Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

A Chimeric ATP-Linked Nucleotide Enables Luminescence Signaling of Damage Surveillance by MTH1, a Cancer Target.

ABSTRACT: The enzyme MTH1 cleanses the cellular nucleotide pool of oxidatively damaged 8-oxo-dGTP, preventing mutagenesis by this nucleotide. The enzyme is considered a promising therapeutic target; however, methods to measure its activity are indirect and laborious and have low sensitivity. Here we describe a novel ATP-linked chimeric nucleotide (ARGO) that enables luminescence signaling of the enzymatic reaction, greatly simplifying the measurement of MTH1 activity. We show that the reporting system can be used to identify inhibitors of MTH1, and we use it to quantify enzyme activity in eight cell lines and in colorectal tumor tissue. The ARGO reporter is likely to have considerable utility in the study of the biology of MTH1 and potentially in analyzing patient samples during clinical testing.

SUBMITTER: Ji D

PROVIDER: S-EPMC5500214 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

A Chimeric ATP-Linked Nucleotide Enables Luminescence Signaling of Damage Surveillance by MTH1, a Cancer Target.

Ji Debin D Beharry Andrew A AA Ford James M JM Kool Eric T ET

Journal of the American Chemical Society 20160718 29

The enzyme MTH1 cleanses the cellular nucleotide pool of oxidatively damaged 8-oxo-dGTP, preventing mutagenesis by this nucleotide. The enzyme is considered a promising therapeutic target; however, methods to measure its activity are indirect and laborious and have low sensitivity. Here we describe a novel ATP-linked chimeric nucleotide (ARGO) that enables luminescence signaling of the enzymatic reaction, greatly simplifying the measurement of MTH1 activity. We show that the reporting system can ...[more]

PMID: 27413803

Similar Datasets

ATP-Linked Chimeric Nucleotide as a Specific Luminescence Reporter of Deoxyuridine Triphosphatase.

Project description:Nucleotide surveillance enzymes play important roles in human health, by monitoring damaged monomers in the nucleotide pool and deactivating them before they are incorporated into chromosomal DNA or disrupt nucleotide metabolism. In particular, deamination of cytosine, leading to uracil in DNA and in the nucleotide pool, can be deleterious, causing DNA damage. The enzyme deoxyuridine triphosphatase (dUTPase) is currently under study as a therapeutic and prognostic target for cancer. Measuring the activity of this enzyme is important both in basic research and in clinical applications involving this pathway, but current methods are nonselective, detecting pyrophosphate, which is produced by many enzymes. Here we describe the design and synthesis of a dUTPase enzyme-specific chimeric dinucleotide (DUAL) that replaces the pyrophosphate leaving group of the native substrate with ATP, enabling sensitive detection via luciferase luminescence signaling. The DUAL probe functions sensitively and selectively to quantify enzyme activities in vitro and in cell lysates. We further report the first measurements of dUTPase activities in eight different cell lines, which are found to vary by a factor of 7-fold. We expect that the new probe can be of considerable utility in research involving this clinically significant enzyme.

| S-EPMC6484856 | biostudies-literature

Measuring deaminated nucleotide surveillance enzyme ITPA activity with an ATP-releasing nucleotide chimera.

Project description:Nucleotide quality surveillance enzymes play important roles in human health, by detecting damaged molecules in the nucleotide pool and deactivating them before they are incorporated into chromosomal DNA or adversely affect metabolism. In particular, deamination of adenine moiety in (deoxy)nucleoside triphosphates, resulting in formation of (d)ITP, can be deleterious, leading to DNA damage, mutagenesis and other harmful cellular effects. The 21.5 kDa human enzyme that mitigates this damage by conversion of (d)ITP to monophosphate, ITPA, has been proposed as a possible therapeutic and diagnostic target for multiple diseases. Measuring the activity of this enzyme is useful both in basic research and in clinical applications involving this pathway, but current methods are nonselective and are not applicable to measurement of the enzyme from cells or tissues. Here, we describe the design and synthesis of an ITPA-specific chimeric dinucleotide (DIAL) that replaces the pyrophosphate leaving group of the native substrate with adenosine triphosphate, enabling sensitive detection via luciferase luminescence signaling. The probe is shown to function sensitively and selectively to quantify enzyme activity in vitro, and can be used to measure the activity of ITPA in bacterial, yeast and human cell lysates.

| S-EPMC5714213 | biostudies-literature

Birth of MTH1 as a therapeutic target for glioblastoma: MTH1 is indispensable for gliomatumorigenesis.

