Hope for Cirrhosis Patients with Genotype 1 Hepatitis C Virus Who Failed the Previous Treatment / 대한소화기학회지

No abstract available.

Biochemical properties of full-length hepatitis C virus RNA-dependent RNA polymerase expressed in insect cells

The hepatitis C virus (HCV) RNA-dependent RNA polymerase, NS5B protein, is the key viral enzyme responsible for replication of the HCV viral RNA genome. Although several full-length and truncated forms of the HCV NS5B proteins have been expressed previously in insect cells, contamination of host terminal transferase (TNTase) has hampered analysis of the RNA synthesis initiation mechanism using natural HCV RNA templates. We have expressed the HCV NS5B protein in insect cells using a recombinant baculovirus and purified it to near homogeneity without contaminated TNTase. The highly purified recombinant HCV NS5B was capable of copying 9.6-kb full-length HCV RNA template, and mini-HCV RNA carrying both 5'- and 3'-untranslated regions (UTRs) of the HCV genome. In the absence of a primer, and other cellular and viral factors, the NS5B could elongate over HCV RNA templates, but the synthesized products were primarily in the double stranded form, indicating that no cyclic replication occurred with NS5B alone. RNA synthesis using RNA templates representing the 3'-end region of HCV minus-strand RNA and the X-RNA at the 3'-end of HCV RNA genome was also initiated de novo. No formation of dimersize self-primed RNA products resulting from extension of the 3'-end hydroxyl group was observed. Despite the internal de novo initiation from the X-RNA, the NS5B could not initiate RNA synthesis from the internal region of oligouridylic acid (U)20, suggesting that HCV RNA polymerase initiates RNA synthesis from the selected region in the 3'-UTR of HCV genome.

A molecular dynamics study based post facto free energy analysis of the binding of bovine angiogenin with UMP and CMP ligands.

Angiogenin is a protein belonging to the superfamily of RNase A. The RNase activity of this protein is essential for its angiogenic activity. Although members of the RNase A family carry out RNase activity, they differ markedly in their strength and specificity. In this paper, we address the problem of higher specificity of angiogenin towards cytosine against uracil in the first base binding position. We have carried out extensive nano-second level molecular dynamics(MD) computer simulations on the native bovine angiogenin and on the CMP and UMP complexes of this protein in aqueous medium with explicit molecular solvent. The structures thus generated were subjected to a rigorous free energy component analysis to arrive at a plausible molecular thermodynamic explanation for the substrate specificity of angiogenin.

Leflunomide: A New Disease Modifying Anti-Rheumatic Drug / 대한류마티스학회지

Leflunomide is a novel, isoxazol based disease-modifying anti-rheumatic drug (DMARD) for the treatment of rheumatoid arthritis (RA). Its mechanism differs from other DMARDs in that it inhibits de novo pyrimidine synthesis by inhibiting the enzyme dihydroorotate dehydrogenase (DHODH). It is a pro-drug and undergoes rapid conversion to its active form in vivo, A77-1726. A77-1726 inhibits the mitochondrial enzyme DHODH, which plays a key role in the de novo synthesis of pyrimidine ribonucleotide uridine monophosphate (rUMP). Leflunomide prevents clonal expansion of activated lymphocytes by interfering with the cell cycle progression due to inadequate production of rUMP and utilizing mechanisms involving p53. The relative lack of toxicity of A77-1726 on non-lymphoid cells may be due to the ability of these cells to fulfill their ribonucleotide requirements by use of salvage pyrimidine pathway, which makes them less dependent on de novo synthesis. Several phase II clinical trials of patients with RA revealed that leflunomide was effective and well tolerated. Large-scale phase III clinical trials have shown that leflunomide (20mg/day) provided a statistically significant clinical benefit and prevention of radiographic progression in comparison to placebo. The clinical benefits of leflunomide were similar to or greater than methotrexate and sulfasalazine. Now, many multi-national studies are in progress and planning, including combination therapy with other DMARD. In future, those studies will provide us more information about the effectiveness and potential adverse effect of leflunomide.

