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Cell Reports Apr 2022The innate immune recognition of the malaria-causing pathogen Plasmodium falciparum (P. falciparum) is not fully explored. Here, we identify the nucleoside...
The innate immune recognition of the malaria-causing pathogen Plasmodium falciparum (P. falciparum) is not fully explored. Here, we identify the nucleoside 5'-methylthioinosine (MTI), a Plasmodium-specific intermediate of the purine salvage pathway, as a pathogen-derived Toll-like receptor 8 (TLR8) agonist. Co-incubation of MTI with the TLR8 enhancer poly(dT) as well as synthetic or P. falciparum-derived RNA strongly increase its stimulatory activity. Of note, MTI generated from methylthioadenosine (MTA) by P. falciparum lysates activates TLR8 when MTI metabolism is inhibited by immucillin targeting the purine nucleoside phosphorylase (PfPNP). Importantly, P. falciparum-infected red blood cells incubated with MTI or cultivated with MTA and immucillin lead to TLR8-dependent interleukin-6 (IL-6) production in human monocytes. Our data demonstrate that the nucleoside MTI is a natural human TLR8 ligand with possible in vivo relevance for innate sensing of P. falciparum.
Topics: Humans; Malaria, Falciparum; Methylthioinosine; Nucleosides; Plasmodium falciparum; Purine-Nucleoside Phosphorylase; Purines; Toll-Like Receptor 8
PubMed: 35417716
DOI: 10.1016/j.celrep.2022.110691 -
Biomedicine & Pharmacotherapy =... Dec 2018Colon cancer is one of the most common digestive malignant tumors that leads to high mortality worldwide, and metastasis is the primary cause of cancer-related death. It...
Colon cancer is one of the most common digestive malignant tumors that leads to high mortality worldwide, and metastasis is the primary cause of cancer-related death. It is well accepted that the epithelial-mesenchymal transition (EMT) plays a key role in the process of metastasis. As a cytokine that macrophage secretes, IL-6 is involved in the progression of tumors, including the invasion and metastasis via kinds of signaling pathways. However, the mechanism of interactions between IL-6, macrophage, EMT and colon cancer is not fully understood. Increased CD68 macrophages and IL-6 level were found in colon tumor as compared to normal colon tissue. Metastatic lymph node showed even more CD68 macrophages and higher IL-6 level than the primary tumor. These results suggested that macrophages and IL-6 play an important role in EMT of colon cancer. In order to investigate the effect of macrophage and IL-6 on EMT of colon cancer, we cultured human colon carcinoma cell line SW48 with conditioned medium (CM) from PMA-stimulated monocyte THP-1 cells and tested for IL-6 dependent EMT pathways. Wound healing assay and Transwell assay were used to analyze cell migration and invasion. Results showed that CM-treated SW48 cells increased IL-6 production and displayed elevated capacity of migration and invasion compared to untreated cells. Increased expressions of EMT markers (N-cadherin, Vimentin and β-catenin) and decreased expression of EMT marker(E-cadherin) were found in CM-treated SW48 cells by Western Blot. The addition of an anti-IL-6 antibody significantly inhibited the increase of EMT markers (Vimentin and β-catenin) as well as cell migration and invasion, suggesting that IL-6 played a critical role in promoting EMT of CM-treated SW48 cells. In addition, we found that the levels of p-STAT3 and p-ERK increased in CM-treated SW48 compared to untreated cells, which can be reversed by AG490, an inhibitor of JAK. In the meantime, the suppression of JAK-associated signaling pathways caused a decrease of β-catenin. In summary, our study suggested that macrophage-induced IL-6 promotes migration and invasion of colon cancer cell via Wnt/β-catenin pathway in STAT3/ERK-dependent way.
Topics: 6-Aminonicotinamide; Antineoplastic Combined Chemotherapy Protocols; Aspartic Acid; Cell Line, Tumor; Cell Movement; Colonic Neoplasms; Epithelial-Mesenchymal Transition; Humans; Interleukin-6; MAP Kinase Signaling System; Macrophages; Methylthioinosine; STAT3 Transcription Factor; THP-1 Cells; Wnt Signaling Pathway; beta Catenin
PubMed: 30243096
DOI: 10.1016/j.biopha.2018.09.067 -
Biochemistry Feb 2011The PA3004 gene of Pseudomonas aeruginosa PAO1 was originally annotated as a 5'-methylthioadenosine phosphorylase (MTAP). However, the PA3004 encoded protein uses...
