Faculty Profiles - UEDA Yuki

Papers - UEDA Yuki

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Ribavirin-induced down-regulation of CCAAT/enhancer-binding protein α leads to suppression of lipogenesis. Reviewed

Satoh S, Onomura D, Ueda Y, Dansako H, Honda M, Kaneko S, Kato N

The Biochemical journal 476 ( 1 ) 137 - 149 2019.1

Publishing type：Research paper (scientific journal) Publisher：Portland Press Ltd.

<title>Abstract</title>
Recently, we demonstrated that the anti-viral drug ribavirin (RBV) had the ability to suppress lipogenesis through down-regulation of retinoid X receptor α (RXRα) under the control of the intracellular GTP-level and AMP-activated protein kinase-related kinases, especially microtubule affinity regulating kinase 4 (MARK4). RXRα-overexpression attenuated but did not abolish lipogenesis suppression by RBV, implying that additional factor(s) were involved in this suppressive effect. In the present study, we found that the protein level, but not the mRNA level, of CCAAT/enhancer-binding protein α (C/EBPα) was down-regulated by RBV in hepatic cells. Treatment with proteasome inhibitor attenuated RBV-induced down-regulation of C/EBPα, suggesting that RBV promoted degradation of C/EBPα protein via the ubiquitin–proteasome pathway. Depletion of intracellular GTP through inosine monophosphate dehydrogenase inhibition by RBV led to down-regulation of C/EBPα. In contrast, down-regulation of C/EBPα by RBV was independent of RXRα and MARK4. Knockdown of C/EBPα reduced the intracellular neutral lipid levels and the expression of genes related to the triglyceride (TG) synthesis pathway, especially glycerol-3-phosphate acyltransferase, mitochondrial (GPAM), which encodes the first rate-limiting TG enzyme. Overexpression of C/EBPα yielded the opposite results. We also observed that RBV decreased GPAM expression. Moreover, overexpression of GPAM attenuated RBV-induced reduction in the intracellular neutral lipid levels. These data suggest that down-regulation of C/EBPα by RBV leads to the reduction in GPAM expression, which contributes to the suppression of lipogenesis. Our findings about the mechanism of RBV action in lipogenesis suppression will provide new insights for therapy against the active lipogenesis involved in hepatic steatosis and hepatocellular carcinomas.

DOI： 10.1042/BCJ20180680

Daunorubicin, a topoisomerase II poison, suppresses viral production of hepatitis B virus by inducing cGAS-dependent innate immune response. Reviewed International journal

Imai H, Dansako H, Ueda Y, Satoh S, Kato N

Biochemical and biophysical research communications 504 ( 4 ) 672 - 678 2018.10

Language：English Publishing type：Research paper (scientific journal)

Hepatitis B virus (HBV) causes hepatic diseases such as chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. These diseases are closely associated with persistent HBV infection. To prevent the progression of hepatic diseases, it is thus important to suppress persistent HBV infection. Daunorubicin (DNR), a topoisomerase II (Top II) poison, is a clinically used anticancer agent with a wide spectrum of activity against malignancies. DNR was recently reported to cause DNA damage-dependent interferon (IFN)-β induction through exogenous cyclic GMP-AMP synthetase (cGAS) and subsequently to suppress Ebola virus replication. In the present study, we demonstrated that DNR caused the inhibition of cell proliferation, but not cell death, through the DNA damage response in immortalized human hepatocyte NKNT-3/NTCP cells. Interestingly, DNR triggered the endogenous cGAS-dependent innate immune response and subsequently suppressed viral production of HBV in NKNT-3/NTCP cells. Top II poisons may be anti-HBV drug candidates.

DOI： 10.1016/j.bbrc.2018.08.195

Multiple antiviral activities of the antimalarial and anti-hepatitis C drug candidates N-89 and N-251. Reviewed International journal

Youki Ueda, Weilin Gu, Hiromichi Dansako, Hye-Sook Kim, Sayaka Yoshizaki, Nobuaki Okumura, Tomohiro Ishikawa, Hironori Nishitsuji, Fumihiro Kato, Takayuki Hishiki, Shinya Satoh, Koji Ishii, Michiaki Masuda, Kunitada Shimotohno, Masanori Ikeda, Nobuyuki Kato

Biochemistry and biophysics reports 15 1 - 6 2018.9

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal)