Project description:Malignant glioma is the most common primary tumor of the central nervous system. Chemotherapy and radiotherapy are the most common therapeutic approaches in glioma therapy. Both processes mainly kill cancer cells through generating high Reactive Oxygen Species (ROS) and lead to oxidative DNA damage. However, tumor resistance to ROS is always a challenge for cancer treatment. Human Mut T homolog 1 (MTH1, also known as NUDT1) is regarded as a protector of nucleotides against oxidization. Recent reports have verified that overexpression of MTH1 could remove oxidized dNTP pools. Here, we find that MTH1 is overexpressed both at mRNA and protein levels in GBM. MTH1 silencing inhibits colony formation; tumor spheres formation and xenograft tumor growth, and more importantly, the viability of glioma cells is significantly decreased in H2O2 after MTH1 was knocked down in glioma. PI staining show that H2O2 cause more glioma cell death after MTH1 silencing. So we speculate that overexpression of MTH1 is crucial for glioma survival, suppression of its expression can inhibit cancer cell survival in vitro and in vivo, MTH1 may be a potential target for human glioma therapy in future.

| S-EPMC4931174 | biostudies-literature

MTH1 as a Chemotherapeutic Target: The Elephant in the Room.

Project description:Many tumors sustain elevated levels of reactive oxygen species (ROS), which drive oncogenic signaling. However, ROS can also trigger anti-tumor responses, such as cell death or senescence, through induction of oxidative stress and concomitant DNA damage. To circumvent the adverse consequences of elevated ROS levels, many tumors develop adaptive responses, such as enhanced redox-protective or oxidatively-generated damage repair pathways. Targeting these enhanced oxidative stress-protective mechanisms is likely to be both therapeutically effective and highly specific to cancer, as normal cells are less reliant on such mechanisms. In this review, we discuss one such stress-protective protein human MutT Homolog1 (MTH1), an enzyme that eliminates 8-oxo-7,8-dihydro-2'-deoxyguanosine triphosphate (8-oxodGTP) through its pyrophosphatase activity, and is found to be elevated in many cancers. Our studies, and subsequently those of others, identified MTH1 inhibition as an effective tumor-suppressive strategy. However, recent studies with the first wave of MTH1 inhibitors have produced conflicting results regarding their cytotoxicity in cancer cells and have led to questions regarding the validity of MTH1 as a chemotherapeutic target. To address the proverbial "elephant in the room" as to whether MTH1 is a bona fide chemotherapeutic target, we provide an overview of MTH1 function in the context of tumor biology, summarize the current literature on MTH1 inhibitors, and discuss the molecular contexts likely required for its efficacy as a therapeutic target.

| S-EPMC5447957 | biostudies-literature

The Existence of MTH1-independent 8-oxodGTPase Activity in Cancer Cells as a Compensatory Mechanism against On-target Effects of MTH1 Inhibitors.