The PA3004 gene of Pseudomonas aeruginosa PAO1 was originally annotated as a 5'-methylthioadenosine phosphorylase (MTAP). However, the PA3004 encoded protein uses 5'-methylthioinosine (MTI) as a preferred substrate and represents the only known example of a specific MTI phosphorylase (MTIP). MTIP does not utilize 5'-methylthioadenosine (MTA). Inosine is a weak substrate with a k(cat)/K(m) value 290-fold less than MTI and is the second best substrate identified. The crystal structure of P. aeruginosa MTIP (PaMTIP) in complex with hypoxanthine was determined to 2.8 Å resolution and revealed a 3-fold symmetric homotrimer. The methylthioribose and phosphate binding regions of PaMTIP are similar to MTAPs, and the purine binding region is similar to that of purine nucleoside phosphorylases (PNPs). The catabolism of MTA in P. aeruginosa involves deamination to MTI and phosphorolysis to hypoxanthine (MTA → MTI → hypoxanthine). This pathway also exists in Plasmodium falciparum, where the purine nucleoside phosphorylase (PfPNP) acts on both inosine and MTI. Three tight-binding transition state analogue inhibitors of PaMTIP are identified with dissociation constants in the picomolar range. Inhibitor specificity suggests an early dissociative transition state for PaMTIP. Quorum sensing molecules are associated with MTA metabolism in bacterial pathogens suggesting PaMTIP as a potential therapeutic target.
Topics: Amino Acid Sequence; Crystallography, X-Ray; Methylthioinosine; Models, Biological; Models, Molecular; Molecular Sequence Annotation; Molecular Sequence Data; Phosphorylases; Pseudomonas aeruginosa; Quorum Sensing; Sequence Homology, Amino Acid; Structure-Activity Relationship
PubMed: 21197954
DOI: 10.1021/bi101642d -
Journal of Pharmaceutical and... Aug 2022Mercaptopurine (6-MP) is an indispensable, first-line, drug in the treatment of pediatric acute lymphoblastic leukemia (ALL). However, 6-MP has several intrinsic...
Mercaptopurine (6-MP) is an indispensable, first-line, drug in the treatment of pediatric acute lymphoblastic leukemia (ALL). However, 6-MP has several intrinsic drawbacks, such as large individual variability in the drug response, undesirable adverse reactions, and drug resistance in patients with release ALL, which requires therapeutic drug monitoring (TDM). Several studies analyzed the total concentration of thiopurine nucleotides in red blood cells (RBCs) after hydrolysis, and two studies detected them separately and accurately by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In this study, we developed a rapid and robust LC-MS/MS method for simultaneous quantitation of mono-, di-, and triphosphates of thioguanosine and methylthioinosine. Not only EDTA and DTT were added, but also EHT1864, a new Rac family small GTPases inhibitor, was innovatively added to ensure the stability of the analytes. Commercial availability and relatively low cost compound methotrexate-D3 was selected as internal standards. The linearity, accuracy, precision, recovery, matrix effect and stability of the method were all in line with the guidelines. This method provide an accurate and robust new solution for the determination of 6 metabolites of MP in RBCs from ALL patients with maintenance therapy.
Topics: Child; Chromatography, Liquid; Humans; Mercaptopurine; Methylthioinosine; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Tandem Mass Spectrometry
PubMed: 35550492
DOI: 10.1016/j.jpba.2022.114813 -
PloS One 2014Plasmodium parasites rely upon purine salvage for survival. Plasmodium purine nucleoside phosphorylase is part of the streamlined Plasmodium purine salvage pathway that...
Plasmodium parasites rely upon purine salvage for survival. Plasmodium purine nucleoside phosphorylase is part of the streamlined Plasmodium purine salvage pathway that leads to the phosphorylysis of both purines and 5'-methylthiopurines, byproducts of polyamine synthesis. We have explored structural features in Plasmodium falciparum purine nucleoside phosphorylase (PfPNP) that affect efficiency of catalysis as well as those that make it suitable for dual specificity. We used site directed mutagenesis to identify residues critical for PfPNP catalytic activity as well as critical residues within a hydrophobic pocket required for accommodation of the 5'-methylthio group. Kinetic analysis data shows that several mutants had disrupted binding of the 5'-methylthio group while retaining activity for inosine. A triple PfPNP mutant that mimics Toxoplasma gondii PNP had significant loss of 5'-methylthio activity with retention of inosine activity. Crystallographic investigation of the triple mutant PfPNP with Tyr160Phe, Val66Ile, andVal73Ile in complex with the transition state inhibitor immucillin H reveals fewer hydrogen bond interactions for the inhibitor in the hydrophobic pocket.
