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Scientific Publications

Boehringer Ingelheim offers an overview of scientific publications on the following pages. This overview represents all publications of the last three years (YTD) where employees of Boehringer Ingelheim worldwide were involved.

49 publications regarding Virology
  • Author:
    Lemke CT; Titolo S; von Schwedler U; Goudreau N; Mercier J-F; Wardrop E; Faucher A-M; Coulombe R; Banik SSR; Fader L; Gagnon A; Kawai SH; Rancourt J; Tremblay M; Yoakim C; Simoneau B; Archambault J; Mason SW; Sundquist WI
    Title:
    Distinct effect of two HIV-1 capsid assembly inhibitor families that bind the same site within the N-termninal domain of the viral CA protein.
    Source:
    J Virol 86 (12), 6643-6655 (2012)
    Abstract:
    The emergence of resistance to existing classes of antiretroviral drugs necessitates finding new HIV-1 targets for drug discovery. The viral capsid (CA) protein represents one such potential new target. CA is sufficient to form mature HIV-1 capsids in vitro, and extensive structure-function and mutational analyses of CA have shown that the proper assembly, morphology, and stability of the mature capsid core are essential for the infectivity of HIV-1 virions. Here we describe the development of an in vitro capsid assembly assay based on the association of CA-NC subunits on immobilized oligonucleotides. This assay was used to screen a compound library, yielding several different families of compounds that inhibited capsid assembly. Optimization of two chemical series, termed the benzodiazepines (BD) and the benzimidazoles (BM), resulted in compounds with potent antiviral activity against wild-type and drug-resistant HIV-1. Nuclear magnetic resonance (NMR) spectroscopic and X-ray crystallographic analyses showed that both series of inhibitors bound to the N-terminal domain of CA. These inhibitors induce the formation of a pocket that overlaps with the binding site for the previously reported CAP inhibitors but is expanded significantly by these new, more potent CA inhibitors. Virus release and electron microscopic (EM) studies showed that the BD compounds prevented virion release, whereas the BM compounds inhibited the formation of the mature capsid. Passage of virus in the presence of the inhibitors selected for resistance mutations that mapped to highly conserved residues surrounding the inhibitor binding pocket, but also to the C-terminal domain of CA. The resistance mutations selected by the two series differed, consistent with differences in their interactions within the pocket, and most also impaired virus replicative capacity. Resistance mutations had two modes of action, either directly impacting inhibitor binding affinity or apparently increasing the overall stability of the viral capsid without affecting inhibitor binding. These studies demonstrate that CA is a viable antiviral target and demonstrate that inhibitors that bind within the same site on CA can have distinct binding modes and mechanisms of action.
  • Author:
    El-Far M; Szelei J; Yu O; Fediere G; Bergoin M; Tijssen P
    Title:
    Organization of the ambisense genome of the Helicoverpa armigera densovirus.
    Source:
    J Virol 86 (12), 7024 (2012)
    Abstract:
    A natural densovirus (DNV) of a serious phytophagous pest, Helicoverpa armigera, was isolated. The genome of HaDNV contained 6,039 nucleotides (nt) and included inverted terminal repeats (ITRs) of 545 nt with terminal Y-shaped hairpins of 126 nt. Its DNA sequence and ambisense organization with four typical open reading frames (ORFs) demonstrated that it belonged to the genus Densovirus in the subfamily Densovirinae of the family Parvoviridae
  • Author:
    Wang X-J; Zhang L; Sun X; Lee H; Krishnamuthry D; O'Meara JA; Landry S; Yoakim C; Simoneau B; Yee NK; Senanayake CH
    Title:
    Practical synthesis of a benzophenone-based NNRT inhibitor of HIV-1.
    Source:
    Org Process Res Dev 16 (4), 561-566 (2012)
  • Author:
    Palmer S; Boltz VF; Chow JY; Martinson NA; McIntyre JA; Gray GE; Hopley MJ; Mayers D; Robinson P; Hall DB; Maldrelli F; Coffin JM; Mellors JW
    Title:
    Short-course combivir after single-dose nevirapine reduces but does not eliminate the emergence of nevirapine resistance in women.
