Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.1.21 (thymidine kinase)
 7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

It is well-known that viral thymidine kinase (TK) expression is important for the maximum demonstration of virulence of herpes simplex virus (HSV). In this study, we have investigated interactions of a TK- mutant of virulent HSV type 2 (HSV-2) (syn+) and a nonneuroinvasive HSV-1 (syn) in mice. When the mice were inoculated with each virus alone in their rear footpads, no mice were killed even after infection with high doses of viruses (greater than 10(6) PFU per mouse), whereas 100% of the mice died when inoculated with 10(5) PFU of a 1:1 mixture of HSV-2 TK- mutant and nonneuroinvasive HSV-1. The 1:1 mixture exhibited even more virulence than the parental HSV-2; the mean survival time of coinfected mice was significantly shorter than that of mice inoculated with 10(5) PFU of the virulent HSV-2. We have also examined the genotypes and phenotypes of viruses isolated from the central nervous system of coinfected mice. Of 50 isolates, 7 were judged to be recombinants from their restriction endonuclease cleavage patterns, but all were nonneuroinvasive. In addition, all syn+ viruses (23 clones) tested were found to have TK- phenotypes, indicating that the majority of viruses present in the central nervous system were TK- viruses, since about 90% of viruses detected in spinal cords and brains exhibited syn+ phenotypes. These results strongly suggest that the lethal invasion of the central nervous system by HSV-2 TK- and nonneuroinvasive HSV-1 was the consequence of in vivo complementation between the two viruses.
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PMID:Complementary lethal invasion of the central nervous system by nonneuroinvasive herpes simplex virus types 1 and 2. 164 47

A 2-kilobase fragment from the 5'-flanking region of the human myoglobin gene extending from -2038 to +7 relative to the cap site regulates expression of a heterologous reporter gene in a cell-specific and developmentally regulated manner. Functional analyses of 5' and internal deletions indicate that sequences located between -261 and -205 are essential for muscle-specific expression in cooperation with the myoglobin core promoter. A 167-base pair fragment containing these sequences (-371 to -205) enhances expression from myoglobin core promoter elements in a manner that is independent of its orientation and position relative to the cap site. When linked to the herpes simplex virus thymidine kinase promoter, this 167-base pair fragment of the myoglobin gene also enhances expression in differentiated myotubes but not in undifferentiated myoblasts or fibroblasts. Nucleotide sequences within the region that is essential for enhancing activity (-261 to -205) are conserved in other mammalian myoglobin genes (seal, mouse) and resemble sequences within control regions of other genes that are expressed selectively in striated myocytes. These data indicate that the 5'-flanking region of the human myoglobin gene contains an enhancer-like element that is important for transcriptional activation during myocyte differentiation.
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PMID:Identification of a muscle-specific enhancer within the 5'-flanking region of the human myoglobin gene. 276 49

In this study, a pre-existing "library" of Herpes Simplex Virus (HSV) intertypic recombinants was found to be not useful for mapping HSV genes controlling viral neurovirulence in mice. Most of these agents were significantly less virulent than either parental type following intracranial inoculation, and in the general case this lessened virulence could be attributed to inefficient replication in any cell type at 38.5 degrees (the normal temperature of the mouse). One agent tested (recombinant RE6) was completely non-neurovirulent following intracranial inoculation of as much as 3.2 X 10(7) PFU. Since about 10 PFU of either 17 Syn+ or HG52 (the "parental" strains of this recombinant) were lethal for mice, RE6 is at least 10 million-fold less neurovirulent than the wild-type strains from which it was produced. The function of the defective gene(s) in RE6 is not yet known, but it is not required for the expression of viral thymidine kinase, efficient replication in cultured cells at 38.5 degrees, or replication in non-neural mouse tissue in vivo. Therefore, the defect in RE6 is in an HSV gene function(s) which is absolutely required for neurovirulence but not for general viral replication. Several possibilities for the molecular nature of the defect in RE6 are discussed.
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PMID:Biological characterization of a herpes simplex virus intertypic recombinant which is completely and specifically non-neurovirulent. 631 49

The murine T cell hybridoma line which specifically binds antigen (ovalbumin) was established using a cell fusion technique with Sendai virus. Regional lymph node cells from ovalbumin (OVA) immunized C57BL/6 mice were fused with thymidine kinase deficient variant cells of the EL-4 cell line (originating from a thymoma of a C57BL/6 mouse). Approximately one hundred cell lines were established and the antigen binding activity was determined by rosette formation with OVA coated sheep red blood cells (SRBC). One hybridoma cell line, MMH-77, could form rosettes and this formation was specifically inhibited by the addition of free OVA. The ability of the cell line to form rosettes varied from one stage of the cell cycle to the other with the maximum ability in the S phase.
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PMID:Fluctuation of antigen binding activity during the cell cycle in the synchronized population of the murine T hybridoma cell line. 698 84