Project description:Investigations into the human 8-oxodGTPase, MutT Homolog 1 (MTH1), have risen sharply since the first-in-class MTH1 inhibitors were reported to be highly tumoricidal. However, MTH1 as a cancer therapeutic target is currently controversial because subsequently developed inhibitors did not exhibit similar cytotoxic effects. Here, we provide the first direct evidence for MTH1-independent 8-oxodGTPase function in human cancer cells and human tumors, using a novel ATP-releasing guanine-oxidized (ARGO) chemical probe. Our studies show that this functionally redundant 8-oxodGTPase activity is not decreased by five different published MTH1-targeting small molecules or by MTH1 depletion. Significantly, while only the two first-in-class inhibitors, TH588 and TH287, reduced cancer cell viability, all five inhibitors evaluated in our studies decreased 8-oxodGTPase activity to a similar extent. Thus, the reported efficacy of the first-in-class MTH1 inhibitors does not arise from their inhibition of MTH1-specific 8-oxodGTPase activity. Comparison of DNA strand breaks, genomic 8-oxoguanine incorporation, or alterations in cellular oxidative state by TH287 versus the noncytotoxic inhibitor, IACS-4759, contradict that the cytotoxicity of the former results solely from increased levels of oxidatively damaged genomic DNA. Thus, our findings indicate that mechanisms unrelated to oxidative stress or DNA damage likely underlie the reported efficacy of the first-in-class inhibitors. Our study suggests that MTH1 functional redundancy, existing to different extents in all cancer lines and human tumors evaluated in our study, is a thus far undefined factor which is likely to be critical in understanding the importance of MTH1 and its clinical targeting in cancer.

| S-EPMC7079300 | biostudies-literature

MTH1 protects platelet mitochondria from oxidative damage and regulates platelet function and thrombosis.

Project description:Human MutT Homolog 1 (MTH1) is a nucleotide pool sanitization enzyme that hydrolyzes oxidized nucleotides to prevent their mis-incorporation into DNA under oxidative stress. Expression and functional roles of MTH1 in platelets are not known. Here, we show MTH1 expression in platelets and its deficiency impairs hemostasis and arterial/venous thrombosis in vivo. MTH1 deficiency reduced platelet aggregation, phosphatidylserine exposure and calcium mobilization induced by thrombin but not by collagen-related peptide (CRP) along with decreased mitochondrial ATP production. Thrombin but not CRP induced Ca2+-dependent mitochondria reactive oxygen species generation. Mechanistically, MTH1 deficiency caused mitochondrial DNA oxidative damage and reduced the expression of cytochrome c oxidase 1. Furthermore, MTH1 exerts a similar role in human platelet function. Our study suggests that MTH1 exerts a protective function against oxidative stress in platelets and indicates that MTH1 could be a potential therapeutic target for the prevention of thrombotic diseases.

| S-EPMC10415391 | biostudies-literature

Development of MTH1-Binding Nucleotide Analogs Based on 7,8-Dihalogenated 7-Deaza-dG Derivatives.

Project description:MTH1 is an enzyme that hydrolyzes 8-oxo-dGTP, which is an oxidatively damaged nucleobase, into 8-oxo-dGMP in nucleotide pools to prevent its mis-incorporation into genomic DNA. Selective and potent MTH1-binding molecules have potential as biological tools and drug candidates. We recently developed 8-halogenated 7-deaza-dGTP as an 8-oxo-dGTP mimic and found that it was not hydrolyzed, but inhibited enzyme activity. To further increase MTH1 binding, we herein designed and synthesized 7,8-dihalogenated 7-deaza-dG derivatives. We successfully synthesized multiple derivatives, including substituted nucleosides and nucleotides, using 7-deaza-dG as a starting material. Evaluations of the inhibition of MTH1 activity revealed the strong inhibitory effects on enzyme activity of the 7,8-dihalogenated 7-deaza-dG derivatives, particularly 7,8-dibromo 7-daza-dGTP. Based on the results obtained on kinetic parameters and from computational docking simulating studies, these nucleotide analogs interacted with the active site of MTH1 and competitively inhibited the substrate 8-oxodGTP. Therefore, novel properties of repair enzymes in cells may be elucidated using new compounds.

| S-EPMC7866122 | biostudies-literature

Echinacoside induces apoptotic cancer cell death by inhibiting the nucleotide pool sanitizing enzyme MTH1.