    Source:
    Antiviral Ther 17 (2), 327-336 (2012)
  • Author:
    Marra A; Lamb L; Medina I; George D; Gibson G; Hardink J; rugg J; Van Deusen J; O'Donnell J P
    Title:
    Effect of linezolid on the 50% lethal dose and 50% protective dose in treatment of infections by gram-negative pathogens in naive and immunosuppressed mice and on the efficacy of ciprofloxacin in an acute murine model of septicemia
    Source:
    Antimicrob Agents Chemother 56 (9), 4671-4675 (2012)
    Abstract:
    Murine models of infection were used to study the effect of linezolid on the virulence of Gram-negative bacteria and to assess potential pharmacodynamic interactions with ciprofloxacin in the treatment of these infections, prompted by observations from a recent clinical trial. Naive and immunosuppressed mice were challenged with Klebsiella pneumoniae 53A1109, K. pneumoniae GC6658, and Pseudomonas aeruginosa UC12120 in acute sepsis and pulmonary infection models, using different serial dilutions of these pathogens (groups of 8 animals each). Linezolid (100 mg/kg/dose) was administered orally at 0.5 and 4.0 h postchallenge in the sepsis model and at 4 h postchallenge followed by 2 days of twice-daily treatment in the pulmonary model. Further, ciprofloxacin alone and in combination with oral linezolid was investigated in the sepsis model. Survival was assessed for 4 and 10 days postchallenge in the systemic and respiratory models, respectively. The data were fitted to a nonlinear regression analysis to determine 50% lethal doses (LD 50s) and 50% protective doses (PD 50s). A clinically relevant, high-dose regimen of linezolid had no significant effect on LD 50 in these models. This lack of effect was independent of immune status. A combination of oral ciprofloxacin with linezolid yielded lower PD 50s than oral ciprofloxacin alone (ciprofloxacin in combination, 8.4 to 32.7 mg/kg; oral ciprofloxacin, 39.4 to 88.3 mg/kg). Linezolid did not improve the efficacy of subcutaneous ciprofloxacin (ciprofloxacin in combination, 2.0 to 2.4 mg/kg; subcutaneous ciprofloxacin, 2.0 to 2.8 mg/kg). In conclusion, linezolid does not seem to potentiate infections caused by Gram-negative pathogens or to interact antagonistically with ciprofloxacin. Copyright © 2012, American Society for Microbiology. All Rights Reserved.
  • Author:
    Sun J-H; O'Boyle II DR; Zhang Y; Wang C; Nower P; Valera L; Roberts S; Nettles R E; Fridell R A; Gao M
    Title:
    Impact of a baseline polymorphism on the emergence of resistance to the hepatitis C virus nonstructural protein 5a replication complex inhibitor, BMS-790052.
    Source:
    Hepatology 55 (6), 1692-1699 (2012)
    Abstract:
    The influence of naturally occurring polymorphisms on the potency of the HCV nonstructural protein 5A (NS5A) replication complex inhibitor, BMS-790052, was investigated by evaluating hybrid replicons in which the entire NS5A coding region of genotype (GT) la and 1b laboratory (lab) strains (H77c and Con1) were replaced with the corresponding regions of specimens collected from 10 GT-1a- and 6 GT-1b-infected subjects. For baseline (BL) specimens, with no previously observed resistance variants identified by population sequencing, the median 50% effective concentration (EC 50) values for BMS-790052 were similar for the clinically derived and lab strains. A Q30R variant was observed at viral breakthrough (VBT) in one of the GT-1a-infected subjects. Because the lowest plasma exposure of BMS-790052 observed in this subject was 117 nM and the median 50% effective concentration value for a GT-1a H77c replicon containing a Q30R substitution is ?7 nM, a rigorous investigation was initiated to determine the basis for resistance. Three approaches were used: (1) replacement of the entire H77c NS5A or (2) replacement of the N-terminal region of NS5A, with sequence from BL and day 14, and (3) substitution of specific amino acids. A BL polymorphism (E62D) did not contribute resistance to BMS-790052; however, the linked variant, Q30R-E62D, conferred high-level resistance in vitro and is likely responsible for VBT in vivo. Conclusion: Our data show that a BL polymorphism with minimal effect on the anti-HCV effect of BMS-790052 can affect the emergence of resistance and significantly affect clinical outcome. This work establishes a clear, systematic approach to monitor resistance to NS5A inhibitors in the clinic. © 2012 American Association for the Study of Liver Diseases.
  • Author:
    Han Y-S; Quashie P; Mesplede T; Xu H; Mekhssian K; Fenwick C; Wainberg MA
    Title:
    A high-throughput assay for HIV-1 integrase 3'-processing activity using time-resolved fluorescence.
    Source:
    J Virol Methods 184 (1-2), 34-40 (2012)
    Abstract:
    HIV-1 integrase (HIV-1 IN), a well-validated antiviral drug target, catalyzes multistep reactions to incorporate viral DNA into the genome of the host cell; these include both a 3'-processing (3'P) reaction and a strand transfer reaction. These enzymatic activities can be measured in vitro with short DNA oligonucleotides that mimic a single viral LTR DNA end and purified IN. A highly sensitive and reproducible time-resolved fluorescence (TRF)-based assay for HIV-1 IN 3'P activity is now reported. This assay was optimized with respect to time and concentrations of metal ions, substrate and enzyme. The assay has now been used successfully to measure HIV-1 IN 3'P activity and has been shown to detect the anti-IN activity of several known 3'P inhibition compounds accurately. This assay, which is amenable to high-throughput screening, will be useful for identification of additional HIV-1 IN 3'P inhibitors. © 2012 Elsevier B.V.
