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Neoplasia (New York, N.Y.) 2000A variety of imaging technologies are being investigated as tools for studying gene expression in living subjects. Noninvasive, repetitive and quantitative imaging of... (Review)
Review
A variety of imaging technologies are being investigated as tools for studying gene expression in living subjects. Noninvasive, repetitive and quantitative imaging of gene expression will help both to facilitate human gene therapy trials and to allow for the study of animal models of molecular and cellular therapy. Radionuclide approaches using single photon emission computed tomography (SPECT) and positron emission tomography (PET) are the most mature of the current imaging technologies and offer many advantages for imaging gene expression compared to optical and magnetic resonance imaging (MRI)-based approaches. These advantages include relatively high sensitivity, full quantitative capability (for PET), and the ability to extend small animal assays directly into clinical human applications. We describe a PET scanner (microPET) designed specifically for studies of small animals. We review "marker/reporter gene" imaging approaches using the herpes simplex type 1 virus thymidine kinase (HSV1-tk) and the dopamine type 2 receptor (D2R) genes. We describe and contrast several radiolabeled probes that can be used with the HSV1-tk reporter gene both for SPECT and for PET imaging. We also describe the advantages/disadvantages of each of the assays developed and discuss future animal and human applications.
Topics: Animals; Gene Expression; Herpesvirus 1, Human; Humans; Mice; Models, Biological; Radionuclide Imaging; Receptors, Dopamine D2; Thymidine Kinase; Tomography, Emission-Computed; Transgenes
PubMed: 10933072
DOI: 10.1038/sj.neo.7900083 -
European Journal of Medicinal Chemistry Jul 2015A library of sixteen 2nd generation amino- and amido-substituted carboranyl pyrimidine nucleoside analogs, designed as substrates and inhibitors of thymidine kinase 1...
A library of sixteen 2nd generation amino- and amido-substituted carboranyl pyrimidine nucleoside analogs, designed as substrates and inhibitors of thymidine kinase 1 (TK1) for potential use in boron neutron capture therapy (BNCT) of cancer, was synthesized and evaluated in enzyme kinetic-, enzyme inhibition-, metabolomic-, and biodistribution studies. One of these 2nd generation carboranyl pyrimidine nucleoside analogs (YB18A [3]), having an amino group directly attached to a meta-carborane cage tethered via ethylene spacer to the 3-position of thymidine, was approximately 3-4 times superior as a substrate and inhibitor of hTK1 than N5-2OH (2), a 1st generation carboranyl pyrimidine nucleoside analog. Both 2 and 3 appeared to be 5'-monophosphorylated in TK1(+) RG2 cells, both in vitro and in vivo. Biodistribution studies in rats bearing intracerebral RG2 glioma resulted in selective tumor uptake of 3 with an intratumoral concentration that was approximately 4 times higher than that of 2. The obtained results significantly advance the understanding of the binding interactions between TK1 and carboranyl pyrimidine nucleoside analogs and will profoundly impact future design strategies for these agents.
Topics: Animals; Boron Compounds; Boron Neutron Capture Therapy; Cell Line, Tumor; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Glioma; Molecular Structure; Protein Kinase Inhibitors; Pyrimidine Nucleosides; Rats; Structure-Activity Relationship; Thymidine Kinase
PubMed: 26087030
DOI: 10.1016/j.ejmech.2015.05.042 -
Cancer Science Aug 2023Immune cells can recognize tumor-associated antigens released from dead tumor cells, which elicit immune responses, potentially resulting in tumor regression. Tumor cell...
