Jin Soo Shin

Inhibition of influenza virus internalization by (−)-epigallocatechin-3-gallate

(-)-Epigallocatechin-3-gallate (EGCG), one of the major flavonoid components of green tea, is known to have a broad antiviral activity against several enveloped viruses, including the influenza virus. However, its mode of action and the mechanism that allows it to target influenza virus molecules have not been fully elucidated. Thus, this study investigated the molecular mechanism by which EGCG suppresses influenza virus infections. EGCG was found to block an early step in the influenza viral life cycle, but it did not affect viral adsorption to target cells or viral RNA replication. However, EGCG inhibited hemifusion events between virus particles and the cellular membrane by reducing the viral membrane integrity, thereby resulting in the loss of the cell penetration capacity of the influenza virus. EGCG also marginally suppressed the viral and nonviral neuraminidase (NA) activity in an enzyme-based assay system. In conclusion, it is suggested that the anti-influenza viral efficacy of EGCG is attributable to damage to the physical properties of the viral envelope and partial inhibition of the NA surface glycoprotein. These results may facilitate future investigations of the antiviral activity of EGCG against other enveloped viruses as well as influenza virus.

Protection of pregnant mice, fetuses and neonates from lethality of H5N1 influenza viruses by maternal vaccination.

The highly pathogenic H5N1 influenza viruses are one of candidates for the next pandemic. Information on protective immunity for pregnant animals by vaccination against the H5N1 influenza virus is limited. Here, we show that the immunization of pregnant mice with inactivated H5N1 influenza vaccine protects them, their fetuses, and their infant mice from H5N1 influenza viruses. Pregnant mice immunized with two doses of H5N1 influenza vaccine were protected from homologous infections of H5N1 influenza viruses with no viruses detected in fetuses, and that they were protected upto 30% from heterologous infections of H5N1 influenza viruses with viruses detected in fetuses. The infant mice born to mothers immunized with H5N1 influenza vaccine were fully protected from infections of H5N1 influenza viruses for upto 4 weeks of age. The protection of infant mice was closely related to the presence of IgG2a antibody in lung, heart, and rectum tissues. Our results suggest that maternal vaccination may be critical for protecting pregnant animals, their fetuses, and their infant mice from lethal infections of H5N1 influenza viruses.

Identification and evaluation of potent Middle East respiratory syndrome coronavirus (MERS-CoV) 3CLPro inhibitors

ABSTRACT Middle East respiratory syndrome coronavirus (MERS‐CoV) causes severe acute respiratory illness with fever, cough and shortness of breath. Up to date, it has resulted in 1826 human infections, including 649 deaths. Analogous to picornavirus 3C protease (3Cpro), 3C‐like protease (3CLpro) is critical for initiation of the MERS‐CoV replication cycle and is thus regarded as a validated drug target. As presented here, our peptidomimetic inhibitors of enterovirus 3Cpro (6b, 6c and 6d) inhibited 3CLpro of MERS‐CoV and severe acute respiratory syndrome coronavirus (SARS‐CoV) with IC50 values ranging from 1.7 to 4.7 &mgr;M and from 0.2 to 0.7 &mgr;M, respectively. In MERS‐CoV‐infected cells, the inhibitors showed antiviral activity with EC50 values ranging from 0.6 to 1.4 &mgr;M, by downregulating the viral protein production in cells as well as reducing secretion of infectious viral particles into culture supernatants. They also suppressed other &agr;‐ and &bgr;‐CoVs from human and feline origin. These compounds exhibited good selectivity index (over 70 against MERS‐CoV) and could lead to the development of broad‐spectrum antiviral drugs against emerging CoVs and picornaviruses. Graphical abstract Figure. No Caption available. HighlightsAldehyde‐containing peptidomimetics were identified to be potent inhibitors against MERS‐CoV 3CLpro.The active inhibitor showed sub‐&mgr;M EC50 in killing MERS‐CoV.Compounds were also effective against other &agr; and &bgr;‐CoVs of both human and feline origin.We identified broad‐spectrum antiviral agents effective against both coronaviruses and picornaviruses.