  • Author:
    Ferris NP; Clavijo A; Yang M; Velazquez-Salinas L; Nordengrahn A; Hutchings GH; Kristersson T; Merza M
    Title:
    Development and laboratory evaluation of two lateral flow devices for the detection of vesicular stromatitis virus in clinical samples.
    Source:
    J Virol Methods 180 (1-2), 96-100 (2012)
  • Author:
    Arasteh K; Ward D; Plettenberg A; Livrozet JM; Orkin C; Cordes C; Guo J; Wang E; Yong CL; Robinson P; Quinson A
    Title:
    Twenty-four-week efficacy and safety of switching virologically suppressed HIV-1-infected patients from nevirapine immediate release 200mg twice daily to nevirapine extended release 400mg once daily (TRANxITION).
    Source:
    HIV Med 13 (4), 236-244 (2012)
  • Author:
    Larrey D; Lohse AW; de Ledinghen V; Trepo C; Gerlach T; Zarski JP; Tian A; Mathurin P; Thimme R; Arasteh K; Trautwein C; Cerny A; Dikopoulos N; Schuchmann M; Heim MH; Gerken G; Stern JO; Wu K; Abdallah N; Grilich B; Scherer J; Berger F; Marquis M; Kukkolj G; Boecher W; Steffgen J
    Title:
    Rapid and strong antiviral of the non-nucleosidic NSSB polymerase inhibitor BI207127 in combination with peginterfon alfa 2a and ribavirin.
    Source:
    J Hepatol Article in Press (2012)
  • Author:
    Ibrisimovic M; Nagl U; Kneidinger D; Rauch M; Lion T; Klein R
    Title:
    Targeted expression of herpes simplex virus thymidine kinase in adenovirus-infected cells reduces virus titers upon treatment with ganciclovir in vitro.
    Source:
    J Gene Med 14 (1), 3-19 (2012)
    Abstract:
    Background: Adenoviruses are a frequent cause of life-threatening infections in immunocompromised patients. Available therapeutics still cannot completely prevent fatal outcomes. By contrast, herpes viruses are well treatable with prodrugs such as ganciclovir (GCV), which are selectively activated in virus-infected cells by virus-encoded thymidine kinases. This effective group of prodrugs is not applicable to adenoviruses and other DNA viruses because they lack those kinases. Methods: To render adenoviruses amenable to GCV treatment, we generated an adenoviral vector-based delivery system for targeted expression of herpes simplex virus thymidine kinase (HSV-TK) in wild-type adenovirus 5 (wt Ad5)-infected cells. HSV-TK expression was largely restricted to wt virus-infected cells by transcription of the gene from the Ad5 E4 promoter. Its activity is dependent on the adenoviral E1A gene product which is not produced by the vector but is only provided in cells infected with wt adenovirus. The anti-adenoviral effect of HSV-TK expression and concomitant treatment with GCV was assessed in vitro in four different cell lines or primary cells. Results: E4 promoter-mediated HSV-TK background expression was sufficiently low to prevent cytotoxicity in the presence of low-levels GCV in cells not infected with wt Ad5. However, expression was several-fold increased in wt Ad5-infected cells and treatment with low levels of GCV efficiently inhibited wt Ad5 DNA replication. Genome copy numbers and output of infectious particles were reduced by up to>99.99% and cell viability was greatly increased. Conclusions: We extended the concept of enzyme/prodrug therapy to adenovirus infections by selectively sensitizing adenovirus-infected cells to treatment with GCV.
  • Author:
    Sulkowski MS; Ceasu E; Asselah T; Caruntu FA; Streinu-Cercel A; Preotescu L; Lalezari J; Ferenci P; Fainboim H; Tanno H; Leggett B; Bessone F; Mauss S; Stern JO; Haefner C; Datsenko Y; Nehmiz G; Boecher W; Steinmann G
    Title:
    SILEN-C1: Sustained virologic response (SVR) and safety of BI201335 combined with peginterforon ALFA-2A and ribavirin (P/R) in treatment naive patients with chronic genotype 1 HCV infection.