Topics: Amino Acid Sequence; Catalytic Domain; Kinetics; Methylthioinosine; Models, Molecular; Molecular Sequence Data; Mutagenesis, Site-Directed; Mutation; Plasmodium falciparum; Purine-Nucleoside Phosphorylase; Pyrimidinones; Structure-Activity Relationship; Substrate Specificity
PubMed: 24416224
DOI: 10.1371/journal.pone.0084384 -
Faraday Discussions Apr 2018S6-Methylthioinosine and O6-methylguanosine are byproducts resulting from the enzymatic reactions of sulfur-substituted prodrugs in cells and from the interaction of...
S6-Methylthioinosine and O6-methylguanosine are byproducts resulting from the enzymatic reactions of sulfur-substituted prodrugs in cells and from the interaction of alkylating agents with cellular DNA, respectively. Their photochemistry has not been investigated, and it is currently unknown whether light absorption by these byproducts may pose any threat to the cell. In this contribution, their photoinduced processes upon absorption of UVB radiation are reported using broadband transient absorption spectroscopy. Plausible electronic relaxation mechanisms are proposed for both biological molecules, which are supported by steady-state absorption and emission measurements, and by singlet and triplet vertical excitation energies performed on a large subset of ground-state optimized conformational isomers in solution. The results are compared to the body of knowledge gathered in the scientific literature about the light-induced processes in the sulfur-substituted and canonical purine monomers. In particular, it is shown that S6-methylation decreases the rate to populate the lowest-energy triplet state and blueshifts the ground-state absorption spectrum compared to those for the sulfur-substituted prodrugs and for the 6-thioguanosine metabolite. Similarly, O6-methylation decreases the rate of internal conversion to the ground state observed in the guanine monomers by more than 10-fold in acetonitrile and 40-fold in aqueous solution, while it redshifts the ground-state absorption spectrum. Collectively, this investigation provides relevant new insights about the relationship between structural modifications of the purine chromophore and the electronic relaxation mechanisms in this important group of biological molecules.
Topics: Guanosine; Methylthioinosine; Photochemical Processes; Solutions; Spectrophotometry, Ultraviolet
PubMed: 29372193
DOI: 10.1039/c7fd00193b -
Biochemical and Biophysical Research... Apr 1983Viable human and murine lymphoblasts, and normal human tissue extracts, converted the thioether nucleosides 5'-methylthioadenosine (MeSAdo) and 5'-methylthioinosine...
Viable human and murine lymphoblasts, and normal human tissue extracts, converted the thioether nucleosides 5'-methylthioadenosine (MeSAdo) and 5'-methylthioinosine (MeSIno) to methionine. Both MeSAdo and MeSIno, but not homocysteine, supported the short-term growth of human or murine lymphoblasts in methionine deficient medium. However, MeSAdo at concentrations greater than 25 microM inhibited cell growth. MeSIno was non-toxic at concentrations up to 200 microM, and supported the long-term growth of lymphoblasts in methionine-free medium.
Topics: Adenosine; Animals; B-Lymphocytes; Cell Division; Cell Line; Deoxyadenosines; Humans; Inosine; Leukemia L1210; Methionine; Methylthioinosine; Mice; Thionucleosides
PubMed: 6601949
DOI: 10.1016/0006-291x(83)91476-6 -
Marine Drugs Feb 2014A new actinomycete strain Micromonospora sp. K310 was isolated from Ghanaian mangrove river sediment. Spectroscopy-guided fractionation led to the isolation of two new... (Comparative Study)
Comparative Study
A new actinomycete strain Micromonospora sp. K310 was isolated from Ghanaian mangrove river sediment. Spectroscopy-guided fractionation led to the isolation of two new compounds from the fermentation culture. One of the compounds is butremycin (2) which is the (3-hydroxyl) derivative of the known Streptomyces metabolite ikarugamycin (1) and the other compound is a protonated aromatic tautomer of 5'-methylthioinosine (MTI) (3). Both new compounds were characterized by 1D, 2D NMR and MS data. Butremycin (2) displayed weak antibacterial activity against Gram-positive S. aureus ATCC 25923, the Gram-negative E. coli ATCC 25922 and a panel of clinical isolates of methicillin-resistant S. aureus (MRSA) strains while 3 did not show any antibacterial activity against these microbes.