Immune cells can recognize tumor-associated antigens released from dead tumor cells, which elicit immune responses, potentially resulting in tumor regression. Tumor cell death induced by chemotherapy has also been reported to activate immunity. However, various studies have reported drug-induced immunosuppression or suppression of inflammation by apoptotic cells. Thus, this study aimed to investigate whether apoptotic tumor cells trigger antitumor immunity independent of anticancer treatment. Local immune responses were evaluated after direct induction of tumor cell apoptosis using a Herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV) system. The inflammatory response was significantly altered at the tumor site after apoptosis induction. The expression of cytokines and molecules that activate and suppress inflammation simultaneously increased. The HSV-tk/GCV-induced tumor cell apoptosis resulted in tumor growth suppression and promoted T lymphocyte infiltration into tumors. Therefore, the role of T cells after inducing tumor cell death was explored. CD8 T cell depletion abrogated the antitumor efficacy of apoptosis induction, indicating that tumor regression was mainly dependent on CD8 T cells. Furthermore, CD4 T cell depletion inhibited tumor growth, suggesting the potential role of CD4 T cells in suppressive tumor immunity. Tumor tissues were evaluated after tumor cell apoptosis and CD4 T cell depletion to elucidate this immunological mechanism. Foxp3 and CTLA4, regulatory T-cell markers, decreased. Furthermore, arginase 1, an immune-suppressive mediator induced by myeloid cells, was significantly downregulated. These findings indicate that tumors accelerate CD8 T cell-dependent antitumor immunity and CD4 T cell-mediated suppressive immunity. These findings could be a therapeutic target for immunotherapy in combination with cytotoxic chemotherapy.
Topics: Humans; Ganciclovir; Thymidine Kinase; Simplexvirus; Genetic Therapy; Apoptosis; Neoplasms; CD8-Positive T-Lymphocytes; CD4-Positive T-Lymphocytes; Inflammation; Antiviral Agents
PubMed: 37322820
DOI: 10.1111/cas.15843 -
Journal of Clinical Laboratory Analysis Jul 2019Thymidine kinase 1 (TK1) is a key enzyme in the pyrimidine salvage pathway. Increased TK1 concentration correlates with cell division. TK1 is an emerging biomarker in...
OBJECTIVE
Thymidine kinase 1 (TK1) is a key enzyme in the pyrimidine salvage pathway. Increased TK1 concentration correlates with cell division. TK1 is an emerging biomarker in cancer diagnosis; however, its effectiveness in diagnosis and management for malignant pleural effusion (MPE) is unclear. We evaluated the diagnostic efficiency and prognostic value of pleural effusion TK1 (pTK1) concentration for MPE.
METHODS
From 2013 to 2017, 210 pleural effusion samples were collected from 160 patients diagnosed with MPE and 50 patients diagnosed with benign pleural effusion (BPE). TK1 concentrations in pleural effusion were measured by chemiluminescence dot blot assays. The median follow-up was 12 months. We constructed a receiver-operating characteristic (ROC) curve to find the optimal cutoff value for MPE diagnosis. The hazard ratios were estimated using a multivariable Cox proportional hazard model. A nomogram was drawn to illustrate the prognostic characteristics of MPE.
RESULTS
The TK1 concentration in pleural effusion was significantly higher in MPE than BPE (P < 0.001), and patients with MPE could be distinguished by an optimal cutoff value of 3.10 pmol/L with a sensitivity of 0.894 and a specificity of 0.800. The multivariate analysis suggested that pTK1 concentration was an independent predictor of survival in patients with MPE.
CONCLUSIONS
The diagnostic and prognostic prediction of MPE may be improved by measuring pTK1 concentration and utilizing a multivariate nomogram.
Topics: Aged; Biomarkers, Tumor; Female; Humans; Kaplan-Meier Estimate; Male; Middle Aged; Nomograms; Pleural Effusion; Pleural Effusion, Malignant; Reproducibility of Results; Thymidine Kinase
PubMed: 30985967
DOI: 10.1002/jcla.22901 -
Proceedings of the National Academy of... Sep 1980A mixture of two recombinant plasmids was microinjected into mouse thymidine kinase-negative fibroblasts (L cells). One plasmid contained the thymidine kinase gene of...
A mixture of two recombinant plasmids was microinjected into mouse thymidine kinase-negative fibroblasts (L cells). One plasmid contained the thymidine kinase gene of herpes simplex virus type I and the other contained the human beta globin gene. Seven fibroblast colonies arising from injected cells incubated in hypoxanthine/aminopterin/thymidine medium were analyzed. These microinjected cells were shown to: (i) produce functionally active herpes simplex type I thymidine kinase enzyme, (ii) replicate the human beta globin gene, and (iii) produce human beta globin mRNA sequences at low levels. Thus, the genetic defect (lack of thymidine kinase activity) was corrected by the microinjected thymidine kinase gene, and a coinjected human beta globin gene was replicated and weakly expressed.