    Source:
    J Hepatol 54 (Supp.[1]), s27 MA 60 (2011)
  • Author:
    Kukolj G; Anderson PC; Boes M; Cordingley MG; Coloumbe R; Garneau M; Gillard J; Goulet S; Jolicoeur E; Lagace L; Lamarre D; Laplante S; Marquis M; McKercher G; Pellerin C; Poupart M-A; Beaulieu PL
    Title:
    Preclinical characterization of the hepatitis C virus NS5B polymerase non-nucleoside inhibitor BILB 1941.
    Source:
    J Hepatol 54 (Suppl.1), S480 (2011)
  • Author:
    Cattoli G; Fusaro A; Monne I; Coven F; Joannis T; El-Hamid HSA; Hussein AA; Cornelius C; Amarin NM; Mancin M; Holmes EC; Capua I
    Title:
    Evidence for differing evolutionary dynamics of A/H5N1 viruses among countries applying or not applying avian influenza vaccination in poultry.
    Source:
    Vaccine 29 (50), 9368-9375 (2011)
  • Author:
    Duan J; Garneau M; Amad Ma'An; Bolger G; De Marte J; Montpetit H; Otis F; Jutras M; Rheaume M; White PW; Bethell RC; Yong Ch-L; Llinas-Brunet M; Cordingley MG
    Title:
    Cross-species absorption, metabolism, distribution and pharmacokinetics of BI 201335, a potent HCV genotype 1 NS3/4A protease inhibtor.
    Source:
    Xenobiotica 42 (2), 164-172 (2012)
  • Author:
    Lavilla-Alonso S; Bauer MMT; Abo-Ramadan U; Ristimaeki A; Halavaara J; Desmond RA; Wang D; Escutenaire S; Ahtiainen L; Saksela K; Tatlisumak T; Hemminki A; Pesonen S
    Title:
    Macrophage metalloelastase (MME) as adjuvant for intra-tumoral injection of oncolytic adenovirus and its influence on metastases development.
    Source:
    Cancer Gene Ther 19 (2), 126-134 (2012)
    Abstract:
    Oncolytic adenoviruses are a promising treatment alternative for many advanced cancers, including colorectal cancer. However, clinical trials have demonstrated that single-agent therapy in advanced tumor masses is rarely curative. Poor spreading of the virus through tumor tissue is one of the major issues limiting efficacy. As oncolytic viruses kill preferentially cancer cells, high extracellular matrix (ECM) content constitutes potential barriers for viral penetration within tumors. In this study, the ECM-degrading proteases relaxin, hyaluronidase, elastase and macrophage metalloelastase (MME) were tested for their antitumor efficacy alone and in combination with oncolytic adenovirus. MME improved the overall antitumor efficacy of oncolytic adenovirus in subcutaneous HCT116 xenografts. In a liver metastatic colorectal cancer model, intra-tumoral treatment of primary tumors from HT29 cells with MME monotherapy or with oncolytic adenovirus inhibited tumor growth. Combination therapy showed no increased mortality in comparison with either monotherapy alone. Contradictory results of effects of MME on tumorigenesis and metastasis formation have been reported in the literature. This study demonstrates for the first time in a metastatic animal model that MME, as a monotherapy or in combination with oncolytic virus, does not increase tumor invasiveness. Co-administration of MME and oncolytic adenovirus may be a suitable approach for further optimization aiming at clinical applications for metastatic colorectal cancer.
  • Author:
    Zeuzem St; Soriano V; Asselah T; Bronowicki J-P; Ceausu E; Lohse AW; Streinu-Cercel A; Preotescu L; Moussalli J; Mullhaupt B; Schuchmann M; Bourliere M; Calinas F; Buti M; Roberts StK; Gane EJ; Stern JO; Mensa FJ; Nehmiz G; Bonaventura H; Boecher WO
    Title:
    Virologic response to an interferon-free regimen of BI201335 and BI207127, with and without ribavirin, in treatment-naive patients with chronic genotype-1 HCV infection: week 12 interim results of the sound-C2 study.
    Source:
    Hepatology 54 (Supp.[1]), 1436 A (2011)
  • Author:
    Csank T; Pistl J; Pollakova J; Holoda E; Harvan M
    Title:
    Prevalence of porcine circovirus 2 infection in pig population in Slovakia.