Topics: Anti-Bacterial Agents; Escherichia coli; Fermentation; Geologic Sediments; Ghana; Lactams, Macrocyclic; Magnetic Resonance Spectroscopy; Mass Spectrometry; Methicillin-Resistant Staphylococcus aureus; Methylthioinosine; Microbial Sensitivity Tests; Micromonospora; Rivers; Staphylococcus aureus
PubMed: 24534843
DOI: 10.3390/md12020999 -
Cancer Mar 1980This review summarizes a body of information suggesting that proper metabolic modulation with certain metabolites can sensitize tumor cells to anti-metabolites, and... (Clinical Trial)
Clinical Trial Comparative Study Review
This review summarizes a body of information suggesting that proper metabolic modulation with certain metabolites can sensitize tumor cells to anti-metabolites, and others can de-sensitize (i.e. protect) normal cells from the toxicity of anti-metabolites. This new approach offers the possibility of increasing the selectivity of drug therapy, with the promise of a real advance in cancer chemotherapy. The metabolite thymidine (TdR), long used as a cell synchronizing agent, is known to exert this effect in vitro by metabolic modulation of a number of enzymes in the salvage pathway to DNA synthesis. Against this biochemical background, in vivo effects of TdR employed as an agent for cancer therapy are reviewed as follows: 1) TdR alone, and in combination with, 2) Methotrexate (MTX), or 3) 5-Fluorouracil (FU), or 4) Cytosine arabinoside (ara-C). TdR is shown in all instances either to protect against host toxicity (eg. MTX), or to potentiate the anti-tumor effect (eg. FU and ara-C). Findings are also presented that a sequential schedule of MTX prior to TdR prior to FU is important for the optimal therapeutic activity of these drugs. The biochemical basis for the MTX leads to FU augmentation is reportedly due to increased activation of FU by MTX (acting indirectely). On the basis of this biochemical insight, a completely different chemotherapeutic agent methyl-mercaptopurine raboside (MMPR) was substituted for MTX, resulting in a dramatic potentiation of anticancer activity. Metabolic modulation with still other metabolites (UR) and a hormone (testosterone) was demonstrated to protect from host toxicity due to certain anti-cancer agents without offsetting anti-tumor activity. The ability to prevent leukopenia by these means was particularly impressive. Clinical trials have been initiated with TdR alone, TdR + MTX, and TdR + FU; the available clinical data are summarized.
Topics: Animals; Antineoplastic Agents; Bone Marrow; Clinical Trials as Topic; Cytarabine; DNA, Neoplasm; Drug Therapy, Combination; Fluorouracil; Humans; Methotrexate; Methylthioinosine; Mice; Neoplasms; Thymidine
PubMed: 6986974
DOI: 10.1002/1097-0142(19800315)45:5+<1117::aid-cncr2820451316>3.0.co;2-s -
The Journal of Biological Chemistry Jan 1984Methylthioinosine (MeSno) is a purine nucleoside analog which is cytotoxic to a number of cultured cell lines including the Reuber H35 hepatoma cells used in the present...
Methylthioinosine (MeSno) is a purine nucleoside analog which is cytotoxic to a number of cultured cell lines including the Reuber H35 hepatoma cells used in the present studies. It has also been observed to cause a rapid profound loss of tyrosine aminotransferase activity in H35 cells well before the onset of any measurable cytotoxicity. The effect is both time and concentration dependent. MeSno does not acutely inhibit synthesis of the enzyme as evidenced by the ability of glucocorticoids or cAMP analogs to induce the enzyme to the same extent in the presence or absence of the drug. The enzyme in extracts of cells treated with the drug is essentially identical with the enzyme from extracts of control cells in terms of thermal stability, immunoprecipitability, and affinities for substrates and cofactor. Addition of MeSno to cell extracts and mixing experiments suggests that the thiopurine does not have any direct effect on enzyme activity. Immunochemical analysis of the rates of synthesis and degradation of the aminotransferase have shown that the enzyme is degraded approximately 3-4 times more rapidly in cells treated with the drug than in control cells. At the same time there is no inhibition of the rate of synthesis of the enzyme.
Topics: Animals; Dexamethasone; Dose-Response Relationship, Drug; Hot Temperature; Inosine; Liver Neoplasms, Experimental; Methylthioinosine; Rats; Tyrosine Transaminase
PubMed: 6141163
DOI: No ID Found