Leflunomide: A New Disease Modifying Anti-Rheumatic Drug / 대한류마티스학회지

Leflunomide is a novel, isoxazol based disease-modifying anti-rheumatic drug (DMARD) for the treatment of rheumatoid arthritis (RA). Its mechanism differs from other DMARDs in that it inhibits de novo pyrimidine synthesis by inhibiting the enzyme dihydroorotate dehydrogenase (DHODH). It is a pro-drug and undergoes rapid conversion to its active form in vivo, A77-1726. A77-1726 inhibits the mitochondrial enzyme DHODH, which plays a key role in the de novo synthesis of pyrimidine ribonucleotide uridine monophosphate (rUMP). Leflunomide prevents clonal expansion of activated lymphocytes by interfering with the cell cycle progression due to inadequate production of rUMP and utilizing mechanisms involving p53. The relative lack of toxicity of A77-1726 on non-lymphoid cells may be due to the ability of these cells to fulfill their ribonucleotide requirements by use of salvage pyrimidine pathway, which makes them less dependent on de novo synthesis. Several phase II clinical trials of patients with RA revealed that leflunomide was effective and well tolerated. Large-scale phase III clinical trials have shown that leflunomide (20mg/day) provided a statistically significant clinical benefit and prevention of radiographic progression in comparison to placebo. The clinical benefits of leflunomide were similar to or greater than methotrexate and sulfasalazine. Now, many multi-national studies are in progress and planning, including combination therapy with other DMARD. In future, those studies will provide us more information about the effectiveness and potential adverse effect of leflunomide.

Effects of Uridine 5'-Triphosphate on the Vascular Tone of Rat Thoracic Aorta

BACKGROUND: Uracil nucleotides are stored in platelets and all other cells, and are released into the extracellular space upon stimulation. They show various biological responses but their actions and mechanism are not well understood. This study was conducted to investigate the effects of uridine 5'-triphosphate(UTP) on vascular tone and to identify the characteristics of their receptors. METHODS: Aortic ring preparation were made from the rat descending thoracic aorta. Endo-thelial cells were preserved or removed by gentle rubbing, The basal tension of aortic ring was lgm and isometric contraction were recorded on polygraph using force transducer. RESULTS: In aortic ring Precontracted by 100nM norepinephrine, UTP induced dual effect with various concentrations. UTP elicited endothelium-dependent relaxation at low concentrations(100nM-10microM), and endothelium-independent contraction at high concentrations(more than 30microM). Among uracil nucleotides, UDP was as much effective as UTP in vascular tone, but UMP and uridine were not. UTP(pA50 6.15) was more potent than ATP(5.17), ITP(4.75) and other nucleotides(TTP, GTP, CTP). At basal tension, UTP induced relaxation at low concentrations and contraction at hige concentrations in endothelium-intact ring. But in endothelium-removed ring, UTP elicited only contraction. Prior treatment of aortic ring with suramin, a non-selective P2-purinoceptor blocker, inhibited UTP-Induced relaxation and contraction. Reactive blue-2, a P2gamma purinoceptor blocker, inhibited relaxation only, but alpha, beta-methylene ATP, a P2x Purinoceptor blocker, enhanced contractile response. ATP inhibited the UPT-induced relaxation, but 2-methylthio ATP did not alter the effects of UTP. It means that UTP and ATP act at the same receptor but 2-methylthio ATP does not. CONCLUSION: These results suggest that UTP-induced relaxation is mediated by nucleotide receptors on endothelium and the contraction is mediated by pyrimidinoceptors on vascular smooth muscle.

Microenvironment at the substrate binding subsite of the active site of UDPglucose 4-epimerase from Kluyveromyces fragilis using a fluorescent analog of UMP.

A chromophorics and fluorescent analog of uridine 5'-monophosphate (UMP), a known competitive inhibitor of UDPglucose 4-epimerase was synthesised. This analog, namely 2',3'-O-(2,4,6-trinitrocyclohexadienylidene) uridine 5'-monophosphate, was found to be a powerful reversible inhibitor of UDPglucose 4-epimerase indicating its interaction with the substrate binding site of the enzyme. The extreme sensitivity of the fluorescence emission spectrum of this analog to solvent polarity makes it an excellent probe for the study of the environment at the active site of the enzyme. We report here the effective use of this UMP analog to demonstrate that the hydroxyl groups of the ribose moiety of UMP and presumably the substrates (UDPgalactose and UDPglucose) do not reside in a hydrophobic milieu.