    Source:
    Acta Virol 55 (3), 267-271 (2011)
    Abstract:
    The prevalence of porcine circovirus 2 (PCV-2) infection in the pig population in Slovakia was investigated. Sera from pigs suspected for post-weaning multisystemic wasting syndrome (PMWS) as well as clinically healthy pigs were tested for viral DNA and specific IgM and IgG antibodies. Pigs (n = 198) were categorized to weaning, grower and fattening ones and sows. The results showed that PCV-2 antibodies were present in 53.4% of PMWS-suspects, in 50.0% of healthy pigs and in 69.0% of sows. In PMWS-suspect grower pigs, 40.7% were positive for IgM+IgG antibodies and 22.2% for viral DNA. In PMWS-suspect fattening pigs, 50.0% were positive for IgM+IgG antibodies and 25.0% for viral DNA. In healthy fattening pigs, almost 90.0% were positive for IgG antibodies and 38.5% for viral DNA. The highest proportion of PMWS-suspects was in grower pigs and specific antibodies were increasing with the age of pigs. A combination of positivities for IgG+IgM antibodies and viral DNA was a highly significant marker of PMWS. Viral DNA was detected in seropositive as well as seronegative PMWS-suspects. Overall, in all categories of pigs tested, specific antibodies and viral DNA were detected in 54.0% and 35.5%, respectively.
  • Author:
    Zeuzem St; Asselah T; Angus PW; Zarski J-PH; Larrey DG; Mullhaupt B; Gane EJ; Schuchmann M; Lohse AW; Pol St; Moussalli J; Bronowicki J-P; Roberts SK; Arasteg K; Zoulim F; Stern JO; Mensa FJ; Nehmiz G; Haefner C; Boecher WO
    Title:
    High sustained virologic response following interferon-free treatment of chronic HCV GT1 infection for 4 weeks with HCV protease inhibitor BI201335, polymerase inhibitor BI207127 and ribavirin, followed by BI201335 and pegifn/ribavirin - the sound-C1 study.
    Source:
    Hepatology 54 (Suppl.[1), 486A-487A (2011)
  • Author:
    Kukolj G; Legace L; Cartier M; Marquis M; Triki I; Sulkowski MS; Asselah T; Lalezari JP; Ferenci P; Bethell R; Scherer J; Stern JO; Datsenko Y; Nehmiz G; Boecher WO; Steinmann G
    Title:
    Characterization of HCV NS3 variants that emerged during virologic breaktrough and relapse from BI201335 phase 2 SILEN-C1 study.
    Source:
    62nd Annual Meeting of the American-Association for the Study of Liver Disease (AASLD), San Francisco (United States), Nov 4-8, 2011. Hepatology 54 ( Suppl.1), 991A (2011)
  • Author:
    Lehr Th; Yuan J; Hall D; Zimdahl-Gelling H; Schaerfer HG; Staab A; Macgregor ThR; Jayadev S
    Title:
    Integration of absorption, distribution, metabolism and elimination genotyping data into a population pharmacokinetic analysis of nevirapine.
    Source:
    Pharmacogenet Genomics 21 (11), 721-730 (2011)
    Abstract:
    OBJECTIVES: The aim of this analysis was to show the applicability of a newly developed algorithm to assess the influence of genetic variants and other covariates on nevirapineg€™s drug disposition. The algorithm combines high-throughput genotyping data and nonlinear mixed effects modeling methods. METHODS: Patients, who participated in the 2NN pharmacokinetic sub study, were reconsented and reenrolled into a clinical trial for genotyping analysis. Overall, 198 single nucleotide polymorphisms located in 45 absorption, distribution, metabolism, and elimination related genes were genotyped using the Illumina BeadArray technology. Data analysis was performed using NONMEM VI and SAS 9.1.3. RESULTS: Overall, 1260 nevirapine plasma concentrations were obtained from 271 genotyped patients. Plasma concentrationg^'time profiles of nevirapine were best described by a one-compartment model with auto-induced first-order elimination process. Nevirapine clearance was 19.4% reduced in Asian/Black patients, compared with Caucasian/Hispanic patients. For single nucleotide polymorphism rs3745274 (CYP2B6 516G>T) heterozygous patients (GT) showed a 15.3% reduced clearance; patients with homozygous CYP2B6 516TT alleles showed a 30.6% reduced clearance compared to patients with homozygous 516GG alleles. Patients carrying the homozygote genotype of rs12768009 (CYP2C19 8403AA), highly linked to rs4244285 (CYP2C19*2), showed a 26.8% reduced clearance compared with patients with CYP2C19 8403 AG and GG alleles. CONCLUSION: By integration of high-throughput genotyping data into a pharmacometric analysis of nevirapine, the impact of the CYP2B6 516G>T polymorphism on nevirapineg€™s exposure was confirmed and quantified. In addition, a new hypothesis with regard to CYP2C19 involvement in nevirapine metabolism has been generated. The analysis presented might help to optimize and individualize the therapy for patients treated with nevirapine to add to their therapeutic benefit.
  • Author:
    Cattoli G; Fusaro A; Monne I; Coven F; Joannis T; El-Hamid HSA; Hussein AA; Cornelius C; Amarin NM; Mancin M; Holmes EC; Capua I
    Title:
    Evidence for differing evolutionary dynamics of A/H5N1 viruses among countries applying or not applying avian influenza vaccination in poultry.