The chemically synthesized endoperoxide compound N-89 and its derivative N-251 were shown to have potent antimalarial activity. We previously demonstrated that N-89 and N-251 potently inhibited the RNA replication of hepatitis C virus (HCV), which belongs to the Flaviviridae family. Since antimalarial and anti-HCV mechanisms have not been clarified, we were interested whether N-89 and N-251 possessed the activity against viruses other than HCV. In this study, we examined the effects of N-89 and N-251 on other flaviviruses (dengue virus and Japanese encephalitis virus) and hepatitis viruses (hepatitis B virus and hepatitis E virus). Our findings revealed that N-89 and N-251 moderately inhibited the RNA replication of Japanese encephalitis virus and hepatitis E virus, although we could not detect those anti-dengue virus activities. We also observed that N-89 and N-251 moderately inhibited the replication of hepatitis B virus at the step after viral translation. These results suggest the possibility that N-89 and N-251 act on some common host factor(s) that are necessary for viral replications, rather than the possibility that N-89 and N-251 directly act on the viral proteins except for HCV. We describe a new type of antiviral reagents, N-89 and N-251, which are applicable to multiple different viruses.

DOI： 10.1016/j.bbrep.2018.05.007

ULBP1 is induced by hepatitis C virus infection and is the target of the NK cell-mediated innate immune response in human hepatocytes Reviewed

Hiromichi Dansako, Hirotaka Imai, Youki Ueda, Shinya Satoh, Takaji Wakita, Nobuyuki Kato

FEBS Open Bio 8 ( 3 ) 361 - 371 2018.3

Language：English Publishing type：Research paper (scientific journal) Publisher：Wiley Blackwell

Natural killer (NK) cells through their NK group 2 member D (NKG2D) receptors recognize NKG2D ligands such as UL16-binding proteins (ULBPs) on virus-infected cells and subsequently trigger the host innate immune response. In the present study, we demonstrated that hepatitis C virus (HCV) induced the cell surface expression of ULBP1 in human immortalized hepatocyte PH5CH8 cells and human hepatoma HuH-7 cell-derived RSc cells. Interestingly, NK cell line NK-92 induced cytotoxicity and interferon-γ mRNA expression and subsequently reduced the levels of HCV RNA replication during co-culture with HCV-infected RSc cells. From these results, we conclude that ULBP1 is a target of the NK cell-mediated innate immune response in HCV-infected human hepatocytes.

DOI： 10.1002/2211-5463.12373

Ribavirin suppresses hepatic lipogenesis through inosine monophosphate dehydrogenase inhibition: Involvement of adenosine monophosphate-activated protein kinase-related kinases and retinoid X receptor α. Reviewed

Satoh S, Mori K, Onomura D, Ueda Y, Dansako H, Honda M, Kaneko S, Ikeda M, Kato N

Hepatology communications 1 ( 6 ) 550 - 563 2017.8

Evaluation of preclinical antimalarial drugs, which can overcome direct acting antivirals-resistant hepatitis C viruses, using the viral reporter assay systems Reviewed

Youki Ueda, Hiromichi Dansako, Shinya Satoh, Hye-Sook Kim, Yusuke Wataya, Hiroyuki Doi, Masanori Ikeda, Nobuyuki Kato

VIRUS RESEARCH 235 37 - 48 2017.5

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：ELSEVIER SCIENCE BV

Persistent hepatitis C virus (HCV) infection causes chronic liver diseases and is a major global health problem. Recently developed treatments with direct-acting antivirals (DAAs) have largely improved the sustained virologic response rate of patients with chronic hepatitis C. However, this approach is still hindered by its great expense and the problem of drug resistance. Using our cell-based HCV assay systems, we reported that the preclinical antimalarial drugs N-89 and N-251 exhibited potent anti-HCV activities. In this study we used our assay systems to evaluate the anti-HCV activities of six kinds of DAAs individually or in combination with N-89 or N-251. The results showed that the DAAs had potent anti-HCV activities and N-89 or N-251 contributed additive or synergistic effect. Using DAA-resistant HCV-RNA-replicating cells, which were prepared by continuous treatment with each DAA, we demonstrated that N-89 and N-251 could overcome all of the DAA-resistant HCVs. These preclinical drugs would have been potential as components of a therapeutic regimen that also included combinations of various DAAs. In addition, sequence analysis of the NS3-NS5B regions of the DAA-resistant HCV genomes newly found several amino acid (aa) substitutions that were suggested to contribute to DAA-resistance in addition to the aa substitutions already known to cause DAA-resistance. Among these new aa substitutions, we found that two substitutions in the NS3 region (D79G and S174Y) contributed to simeprevirand/or asunaprevir-resistance.