Non-invasive imaging of transgene expression requires the appropriate combination of a reporter gene and a reporter probe. [18F]FEAU positron emission tomography (PET) is used for the assessment of herpes simplex virus type-1 thymidine kinase gene expression. Hybrid AAV phage (termed AAVP) can be adapted to transduce mammalian cells by targeting to a specific receptor. We evaluated a targeted AAVP vector using [18F]FEAU PET. This protocol describes [18F]FEAU production and dosing, micro-PET imaging and image analysis. 2-Deoxy-2-trifluoromethanesulfonyl-1,3,5-tri-O-benzoyl-alpha-D-ribofuranose is radio-fluorinated, converted into its 1-bromo derivative and coupled with protected 5-ethyl uracil. The coupled product is hydrolyzed and purified using HPLC. Tumor-bearing animals targeted with either retroviral or AAVP vectors are anesthetized and injected with [18F]FEAU (0.1 mCi per mouse); this is followed 2 h after injection by imaging on a micro-PET. Production of [18F]FEAU requires approximately 3.5 h from the end of bombardment. PET imaging studies require 2-3 h (depending on the number of animals) after synthesis of [18F]FEAU.
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PMID:Molecular PET imaging of HSV1-tk reporter gene expression using [18F]FEAU. 1740 3

Previously, we reported that a recombinant vaccinia virus (VACV) carrying a light-emitting fusion gene enters, replicates in, and reveals the locations of tumors in mice. A new recombinant VACV, GLV-1h68, as a simultaneous diagnostic and therapeutic agent, was constructed by inserting three expression cassettes (encoding Renilla luciferase-Aequorea green fluorescent protein fusion, beta-galactosidase, and beta-glucuronidase) into the F14.5L, J2R (encoding thymidine kinase) and A56R (encoding hemagglutinin) loci of the viral genome, respectively. I.v. injections of GLV-1h68 (1x10(7) plaque-forming unit per mouse) into nude mice with established (approximately 300-500 mm3) s.c. GI-101A human breast tumors were used to evaluate its toxicity, tumor targeting specificity, and oncolytic efficacy. GLV-1h68 showed an enhanced tumor targeting specificity and much reduced toxicity compared with its parental LIVP strains. The tumors colonized by GLV-1h68 exhibited growth, inhibition, and regression phases followed by tumor eradication within 130 days in 95% of the mice tested. Tumor regression in live animals was monitored in real time based on decreasing light emission, hence demonstrating the concept of a combined oncolytic virus-mediated tumor diagnosis and therapy system. Transcriptional profiling of regressing tumors based on a mouse-specific platform revealed gene expression signatures consistent with immune defense activation, inclusive of IFN-stimulated genes (STAT-1 and IRF-7), cytokines, chemokines, and innate immune effector function. These findings suggest that immune activation may combine with viral oncolysis to induce tumor eradication in this model, providing a novel perspective for the design of oncolytic viral therapies for human cancers.
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PMID:Eradication of solid human breast tumors in nude mice with an intravenously injected light-emitting oncolytic vaccinia virus. 1794 38

Duchenne muscular dystrophy (DMD) is a severe genetic neuromuscular disorder that affects 1 in 3,500 boys, and is characterized by progressive muscle degeneration. In patients, the ability of resident muscle satellite cells (SCs) to regenerate damaged myofibers becomes increasingly inefficient. Therefore, transplantation of muscle progenitor cells (MPCs)/myoblasts from healthy subjects is a promising therapeutic approach to DMD. A major limitation to the use of stem cell therapy, however, is a lack of reliable imaging technologies for long-term monitoring of implanted cells, and for evaluating its effectiveness. Here, we describe a non-invasive, real-time approach to evaluate the success of myoblast transplantation. This method takes advantage of a unified fusion reporter gene composed of genes (firefly luciferase [fluc], monomeric red fluorescent protein [mrfp] and sr39 thymidine kinase [sr39tk]) whose expression can be imaged with different imaging modalities. A variety of imaging modalities, including positron emission tomography (PET), single-photon emission computed tomography (SPECT), magnetic resonance imaging (MRI), optical imaging, and high frequency 3D-ultrasound are now available, each with unique advantages and limitations. Bioluminescence imaging (BLI) studies, for example, have the advantage of being relatively low cost and high-throughput. It is for this reason that, in this study, we make use of the firefly luciferase (fluc) reporter gene sequence contained within the fusion gene and bioluminescence imaging (BLI) for the short-term localization of viable C2C12 myoblasts following implantation into a mouse model of DMD (muscular dystrophy on the X chromosome [mdx] mouse). Importantly, BLI provides us with a means to examine the kinetics of labeled MPCs post-implantation, and will be useful to track cells repeatedly over time and following migration. Our reporter gene approach further allows us to merge multiple imaging modalities in a single living subject; given the tomographic nature, fine spatial resolution and ability to scale up to larger animals and humans, PET will form the basis of future work that we suggest may facilitate rapid translation of methods developed in cells to preclinical models and to clinical applications.
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PMID:Molecular imaging to target transplanted muscle progenitor cells. 2356 4