    Source:
    Vaccine 29 (50), 9368-9375 (2011)
    Abstract:
    Highly pathogenic avian influenza (HPAI) H5N1 (clade 2.2) was introduced into Egypt in early 2006. Despite the control measures taken, including mass vaccination of poultry, the virus rapidly spread among commercial and backyard flocks. Since the initial outbreaks, the virus in Egypt has evolved into a third order clade (clade 2.2.1) and diverged into antigenically and genetically distinct subclades. To better understand the dynamics of HPAI H5N1 evolution in countries that differ in vaccination policy, we undertook an in-depth analysis of those virus strains circulating in Egypt between 2006 and 2010, and compared countries where vaccination was adopted (Egypt and Indonesia) to those where it was not (Nigeria, Turkey and Thailand). This study incorporated 751 sequences (Egypt n=309, Indonesia n=149, Nigeria n=106, Turkey n=87, Thailand n=100) of the complete haemagglutinin (HA) open reading frame, the major antigenic determinant of influenza A virus. Our analysis revealed that two main Egyptian subclades (termed A and B) have co-circulated in domestic poultry since late 2007 and exhibit different profiles of positively selected codons and rates of nucleotide substitution. The mean evolutionary rate of subclade A H5N1 viruses was 4.07×10-3 nucleotide substitutions per site, per year (HPD 95%, 3.23-4.91), whereas subclade B possessed a markedly higher substitution rate (8.87×10-3; 95% HPD 7.0-10.72×10-3) and a stronger signature of positive selection. Although the direct association between H5N1 vaccination and virus evolution is difficult to establish, we found evidence for a difference in the evolutionary dynamics of H5N1 viruses among countries where vaccination was or was not adopted. In particular, both evolutionary rates and the number of positively selected sites were higher in virus populations circulating in countries applying avian influenza vaccination for H5N1, compared to viruses circulating in countries which had never used vaccination. We therefore urge a greater consideration of the potential consequences of inadequate vaccination on viral evolution.
  • Author:
    Schibler M; Gerlach D; Martinez Y; van Belle S; Kaiser L; Turin L; Tapparel C
    Title:
    Experimental human rhinovirus and enterovirus interspecies recombination.
    Source:
    J Gen Virol 93 (1), 93-101 (2012)
    Abstract:
    Human rhinoviruses (HRVs) and enteroviruses (HEVs), two important human pathogens, are nonenveloped, positive-sense RNA viruses of the genus Enterovirus within the family Picornaviridae. Intraspecies recombination is known as a driving force for enterovirus and, to a lesser extent, rhinovirus evolution. Interspecies recombination is much less frequent among circulating strains, and supporting evidence for such recombination is limited to ancestral events, as shown by recent phylogenetic analyses reporting ancient HRV-A/HRV-C, HEV-A/HEV-C and HEV-A/HEV-D recombination mainly at the 5'-untranslated region (5' UTR)-polyprotein junction. In this study, chimeric genomes were artificially generated using the 5' UTR from two different clinical HRV-C strains (HRV-Ca and HRV-Cc), an HRV-B strain (HRV-B37) and an HEV-A strain (HEV-A71), and the remaining part of the genome from an HRV-A strain (HRV-A16). Whilst the chimeric viruses were easily propagated in cell culture, the wild-type HRV-A16 retained a replication advantage, both individually and in competition experiments. Assessment of protein synthesis ability did not show a correlation between translation and replication efficiencies. These results reflect the interchangeability of the 5' UTR, including its functional RNA structural elements implicated in both genome translation and replication among different enterovirus species. The 5' UTR-polyprotein junction therefore represents a theoretic interspecies recombination breakpoint. This recombination potential is probably restricted by the need for co-infection opportunities and the requirement for the progeny chimera to outcompete the parental genomes' fitness, explaining the rare occurrence of such events in vivo.
  • Author:
    Elgadi MM; Piliero PJ
    Title:
    Boosted tipranavir versus darunavir in treatment-experienced patients: Observational data from the randomized potent trial.