DOI： 10.1016/j.virusres.2017.03.015

The cyclic GMP-AMP synthetase-STING signaling pathway is required for both the innate immune response against HBV and the suppression of HBV assembly

Hiromichi Dansako, Youki Ueda, Nobuaki Okumura, Shinya Satoh, Masaya Sugiyama, Masashi Mizokami, Masanori Ikeda, Nobuyuki Kato

FEBS JOURNAL 283 ( 1 ) 144 - 156 2016.1

Language：English Publishing type：Research paper (scientific journal) Publisher：WILEY-BLACKWELL

During viral replication, the innate immune response is induced through the recognition of viral replication intermediates by host factor(s). One of these host factors, cyclic GMP-AMP synthetase (cGAS), was recently reported to be involved in the recognition of viral DNA derived from DNA viruses. However, it is uncertain whether cGAS is involved in the recognition of hepatitis B virus (HBV), which is a hepatotropic DNA virus. In the present study, we demonstrated that HBV genome-derived double-stranded DNA induced the innate immune response through cGAS and its adaptor protein, stimulator of interferon genes (STING), in human hepatoma Li23 cells expressing high levels of cGAS. In addition, we demonstrated that HBV infection induced ISG56 through the cGAS-STING signaling pathway. This signaling pathway also showed an antiviral response towards HBV through the suppression of viral assembly. From these results, we conclude that the cGAS-STING signaling pathway is required for not only the innate immune response against HBV but also the suppression of HBV assembly. The cGAS-STING signaling pathway may thus be a novel target for anti-HBV strategies.

DOI： 10.1111/febs.13563

Synthesis and In Vitro Activity of Pyrrolo[3,4-d]pyrimidine-2,5-diones as Potential Non-nucleoside HCV Inhibitors Reviewed

Amany S. Mostafa, Serry A. El Bialy, Waleed A. Bayoumi, Youki Ueda, Masanori Ikeda, Nobuyuki Kato, Ali M. Abdelal

Current Enzyme Inhibition 12 ( 2 ) 170 - 176 2016

Establishment of Hepatitis C Virus RNA-Replicating Cell Lines Possessing Ribavirin-Resistant Phenotype

Shinya Satoh, Kyoko Mori, Youki Ueda, Hiroe Sejima, Hiromichi Dansako, Masanori Ikeda, Nobuyuki Kato

PLOS ONE 10 ( 2 ) e0118313 2015.2

Language：English Publishing type：Research paper (scientific journal) Publisher：PUBLIC LIBRARY SCIENCE

Background
Ribavirin (RBV) is a potential partner of interferon-based therapy and recently approved therapy using direct acting antivirals for patients with chronic hepatitis C. However, the precise mechanisms underlying RBV action against hepatitis C virus (HCV) replication are not yet understood. To clarify this point, we attempted to develop RBV-resistant cells from RBV-sensitive HCV RNA-replicating cells.
Methodology/Principal Findings
By repetitive RBV (100 mu M) treatment (10 weeks) of 3.5-year-cultured OL8 cells, in which genome-length HCV RNA (O strain of genotype 1b) efficiently replicates, dozens of colonies that survived RBV treatment were obtained. These colonies were mixed together and further treated with high doses of RBV (up to 200 mu M). By such RBV treatment, we successfully established 12 RBV-survived genome-length HCV RNA-replicating cell lines. Among them, three representative cell lines were characterized. HCV RNA replication in these cells resisted RBV significantly more than that in the parental OL8 cells. Genetic analysis of HCV found several common and conserved amino acid substitutions in HCV proteins among the three RBV-resistant cell species. Furthermore, using cDNA microarray and quantitative RT-PCR analyses, we identified 5 host genes whose expression levels were commonly altered by more than four-fold among these RBV-resistant cells compared with the parental cells. Moreover, to determine whether viral or host factor contributes to RBV resistance, we developed newly HCV RNA-replicating cells by introducing total RNAs isolated from RBV-sensitive parental cells or RBV-resistant cells into the HCV RNA-cured-parental or -RBV-resistant cells using an electroporation method, and evaluated the degrees of RBV resistance of these developed cells. Consequently, we found that RBV-resistant phenotype was conferred mainly by host factor and partially by viral factor.
Conclusions/Significance
These newly established HCV RNA-replicating cell lines should become useful tools for further understanding the anti-HCV mechanisms of RBV.