Project description:Inhibition of the nucleotide pool sanitizing enzyme MTH1 causes extensive oxidative DNA damages and apoptosis in cancer cells and hence may be used as an anticancer strategy. As natural products have been a rich source of medicinal chemicals, in the present study, we used the MTH1-catalyzed enzymatic reaction as a high-throughput in vitro screening assay to search for natural compounds capable of inhibiting MTH1. Echinacoside, a compound derived from the medicinal plants Cistanche and Echinacea, effectively inhibited the catalytic activity of MTH1 in an in vitro assay. Treatment of various human cancer cell lines with Echinacoside resulted in a significant increase in the cellular level of oxidized guanine (8-oxoguanine), while cellular reactive oxygen species level remained unchanged, indicating that Echinacoside also inhibited the activity of cellular MTH1. Consequently, Echinacoside treatment induced an immediate and dramatic increase in DNA damage markers and upregulation of the G1/S-CDK inhibitor p21, which were followed by marked apoptotic cell death and cell cycle arrest in cancer but not in noncancer cells. Taken together, these studies identified a natural compound as an MTH1 inhibitor and suggest that natural products can be an important source of anticancer agents.

| S-EPMC4677763 | biostudies-literature

MTH1 Inhibitor TH1579 Induces Oxidative DNA Damage and Mitotic Arrest in Acute Myeloid Leukemia.

Project description:Acute myeloid leukemia (AML) is an aggressive hematologic malignancy, exhibiting high levels of reactive oxygen species (ROS). ROS levels have been suggested to drive leukemogenesis and is thus a potential novel target for treating AML. MTH1 prevents incorporation of oxidized nucleotides into the DNA to maintain genome integrity and is upregulated in many cancers. Here we demonstrate that hematologic cancers are highly sensitive to MTH1 inhibitor TH1579 (karonudib). A functional precision medicine ex vivo screen in primary AML bone marrow samples demonstrated a broad response profile of TH1579, independent of the genomic alteration of AML, resembling the response profile of the standard-of-care treatments cytarabine and doxorubicin. Furthermore, TH1579 killed primary human AML blast cells (CD45+) as well as chemotherapy resistance leukemic stem cells (CD45+Lin-CD34+CD38-), which are often responsible for AML progression. TH1579 killed AML cells by causing mitotic arrest, elevating intracellular ROS levels, and enhancing oxidative DNA damage. TH1579 showed a significant therapeutic window, was well tolerated in animals, and could be combined with standard-of-care treatments to further improve efficacy. TH1579 significantly improved survival in two different AML disease models in vivo. In conclusion, the preclinical data presented here support that TH1579 is a promising novel anticancer agent for AML, providing a rationale to investigate the clinical usefulness of TH1579 in AML in an ongoing clinical phase I trial. SIGNIFICANCE: The MTH1 inhibitor TH1579 is a potential novel AML treatment, targeting both blasts and the pivotal leukemic stem cells while sparing normal bone marrow cells.

| S-EPMC9397639 | biostudies-literature

Multi-forms of human MTH1 polypeptides produced by alternative translation initiation and single nucleotide polymorphism.

Project description:The human MTH1 gene for 8-oxo-7,8-dihydrodeoxyguanosine triphosphatase, produces seven types (types 1, 2A, 2B, 3A, 3B, 4A and 4B) of mRNAs. The B-type mRNAs with exon 2b-2c segments have three additional in-frame AUGs in their 5' regions. We report here that these transcripts produce three forms of MTH1 polypeptides (p22, p21 and p18) in in vitro translation reactions. Three polypeptides were also detected in extracts of human cells, using western blotting. B-type mRNAs with a polymorphic alteration (GU-->GC) at the beginning of exon 2c that converts an in-frame UGA to CGA yielding another in-frame AUG further upstream, produced an additional polypeptide (p26) in vitro. Substitution of each AUG abolished the production of each corresponding polypeptide. Cell lines from individuals with the GC allele contain more B-type mRNAs than do those of GT homozygotes, and the former produce all of four polypeptides but the latter lack p26. Amounts of each polypeptide reflected copy number of the GC allele in each cell line. There is an apparent linkage dis-equilibrium between the two polymorphic sites, GT/GC at exon 2c and Val83/Met83 at codon 83 for p18.

| S-EPMC148714 | biostudies-other

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data