    Source:
    Drugs Res Dev 11 (4), 295-302 (2011)
    Abstract:
    tipranavir/ritonavir (TPV/r) to darunavir/ritonavir (DRV/r), each with an optimized background regimen (OBR) in triple-class experienced HIV-1-infected patients with resistance to more than one protease inhibitor (PI). MethodologyPrincipal Findings: POTENT was a prospective, open-label study of triple-class (PI, non-nucleoside reverse transcriptase inhibitors [NNRTI], nucleoside reverse transcriptase inhibitors [NRTI]), treatment-experienced, HIV-positive patients. Subjects were randomized to either TPV/r (500/200 mg twice daily) or DRV/r (600/100 mg twice daily) on a genotype-guided, investigator-selected OBR. CD4+ counts and HIV viral loads were assayed at key timepoints. The primary endpoint was time to virologic failure (viral load .gtoreq.500copies/mL). POTENT was prematurely terminated due to slow enrollment. Thirty-nine patients were treated with either TPV/r (n = 19) or DRV/r (n = 20); 82% were male, 77% White, with mean age of 43.6 years. Mean baseline HIV RNA was 3.9 log(10) copies/mL. Median prior antiretrovirals was 11, with no prior raltegravir or maraviroc exposure. Raltegravir was the most common novel class agent in the OBRs (n = 14 TPV/r; n = 12 DRV/r). In both groups, patients achieved mean viral load decreases .gtoreq.2 log(10) copies/mL by week 8, and by week 12 mean CD4+ counts rose by 40-50 cells/mm3. Total observation time was 32 weeks. Drug-related adverse events were reported in 21% (TPV/r) and 25% (DRV/r) of patients. ConclusionsSignificance: TPV/r- and DRV/r-based regimens showed similar short-term safety and efficacy. These data support the use of next-generation PIs such as tipranavir or darunavir with novel class antiretroviral agents (integrase inhibitors, CCR5 antagonists, or fusion inhibitors). Trial Registration: Clinicaltrials.gov NCT00517192. .COPYRGT. 2011 Elgadi & Piliero, publisher and licensee Adis Data Information BV. This is an open access article published under the terms of the Creative Commons License "Attribution-NonCommercial-NoDerivative 3.0" (http://creativecommons.org/licenses/by-nc-nd/3.0/) which permits non-commercial use, distribution, and reproduction, provided the original work is properly cited and not altered.
  • Author:
    Murtaugh MP; Genzow M
    Title:
    Immunological solutions for treatment and prevention of porcine reproductive and respiratory syndrome (PRRS).
    Source:
    Vaccine 29 (46), 8192-8204 (2011)
    Abstract:
    Vaccination is the principal means used to control and treat porcine reproductive and respiratory syndrome virus (PRRSV) infection. An array of PRRS vaccine products is available in various regions of the world. However, despite extensive efforts, little progress has been made to improve efficacy since the first introduction of a live, attenuated vaccine in 1994 in the USA. Key limitations include: (a) uncertainty about the viral targets of protective immunity that prevents a research focus on individual viral structures and proteins, and frustrates efforts to design novel vaccines; (b) inability to establish clear immunological correlates of protection that requires laborious in vivo challenge models for evaluation of protection against challenge; and (c) the great genetic diversity of PRRSV which requires that challenge experiments be interpreted cautiously since it is not possible to predict how immunological protection against one isolate will translate to broadly cross-protective immunity. Economically significant levels of cross-protection that are provided to a variety of field isolates still cannot assure that effective protection will be conferred to isolates that might emerge in the future. In addition to these substantial barriers to new PRRSV vaccine development, there are enormous gaps in our understanding of porcine immunological mechanisms and processes that provide immunity to PRRSV infection and memory responses for long-term protection. Despite these impediments, we should be confident that progress will be made. Sequencing of the swine genome is providing a rich source of primary knowledge of gene structure and transcriptional regulation that is certain to reveal important insights about the mechanisms of anti-PRRSV immunity, and continued efforts to unravel the details of the interaction of PRRSV with pigs will lead to new insights that overcome the current limitations in the field.
  • Author:
    Battegay M; Arasteh K; Plettenberg A; Bogner JR; Livrozet JM; Witt MD; Mossdorf E; Yong C-L; Zhang W; Macha S; Berger F; Stern J; Robinson P; Quinson A-M
    Title:
    Bioavailability of extended-release nevirapine 400 and 300 mg in HIV-1: A multicenter, open-label study.
