activity in vivo. These results provide a new rationale for the use of selected purine analogues in combination therapy of solid cancer.
Topics: 2-Aminopurine; Adenocarcinoma; Adenosine; Allantois; Animals; Antimetabolites, Antineoplastic; Antineoplastic Agents; Aorta; Brain; Calcium-Calmodulin-Dependent Protein Kinases; Cattle; Cells, Cultured; Chick Embryo; Chorion; Cornea; Depression, Chemical; Endometrial Neoplasms; Endothelium, Vascular; Female; Fibroblast Growth Factor 2; Humans; Mercaptopurine; Methylthioinosine; Mice; Mitogen-Activated Protein Kinase 1; Neoplasm Transplantation; Neovascularization, Physiologic; Phosphorylation; Protein Processing, Post-Translational; Rabbits; Recombinant Fusion Proteins; Signal Transduction; Transfection
PubMed: 10344752
DOI: No ID Found