Phylogenic analysis of reassorted avian influenza viruses isolated from Korean domestic ducks from 2005 to 2007

Ducks have been regarded as animals that can perpetuate most avian influenza viruses since they generally do not show the clear clinical signs such as death and reduced body weight when they are infected. Here, we characterized two H3N2 and one H3N6 avian influenza viruses isolated from ducks on the local farms in Korea from 2005 to 2007. Genetic analysis of these viruses showed that most segments of isolates except NP genes belonged to Eurasian lineage. NP genes of two H3N2 isolates, A/Duck/Korea/S71/07, and A/Duck/Korea/S72/07 belonged to North American lineage. Our results suggest that the genetic reassortment among avian influenza viruses can occur in domestic ducks.

A Novel Series of Highly Potent Small Molecule Inhibitors of Rhinovirus Replication

Human rhinoviruses (hRVs) are the main causative pathogen for common colds and are associated with the exacerbation of asthma. The wide variety in hRV serotypes has complicated the development of rhinovirus replication inhibitors. In the current investigation, we developed a novel series of benzothiophene derivatives and their analogues (6-8) that potently inhibit the replication of both hRV-A and hRV-B strains. Compound 6g inhibited the replication of hRV-B14, A21, and A71, with respective EC50 values of 0.083, 0.078, and 0.015 μM. The results of a time-of-addition study against hRV-B14 and hRV-A16 and resistant mutation analysis on hRV-B14 implied that 6g acts at the early stage of the viral replication process, interacting with viral capsid protein. A molecular docking study suggested that 6g has a capsid-binding mode similar to that of pleconaril. Finally, derivatives of 6 also displayed significant inhibition against poliovirus 3 (PV3) replication, implying their potential inhibitory activities against other enterovirus species.

Protection of ferrets from infection by swine-origin 2009 A (H1N1) influenza virus by the inactivated vaccine.

Swine-origin pandemic 2009 A (H1N1) influenza viruses are still infecting humans, and humans are currently being vaccinated with the inactivated vaccine of 2009 A (H1N1) influenza virus. We wanted to determine the efficacy of 2009 A (H1N1) inactivated vaccine in ferrets. Ferrets immunized with one dose (7.5 microg) of 2009 A (H1N1) inactivated vaccine were not protected from infections of either pandemic H1N1 or seasonal H1N1 influenza viruses, while ferrets immunized with two doses of 2009 A (H1N1) inactivated vaccine were protected from infections of pandemic H1N1, but not seasonal H1N1 influenza viruses. IgG subtype of antibody was dominantly detected in tissues of immunized ferrets. Our study suggests that pandemic H1N1 vaccine may not elicit the antibody cross-reactive to the seasonal H1N1 influenza virus.

IgG Antibodies Mediate Protective Immunity of Inactivated Vaccine for Highly Pathogenic H5N1 Influenza Viruses in Ferrets

Outbreaks of highly pathogenic H5N1 influenza viruses have been reported in many countries worldwide. The possibility of pandemics caused by H5N1 influenza viruses is high since human infections by H5N1 viruses continually occur. In this study we determined the immune response and efficacy of inactivated H5N1 vaccine developed by reverse genetics in ferrets. Ferrets intramuscularly inoculated with two doses of H5N1 vaccine survived the lethal challenge with homologous or heterologous H5N1 influenza viruses, while 75% and 25% of ferrets immunized with one dose of H5N1 vaccine survived the lethal challenge with homologous and heterologous H5N1 influenza viruses, respectively. When we determined antibody subtypes specific for H5N1 influenza viruses in tissues and sera of vaccinated ferrets, IgG antibodies were detected mainly in the trachea, nostril, lung, heart, liver, kidney, intestine, spleen, and serum. Our results suggest that IgG antibodies may play a major role in protecting ferrets immunized with the inactivated H5N1 vaccine from lethal challenge with H5N1 influenza viruses.