Topics: Animals; Clone Cells; Cloning, Molecular; DNA, Recombinant; Escherichia coli; Gene Expression Regulation; Genetic Vectors; Globins; Humans; L Cells; Mice; Microinjections; Nucleic Acid Hybridization; Plasmids; RNA, Messenger; Simplexvirus; Thymidine Kinase; Transformation, Genetic
PubMed: 6254080
DOI: 10.1073/pnas.77.9.5399 -
Proceedings of the National Academy of... Oct 1972The cell line LM(TK(-)) Cl 1D, a derivative of mouse L fibroblasts deficient in thymidine kinase (EC 2.7.1.21) that shows very little thymidine kinase activity in...
The cell line LM(TK(-)) Cl 1D, a derivative of mouse L fibroblasts deficient in thymidine kinase (EC 2.7.1.21) that shows very little thymidine kinase activity in extracts of whole cells as compared to the parental line, and that does not incorporate thymidine or 5-bromodeoxyuridine into nuclear DNA, has maintained the capacity to incorporate these precursors into mitochondrial DNA at a substantial rate. The amount of [methyl-(3)H]thymidine incorporated into mitochondrial DNA of LM(TK(-)) Cl 1D cells after long-term labeling has been conservatively estimated in different experiments to be between 30 and 60% of that incorporated into mitochondrial DNA or nuclear DNA of strain A9, an L-cell derivative without any thymidine-kinase dificiency; by contrast, the incorporation of thymidine into nuclear DNA of Cl 1D cells is less than 1% of that in A9 cells. These results strongly suggest that the loss of thymidine kinase activity in the extramitochondrial compartment of LM(TK(-)) Cl 1D cells has not been accompanied by the loss of the mitochondria-associated enzyme activity, pointing to a different genetic or epigenetic control of the extramitochondrial and mitochondrial enzymes.
Topics: Adenosine; Animals; Bromodeoxyuridine; Carbon Isotopes; Cell Line; Cell Nucleus; Centrifugation, Density Gradient; Connective Tissue; DNA; Ethidium; Genetic Code; L Cells; Mice; Mitochondria; Thymidine; Thymidine Kinase; Tritium
PubMed: 4507611
DOI: 10.1073/pnas.69.10.2874 -
Molecular and Cellular Biology Nov 1984A cDNA containing the entire coding region of the human thymidine kinase gene has been molecularly cloned. The cDNA is under the control of a simian virus 40 promoter... (Comparative Study)
Comparative Study
A cDNA containing the entire coding region of the human thymidine kinase gene has been molecularly cloned. The cDNA is under the control of a simian virus 40 promoter and is expressible in mammalian cells. The complete nucleotide sequence of the human thymidine kinase cDNA has been determined. The cDNA is 1,421 base pairs in length and has a large open reading frame of 702 base pairs capable of specifying a protein with a molecular weight of 25,504. Genomic Southern blotting experiments show that sequences homologous to the human thymidine kinase cDNA are conserved among many vertebrates, including prosimians (lemur), tree shrews, rats, mice, and chickens. Direct comparison of the nucleotide sequences of the human thymidine kinase cDNA and the chicken thymidine kinase gene reveals ca. 70% overall homology. This homology is extended further at the amino acid sequence level, with greater than 74% amino acid residues matched between the human and chicken thymidine kinase proteins.
Topics: Amino Acid Sequence; Animals; Base Sequence; Cloning, Molecular; DNA; Genes; Humans; Species Specificity; Thymidine Kinase; Vertebrates
PubMed: 6549046
DOI: 10.1128/mcb.4.11.2316-2320.1984 -
Asian Journal of Surgery Feb 2024
Topics: Male; Humans; Thymidine Kinase; Prostatic Neoplasms
PubMed: 38135534
DOI: 10.1016/j.asjsur.2023.12.011 -
British Journal of Cancer Oct 1990The enzyme thymidine kinase is associated with DNA synthesis. Thymidine kinase serum levels were studied in normal controls (n = 20), patients with primary breast cancer...