DOI： 10.1371/journal.pone.0118313

Synthesis of 3 ',4 '-difluoro-3 '-deoxyribonucleosides and its evaluation of the biological activities: Discovery of a novel type of anti-HCV agent 3 ',4 '-difluorocordycepin Reviewed

Hisashi Shimada, Kazuhiro Haraguchi, Kumi Hotta, Tomoko Miyaike, Yasuyuki Kitagawa, Hiromichi Tanaka, Ryutaro Kaneda, Hiroshi Abe, Satoshi Shuto, Kyoko Mori, Youki Ueda, Nobuyuki Kato, Robert Snoeck, Graciela Andrei, Jan Balzarini

BIOORGANIC & MEDICINAL CHEMISTRY 22 ( 21 ) 6174 - 6182 2014.11

Language：English Publishing type：Research paper (scientific journal) Publisher：PERGAMON-ELSEVIER SCIENCE LTD

Upon reacting 3',4'-unsaturated cytosine (8 and 9) and adenine nucleosides (13 and 14) with XeF2/BF3 center dot OEt2, the respective novel 3',4'-difluoro-3'-deoxyribofuranosyl nucleosides (10-12 and 15-18) could be obtained. Formation of anti-adducts (11, 16 and 18) revealed that the fluorination involved oxonium ions as incipient intermediates. TBDMS-protected 3',4'-unsaturated adenosine provided the beta-face adducts as sole stereoisomers whereas a-face-selectivity was observed with the TBDPS-protected adenosine 14. The evaluation of the novel 3'-deoxy-3',4'-difluororibofuranosylcytosine-(19-21) and adenine nucleosides (22-25) against antitumor and antiviral activities revealed that 3',4'-difluorocordycepin (24) was found to possess anti-HCV activity. The SI of 24 was comparable to that of the anti-HCV drug ribavirin. However, sofosbuvir, FDA-approved novel anti-HCV drug, showed better SI value. Our finding revealed that the introduction of the fluoro-substituent into the 4'-position of cordycepin derivatives decreased the cytotoxicity to the host cell with retention of the antiviral activity. (C) 2014 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmc.2014.08.024

Synthesis and inhibitory activity on hepatitis C virus RNA replication of 4-(1,1,1,3,3,3-hexafluoro-2-hydroxy-2-propyl)aniline analogs Reviewed

Kenji Matsuno, Youki Ueda, Miwa Fukuda, Kenji Onoda, Minoru Waki, Masanori Ikeda, Nobuyuki Kato, Hiroyuki Miyachi

BIOORGANIC & MEDICINAL CHEMISTRY LETTERS 24 ( 17 ) 4276 - 4280 2014.9

Language：English Publishing type：Research paper (scientific journal) Publisher：PERGAMON-ELSEVIER SCIENCE LTD

Using our recently developed assay system for full-genome-length hepatitis C virus (HCV) RNA replication in human hepatoma-derived Li23 cells (ORL8), we identified 4-(1,1,1,3,3,3-hexafluoro-2-hydroxy-2-propyl)aniline analog la as a novel HCV inhibitor. Structural modifications of la provided a series of sulfonamides 7 with much more potent HCV RNA replication-inhibitory activity than ribavirin. Compound 7a showed an additive anti-HCV effect in combination with standard anti-HCV therapy (IFN-alpha plus ribavirin). Since 7a generated reactive oxygen species (ROS) in the ORL8 system and its anti-HCV activity was blocked by vitamin E, its anti-HCV activity may be mediated at least in part by ROS. (C) 2014 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmcl.2014.07.019

Anti-HCV activity of the Chinese medicinal fungus Cordyceps militaris Reviewed

Youki Ueda, Kyoko Mori, Shinya Satoh, Hiromichi Dansako, Masanori Ikeda, Nobuyuki Kato

