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)

We introduced chromosome-mediated genes into mouse thymidine kinase-deficient FM3A (FM3Atk-) cells, by electroporation. The effects of some parameters on the electric shock-mediated transfection of FM3Atk- cells were investigated. Gene transfer of mouse L929 metaphase chromosome DNA into FM3Atk- resulted in a maximum frequency of (3.0 +/- 0.3) x 10(-5) at a cell density of 2.0 x 10(8)/ml and chromosome dosage of 5.0 x 10(7) cell equivalents/ml in a buffer containing 0.25 M mannitol, 0.5 mM MgCl2, 0.1mM CaCl2, and 1 mM Tris-HCl (pH 7.1). The highest yield of the transformants was obtained at an electric field strength of 1 kV/cm and a capacitance of 35 microF, with a single exponentially decaying pulse at 0 degrees C was optimal for post-shock incubation after electroporation. The tk gene was detected in the transformants by in situ hybridization analysis.
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PMID:A new and efficient method for gene transfer into mouse FM3A cells using metaphase chromosomes by electroporation. 898 67

6-Chloropurine derivatives of gamma-(Z)-ethylidene-2,3-dimethoxybutenolide 3a, gamma-(Z)-ethylidene-2-methoxy-3-(4-nitro)benzyloxybutenolide 3b, gamma-(Z)-ethylidene-2-(4-nitro)benzyloxy-3-methoxybutenolide 3c, gamma-(Z)-ethylidene-2,3-di(4-nitro)benzyloxybutenolide 3d, and dimethylphosphono-gamma-(Z)-ethylidene-2,3-dimethoxybutenolide 11 as well as the adenine derivative of gamma-(Z)-ethylidene-2,3-dimethoxybutenolide 6 were synthesized. The key steps in the high-yield synthesis of 6 involved hydration/dehydration of the C(4)=C(5) in the precursor 3a. In the presence of NH4OH at elevated temperature, 3a underwent a reverse Michael-type addition with water to produce hydrate 5. At 37 degrees C, 6 was also hydrated in the presence of S-adenosyl-L-homocysteine hydrolase to afford 5. Butenolide 6 exhibited an inhibitory property toward the enzyme. Such type II (enzyme-mediated addition of water across C(4)=C(5)) mechanism is the first example of "enzyme-substrate intermediate" inactivation of S-adenosyl-L-homocysteine hydrolase. In contrast with type I mechanism-based inactivation, reduction of enzyme-bound NADP(+) to NADPH was not observed. Upon treatment with HCl, stereoselective dehydration of 5 occurred to give the target molecule 6. At ambident temperature, 3a was hydrated in the presence of NH4OH or pig liver esterase to produce 6-chloropurine derivative 4. An unambiguous proof of the structures of 3-5 was obtained by X-ray crystallographic analysis. For the synthesis of phosphonate derivative 11, the key step involved chlorination of phosphonate 9 by use of CF3SO2Cl and 1,8-diazabicyclo[5.4.0]undec-7-ene in CH2Cl2. 6-Chloropurine-containing butenolide 3d, 6-chloropurine derivative of 4-hydroxybutenolide 4, and adenine-containing 4-hydroxybutenolide 5 did not show anticancer and antiviral activities. 6-Chloropurine-containing ethylidene-2,3-dialkoxybutenolides 3a-c and phosphonate 11, however, exhibited inhibitory activity against murine leukemias (L1210 and P388), breast carcinoma (MCF7), and human T-lymphoblasts (Molt4/C8 and CEM/0) cell lines. They were also notably active toward thymidine kinase-deficient varicella-zoster virus (TK(-)VZV). Adenine-containing ethylidene-2,3-dimethoxybutenolide 6 exhibited marked selectivity in cytostatic activity against the murine leukemia (P388) cell line.
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PMID:Synthesis and biological evaluation of purine-containing butenolides. 1135 10

9-[(3-[18F]Fluoro-1-hydroxy-2-propoxy)methyl]guanine ([18F]FHPG, 2) has been synthesized by nucleophilic substitution of N(2)-(p-anisyldiphenylmethyl)-9-[[1-(p-anisyldiphenylmethoxy)-3-toluenesulfonyloxy-2-propoxy]methyl]guanine (1) with potassium [18F]fluoride/Kryptofix 2.2.2 followed by deprotection with 1 N HCl and purification with different methods in variable yields. When both the nucleophilic substitution and deprotection were carried out at 90 degrees C and the product was purified by HPLC (method A), the yield of compound 2 was 5-10% and the synthesis time was 90 min from EOB. However, if both the nucleophilic substitution and deprotection were carried out at 120 degrees C and the product was purified by HPLC, the yield of compound 2 decreased to 2%. When compound 2 was synthesized at 90 degrees C and purified by Silica Sep-Pak (method B), the yield increased to 10-15% and the synthesis time was 60 min from EOB. Similarly, 9-(4-[18F]fluoro-3-hydroxymethylbutyl)guanine ([18F]FHBG, 4) was synthesized with method A and method B in 9% and 10-15% yield, respectively, in a synthesis time of 90 and 60 min, respectively, from EOB. Compound 2 was relatively unstable in acidic medium at 120 degrees C while compound 4 was stable under the same condition. Both compound 2 and compound 4 had low lipid/water partition coefficient (0.126 +/- 0.022, n=5 and 0.165 +/- 0.023, n=5, respectively). Although it contains non-radioactive ganciclovir ( approximately 5-30 microg) as a chemical by-product, compound 2 synthesized by method B has a similar uptake in 9L glioma cells as that synthesized by method A, and is a potential tracer for imaging herpes simplex virus thymidine kinase gene expression in tumors using PET. Similarly, compound 4 synthesized by method B contains approximately 10-25 microg of penciclovir as a chemical by-product. Thus, the simplified one pot synthesis (method B) is a useful method for synthesizing both compound 2 and compound 4 in good yield for routine clinical use, and the method is readily amenable for automation.
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PMID:A simplified one-pot synthesis of 9-[(3-[18F]fluoro-1-hydroxy-2-propoxy)methyl]guanine([18F]FHPG) and 9-(4-[18F]fluoro-3-hydroxymethylbutyl)guanine ([18F]FHBG) for gene therapy. 1157 10