Comparison of anti-influenza virus activity and pharmacokinetics of oseltamivir free base and oseltamivir phosphate

Influenza viruses are major human respiratory pathogens that cause high morbidity and mortality worldwide. Currently, prophylactic vaccines and therapeutic antiviral agents are used to prevent and control influenza virus infection. Oseltamivir free base (OSV-FB), a modified generic antiviral drug of Tamiflu (oseltamivir phosphate, OSV-P), was launched in the Republic of Korea last year. Here, we examine the bioequivalence of these two compounds by assessing their antiviral efficacy in infected cells and in a mouse model. It was observed that both antivirals showed comparable efficacy against 11 different influenza A and B viruses in vitro. Moreover, in mice infected with influenza A virus (mouse-adapted A/Puerto Rico/8/34), they showed a dose-dependent therapeutic activity and alleviated infection-mediated reductions in body weight, leading to significantly better survival. There was histopathological disappearance of virus-induced inflammatory cell infiltration of the lung after oral treatment with either antiviral agent (at 10 mg/kg). Pharmacokinetic analysis also exhibited similar plasma concentrations of the active drug, oseltamivir carboxylate, metabolised from both OSV-B and OSV-P. This is the first report showing bioequivalence of OSV-FB to its phosphate salt form in the mouse system. The free base drug has some beneficial points including simple drug formulation process and reduced risk of undesirable cation-phosphate precipitation within solution. The long term stability of OSV-FB requires further monitoring when it is provided as a national stock in readiness for an influenza pandemic.

Systematic editing of synthetic RIG-I ligands to produce effective antiviral and anti-tumor RNA immunotherapies

Abstract Retinoic acid-inducible gene I (RIG-I) recognizes double-stranded viral RNAs (dsRNAs) containing two or three 5′ phosphates. A few reports of 5′-PPP-independent RIG-I agonists have emerged, but little is known about the molecular principles underlying their recognition. We recently found that the bent duplex RNA from the influenza A panhandle promoter activates RIG-I even in the absence of a 5′-triphosphate moiety. Here, we report that non-canonical synthetic RNA oligonucleotides containing G-U wobble base pairs that form a bent helix can exert RIG-I-mediated antiviral and anti-tumor effects in a sequence- and site-dependent manner. We present synthetic RNAs that have been systematically modified to enhance their efficacy and we outline the basic principles for engineering RIG-I agonists applicable to immunotherapy.

Enterovirus inhibitory activity of C-8-tert-butyl substituted 4-aryl-6,7,8,9-tetrahydrobenzo[4,5]thieno[3,2-e][1,2,4]triazolo[4,3-a]pyrimidin-5(4H)-ones

Members of a series of 4-aryl-6,7,8,9-tetrahydrobenzo[4,5]thieno[3,2-e][1,2,4]triazolo[4,3-a]pyrimidin-5(4H)-ones (1, Fig. 2) were prepared and tested against representative enteroviruses including Human Coxsackievirus B1 (Cox B1), Human Coxsackievirus B3 (Cox B3), human Poliovirus 3 (PV3), human Rhinovirus 14 (HRV14), human Rhinovirus 21 (HRV 21) and human Rhinovirus 71 (HRV 71). The C-8-tert-butyl group on the tetrahydrobenzene ring in these substances was found to be crucial for their enterovirus activity. One member of this group, 1e, showed single digit micromolar activities (1.6-8.85μM) against a spectrum of viruses screened, and the highest selectivity index (SI) values for Cox B1 (>11.2), for Cox B3 (>11.5), and for PV3 (>51.2), respectively. In contrast, 1p, was the most active analog against the selected HRVs (1.8-2.6μM), and showed the highest selectivity indices among the group of compounds tested. The SI values for 1p were 11.5 for HRV14, 8.4 for HRV21, and 12.1 for HRV71, respectively.