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 447 ( 2 ) 341 - 345 2014.5

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：ACADEMIC PRESS INC ELSEVIER SCIENCE

Persistent hepatitis C virus (HCV) infection causes chronic liver diseases and is a global health problem. Although the sustained virologic response rate in the treatment of genotype 1 using new triple therapy (pegylated-interferon, ribavirin, and telaprevir/boceprevir) has been improved by more than 70%, several severe side effects such as skin rash/ageusia and advanced anemia have become a problem. Under these circumstances, a new type of anti-HCV oral drug with few side effects is needed. Our recently developed HCV drug assay systems, including the HuH-7 cell line-derived OR6 and AH1R, and the Li23 cell line derived ORL8 and ORL11, allow genome-length HCV RNAs (several strains of genotype 1b) encoding renilla luciferase to replicate efficiently. Using these systems as anti-HCV candidates, we have identified numerous existing medicines that can be used against HCV with few side effects, such as statins and teprenon. To obtain additional anti-HCV candidates, we evaluated a number of oral health supplements, and found that the capsule but not the liquid form of Cordyceps militaris (CM) (Ascomycotinanorth, North Chinese caterpillar fungus), which is used as a Chinese herbal medicine, exhibited moderate anti-HCV activity. In combination with interferon-a or ribavirin, CM exhibited an additive inhibitory effect. Among the main components of CM, cordycepin, but not ergosterol, contributed to the anti-HCV activity of CM. In consideration of all these results, we suggest that CM would be useful as an oral anti-HCV agent in combination with interferon-alpha and/or ribavirin. (C) 2014 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2014.03.150

Genetic Characterization of Hepatitis C Virus in Long-Term RNA Replication Using Li23 Cell Culture Systems Reviewed

Nobuyuki Kato, Hiroe Sejima, Youki Ueda, Kyoko Mori, Shinya Satoh, Hiromichi Dansako, Masanori Ikeda

PLOS ONE 9 ( 3 ) 91156 - 91156 2014.3

Language：English Publishing type：Research paper (scientific journal) Publisher：PUBLIC LIBRARY SCIENCE

Background: The most distinguishing genetic feature of hepatitis C virus (HCV) is its remarkable diversity and variation. To understand this feature, we previously performed genetic analysis of HCV in the long-term culture of human hepatoma HuH-7-derived HCV RNA-replicating cell lines. On the other hand, we newly established HCV RNA-replicating cell lines using human hepatoma Li23 cells, which were distinct from HuH-7 cells.
Methodology/Principal Findings: Li23-derived HCV RNA-replicating cells were cultured for 4 years. We performed genetic analysis of HCVs recovered from these cells at 0, 2, and 4 years in culture. Most analysis was performed in two separate parts: one part covered from the 5'-terminus to NS2, which is mostly nonessential for RNA replication, and the other part covered from NS3 to NS5B, which is essential for RNA replication. Genetic mutations in both regions accumulated in a timedependent manner, and the mutation rates in the 5'-terminus-NS2 and NS3-NS5B regions were 4.0-9.0x10(-3) and 2.7-4.0x10(-3) base substitutions/site/year, respectively. These results suggest that the variation in the NS3-NS5B regions is affected by the pressure of RNA replication. Several in-frame deletions (3-105 nucleotides) were detected in the structural regions of HCV RNAs obtained from 2-year or 4-year cultured cells. Phylogenetic tree analyses clearly showed that the genetic diversity of HCV was expanded in a time-dependent manner. The GC content of HCV RNA was significantly increased in a time-dependent manner, as previously observed in HuH-7-derived cell systems. This phenomenon was partially due to the alterations in codon usages for codon optimization in human cells. Furthermore, we demonstrated that these long-term cultured cells were useful as a source for the selection of HCV clones showing resistance to anti-HCV agents.
Conclusions/Significance: Long-term cultured HCV RNA-replicating cells are useful for the analysis of evolutionary dynamics and variations of HCV and for drug-resistance analysis.