The enzyme thymidine kinase is associated with DNA synthesis. Thymidine kinase serum levels were studied in normal controls (n = 20), patients with primary breast cancer (n = 60), patients with systemic breast cancer (n = 20) and as a non-cancer disease control group in patients with inflammatory gastrointestinal disorders (n = 20). Comparison of pretreatment values in the cancer patients with the normal controls showed a significant difference between the three groups in relation to stage of disease: mean values 4.22 (+/- 1.08), 6.22 (+/- 2.24) and 9.79 (+/- 7.56) pmol ml-1 h-1 for normal controls, operable breast cancer and systemic breast cancer respectively (P less than 0.005; analysis of variance). Patients with systemic breast cancer had a significantly elevated serum thymidine kinase level compared to controls (P less than 0.01) and patients with primary operable cancer (P less than 0.05). Patients with primary operable cancer had significantly higher serum thymidine kinase levels over normal controls (P less than 0.01). Mean serum TK in patients with inflammatory gastrointestinal diseases was similar to normal controls but significantly less than both patients with primary operable breast cancer and patients with systemic breast cancer. Twenty patients with operable breast cancer were followed up after primary surgery by serial 3-monthly thymidine kinase levels in the disease free interval. Four patients have developed systemic recurrence with a rise in the mean thymidine kinase value to 14.3 pmol ml-1 h-1. Ten patients with advanced breast cancer had serial thymidine kinase levels measured 2-monthly during the first 6 months of primary hormone therapy. The serum values fell in all five responders (mean 9.12-4.78 pmol ml-1 h-1) and rose in all five progressors (mean 8.62-38.5 pmol ml-1 h-1). Serum thymidine kinase reflects stage of disease in breast cancer. Serial thymidine kinase levels in patients with systemic breast cancer reflected response to systemic therapy.
Topics: Breast Neoplasms; Female; Humans; Middle Aged; Neoplasm Recurrence, Local; Thymidine Kinase
PubMed: 2223587
DOI: 10.1038/bjc.1990.352 -
EBioMedicine Aug 2019TK2 is a nuclear gene encoding the mitochondrial matrix protein thymidine kinase 2 (TK2), a critical enzyme in the mitochondrial nucleotide salvage pathway. Deficiency...
BACKGROUND
TK2 is a nuclear gene encoding the mitochondrial matrix protein thymidine kinase 2 (TK2), a critical enzyme in the mitochondrial nucleotide salvage pathway. Deficiency of TK2 activity causes mitochondrial DNA (mtDNA) depletion, which in humans manifests predominantly as a mitochondrial myopathy with onset typically in infancy and childhood. We previously showed that oral treatment of the Tk2 H126N knock-in mouse model (Tk2) with the TK2 substrates, deoxycytidine (dCtd) and thymidine (dThd), delayed disease onset and prolonged median survival by 3-fold. Nevertheless, dCtd + dThd treated Tk2 mice showed mtDNA depletion in brain as early as postnatal day 13 and in virtually all other tissues at age 29 days.
METHODS
To enhance mechanistic understanding and efficacy of dCtd + dThd therapy, we studied the bioavailability of dCtd and dThd in various tissues as well as levels of the cytosolic enzymes, TK1 and dCK that convert the deoxynucleosides into dCMP and dTMP.
FINDINGS
Parenteral treatment relative to oral treatment produced higher levels of dCtd and dThd and improved mtDNA levels in liver and heart, but did not ameliorate molecular defects in brain or prolong survival. Down-regulation of TK1 correlated with temporal- and tissue-specificity of response to dCtd + dThd. Finally, we observed in human infant and adult muscle expression of TK1 and dCK, which account for the long-term efficacy to dCtd + dThd therapy in TK2 deficient patients.
INTERPRETATIONS
These data indicate that the cytosolic pyrimidine salvage pathway enzymes TK1 and dCK are critical for therapeutic efficacy of deoxynucleoside therapy for Tk2 deficiency. FUND: National Institutes of Health P01HD32062.
Topics: Animals; Biological Availability; Blood-Brain Barrier; DNA, Mitochondrial; Deoxyribonucleosides; Disease Models, Animal; Enzyme Activation; Humans; Mice; Mice, Knockout; Mice, Transgenic; Mitochondria; Muscle, Skeletal; Organ Specificity; Oxidative Phosphorylation; Phenotype; Thymidine Kinase
PubMed: 31383553
DOI: 10.1016/j.ebiom.2019.07.037