DOI： 10.1371/journal.pone.0091156

Peroxisome proliferator-activated receptor delta antagonists inhibit hepatitis C virus RNA replication Reviewed

Shintaro Ban, Youki Ueda, Masao Ohashi, Kenji Matsuno, Masanori Ikeda, Nobuyuki Kato, Hiroyuki Miyachi

Bioorganic and Medicinal Chemistry Letters 23 ( 17 ) 4774 - 4778 2013.9

Language：English Publishing type：Research paper (scientific journal)

It has been reported that ligand-mediated transcription factor peroxisome proliferator-activated receptor alpha (hPPARα) is involved in hepatitis C virus (HCV) RNA replication, whereas hPPARγ is not, and the effect of hPPARδ is unknown. Here, we show that hPPARδ-selective antagonists effectively inhibit HCV RNA replication. We describe the design, synthesis and pharmacological evaluation of a series of biphenyl-4-carboxylic acid-type hPPARδ antagonists, including previously reported compounds, as candidate anti-HCV agents. A representative compound (4c) dose-dependently inhibited HCV RNA replication (EC50 0.22 μM), while exhibiting relatively weak cytotoxicity to the host cells (CC50 2.5 μM). It also showed an additive and dose-dependent effect on the inhibition of HCV RNA replication by pegylated interferon alpha (Peg-IFNα) alone and by both Peg-IFNα and ribavirin (currently the clinical treatment of choice for HCV infection). Thus, combination of a hPPARδ antagonist with current therapy may improve the efficacy of treatment for HCV infection. © 2013 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.bmcl.2013.07.005

New Preclinical Antimalarial Drugs Potently Inhibit Hepatitis C Virus Genotype 1b RNA Replication Reviewed

Youki Ueda, Midori Takeda, Kyoko Mori, Hiromichi Dansako, Takaji Wakita, Hye-Sook Kim, Akira Sato, Yusuke Wataya, Masanori Ikeda, Nobuyuki Kato

PLOS ONE 8 ( 8 ) e72519 2013.8

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：PUBLIC LIBRARY SCIENCE

Background: Persistent hepatitis C virus (HCV) infection causes chronic liver diseases and is a global health problem. Although new triple therapy (pegylated-interferon, ribavirin, and telaprevir/boceprevir) has recently been started and is expected to achieve a sustained virologic response of more than 70% in HCV genotype 1 patients, there are several problems to be resolved, including skin rash/ageusia and advanced anemia. Thus a new type of anti-HCV drug is still needed.
Methodology/Principal Findings: Recently developed HCV drug assay systems using HCV-RNA-replicating cells (e.g., HuH-7-derived OR6 and Li23-derived ORL8) were used to evaluate the anti-HCV activity of drug candidates. During the course of the evaluation of anti-HCV candidates, we unexpectedly found that two preclinical antimalarial drugs (N-89 and its derivative N-251) showed potent anti-HCV activities at tens of nanomolar concentrations irrespective of the cell lines and HCV strains of genotype 1b. We confirmed that replication of authentic HCV-RNA was inhibited by these drugs. Interestingly, however, this anti-HCV activity did not work for JFH-1 strain of genotype 2a. We demonstrated that HCV-RNA-replicating cells were cured by treatment with only N-89. A comparative time course assay using N-89 and interferon-alpha demonstrated that N-89-treated ORL8 cells had more rapid anti-HCV kinetics than did interferon-alpha-treated cells. This anti-HCV activity was largely canceled by vitamin E. In combination with interferon-alpha and/or ribavirin, N-89 or N-251 exhibited a synergistic inhibitory effect.
Conclusions/Significance: We found that the preclinical antimalarial drugs N-89 and N-251 exhibited very fast and potent anti-HCV activities using cell-based HCV-RNA-replication assay systems. N-89 and N-251 may be useful as a new type of anti-HCV reagents when used singly or in combination with interferon and/or ribavirin.

DOI： 10.1371/journal.pone.0072519

Development of a drug assay system with hepatitis C virus genome derived from a patient with acute hepatitis C Reviewed

Kyoko Mori, Youki Ueda, Yasuo Ariumi, Hiromichi Dansako, Masanori Ikeda, Nobuyuki Kato

VIRUS GENES 44 ( 3 ) 374 - 381 2012.6

Language：English Publishing type：Research paper (scientific journal) Publisher：SPRINGER

We developed a new cell culture drug assay system (AH1R), in which genome-length hepatitis C virus (HCV) RNA (AH1 strain of genotype 1b derived from a patient with acute hepatitis C) efficiently replicates. By comparing the AH1R system with the OR6 assay system that we developed previously (O strain of genotype 1b derived from an HCV-positive blood donor), we demonstrated that the anti-HCV profiles of reagents including interferon-gamma and cyclosporine A significantly differed between these assay systems. Furthermore, we found unexpectedly that rolipram, an anti-inflammatory drug, showed anti-HCV activity in the AH1R assay but not in the OR6 assay, suggesting that the anti-HCV activity of rolipram differs depending on the HCV strain. Taken together, these results suggest that the AH1R assay system is useful for the objective evaluation of anti-HCV reagents and for the discovery of different classes of anti-HCV reagents.

DOI： 10.1007/s11262-012-0712-2

Plural assay systems derived from different cell lines and hepatitis C virus strains are required for the objective evaluation of anti-hepatitis C virus reagents Reviewed

Youki Ueda, Kyoko Mori, Yasuo Ariumi, Masanori Ikeda, Nobuyuki Kato

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 409 ( 4 ) 663 - 668 2011.6

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：ACADEMIC PRESS INC ELSEVIER SCIENCE

Persistent hepatitis C virus (HCV) infection causes chronic liver diseases and is a global health problem. HuH-7 hepatoma-derived cells are widely used as the only cell-based HCV replication system for HCV research, including drug assays. Recently, using different hepatoma Li23-derived cells, we developed an HCV drug assay system (ORL8), in which the genome-length HCV RNA (O strain of genotype 1b) encoding renilla luciferase replicates efficiently. In this study, using the HuH-7-derived OR6 assay system that we developed previously and the ORL8 assay system, we evaluated 26 anti-HCV reagents, which other groups had reported as anti-HCV candidates using HuH-7-derived assay systems other than ORB. The results revealed that more than half of the reagents showed different anti-HCV activities from those in the previous studies, and that anti-HCV activities evaluated by the ORB and ORL8 assays were also frequently different. In further evaluation using the HuH-7-derived AH1R assay system, which was developed using the AH1 strain of genotype 1b, several reagents showed different anti-HCV activities in comparison with those evaluated by the OR6 and ORL8 assays. These results suggest that the different activities of anti-HCV reagents are caused by the differences in cell lines or HCV strains used for the development of assay systems. Therefore, we conclude that plural HCV assay systems developed using different cell lines or HCV strains are required for the objective evaluation of anti-HCV reagents. (C) 2011 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2011.05.061

Mechanism of freezing of water in contact with mesoporous silicas MCM-41, SBA-15 and SBA-16: role of boundary water of pore outlets in freezing Reviewed

Shigeharu Kittaka, Yuki Ueda, Fumika Fujisaki, Taku Iiyama, Toshio Yamaguchi

PHYSICAL CHEMISTRY CHEMICAL PHYSICS 13 ( 38 ) 17222 - 17233 2011

Language：English Publishing type：Research paper (scientific journal) Publisher：ROYAL SOC CHEMISTRY

The freezing mechanism of water contacted with mesoporous silicas with uniform pore shapes, both cylindrical and cagelike, was studied by thermodynamic and structural analyses with differential scanning calorimetry (DSC) and X-ray diffraction (XRD) together with adsorption measurements. In the DSC data extra exothermic peaks were found at around 230 K for water confined in SBA-15, in addition to that due to the freezing of pore water. These peaks are most likely to be ascribed to the freezing of water present over the micropore and/or mesopore outlets of coronas in SBA-15. Freezing of water confined in SBA-16 was systematically analysed by DSC with changing the pore size. The freezing temperature was found to be around 232 K, close to the homogeneous nucleation temperature of bulk water, independent of the pore size when the pore diameter (d) < 7.0 nm. Water confined in the cagelike pores of SBA-16 is probably surrounded by a water layer (boundary water) at the outlets of channels to interconnect the pores and of fine corona-like pores, which is similar to that present at the outlet of cylindrical pores in MCM-41 and of cylindrical channels in SBA-15. The presence of the boundary water would be a key for water in SBA-16 to freeze at the homogeneous nucleation temperature. This phenomenon is similar to those well known for water droplets in oil and water droplets of clouds in the sky. The XRD data showed that the cubic ice I(c) was formed in SBA-16 as previously found in SBA-15 when d < 8.0 nm.

DOI： 10.1039/c1cp21458f

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