    Source:
    Clin Ther 33 (9), 1308-1320 (2011)
    Abstract:
    Background: Nevirapine (NVP) is a widely used non-nucleoside reverse transcriptase inhibitor. A once-daily extended-release (XR) formulation would potentially increase adherence and thus efficacy. Objective: The aim of this study was to investigate the steady-state bioavailability of 2 once-daily tablet formulations of NVP XR (containing 25% or 20% hypromellose; NVP XR25 and NVP XR20, respectively) in 400- or 300-mg tablets compared with twice-daily immediate-release (IR) NVP 200-mg tablets. Methods: This Phase Ib multinational, multicenter, open-label trial was conducted in patients aged 18 to 60 years, infected with HIV-1 (viral load, ?50 copies/mL), and treated for ?12 weeks with twice-daily NVP IR 200 mg. Patients were switched to NVP XR25 400 or 300 mg or NVP XR20 400 or 300 mg for 19 days. Plasma samples were collected over 24-hour periods at steady state. Primary end points were AUC0-24,ss, Cmax,ss, and Cmin,ss, analyzed using an ANOVA statistical model on the logarithmic scale and 2-sided 90% CI. Sample size was determined assuming an intrasubject %CV of 20% for Cmax. Adverse events (AEs) and viral loads were monitored. Results: Ninety-two patients were enrolled (NVP XR25 400 mg, 24 patients; NVP XR20 400 mg, 24; NVP XR25 300 mg, 21; NVP XR20 300 mg, 23). Compared with NVP IR, the AUC0-24,ss values of the NVP XR formulations were lower (test/reference ratios: 79.5% [90% CI, 73.0-86.7; P = 0.544], 71.0% [90% CI, 63.3-79.7; P = 0.956], 90.3% [90% CI, 80.4-101.4; P = 0.044], and 83.7% [90% CI, 77.9-89.9; P = 0.148] with NVP XR25 400 mg, NVP XR20 400 mg, NVP XR25 300 mg, and NVP XR20 300 mg, respectively). The relative bioavailability of NVP XR25 was greater compared with that of NVP XR20. Cmax,ss values were lower with all NVP XR formulations compared with NVP IR. For Cmin,ss, NVP XR25 400 and 300 mg were not significantly different from NVP IR, with 90% CIs within the range of 80% to 125% (P = 0.039 and P = 0.017, respectively). All AEs were mild or moderate, with no significant differences between treatment groups. No virologic failures (viral load, >50 copies/mL over 2 consecutive readings) were observed. Conclusions: Extent of bioavailability was lower and tmax,ss was delayed with all NVP XR formulations compared with NVP IR. The bioavailability of the NVP XR25 formulations was greater than that of the NVP XR20 formulations. Cmin,ss with NVP XR25 was similar to that with NVP IR. All of the NVP XR formulations were well tolerated. The 400-mg NVP XR25 formulation was selected for further development.
  • Author:
    Quinson A-M; Scherer J; Nguyen T; Robinson P; Hall DB; Bethell R; Gathe J; Podzcamzcer D
    Title:
    Resistance development in ART-naive patients randomized to receive immediate or extended release formulations of nevirapine (NVP) in combination with tenofovir (TDF) and emtricitabine (FTC): (VERxVE).
    Source:
    International Workshop on HIV and Hepatitis Virus Drug Resistance and Curative Strategies, Los Cabos (Mexico), Jun 7-11, 2011Antiviral Ther 16 ( 4), A73 (2011)
    Abstract:
    This randomized, placebo-controlled VERxVE study (NCT00561925) evaluated risk of early virological failure and development of resistance in 1068 antiretroviral therapy (ART)-naive HIV-1 patients treated with immediate (IE) or extended release (XE) formulations of nevirapine (NVP) in combination with tenofovir (TDF) and emtricitabine (FTC). The regimen of NVP+TDF+FTC was efficacious. The majority of those who failed to achieve sustained response discontinued for reasons other than efficacy and had little emergent resistance. Clade C virological failures differed from other clades with increased prevalence of K103N and decreased prevalence of Y181C and TDF/FTC resistance. Methods 1068 ART-naive HIV-1 patients with >50 to <250 CD4 cells/ml (female), >50 to <400 CD4 cells/ml (male) were enrolled. After 2 wk of NVP 200 mg QD and TDF+FTC, were randomized to NVP IE 200 mg BID or NVP XE 400 mg QD for 48 wk. Treatment was blinded with placebos for both formulations. Results 96/1068 Patients discontinued due to lack of efficacy or virological failure and 17% for reasons other than efficacy. 100/278 Patients were evaluable (59/96 virological failures and 41/182 with discontinuation for other reasons) based on successful genotypic sequencing of on-treatment specimens. 16 Patients were evaluable based on post-wk 48 rebound or transient elevations. Virological failure was characterized by development of resistance, 46/59 had resistance to one or both of NVP or TDF/FTC. Late or transient rebound had resistance for 5/16 of patients. 5% Of treatment failures not due to efficacy developed resistance. The only difference between XE and IE was in the number of evaluable patients, 45 for XE and 71 for IE.
  • Author:
    Fenwick CW; Tremblay S; Wardrop E; Bethell R; Coulomb R; Elston R; Faucher A-M; Mason S; Simoneau B; Tsantrizos Y; Yoakim C
    Title:
    Resistance studies with HIV-1 non-catalytic site integrase inhibitors.
    Source:
    Antiviral Ther 16 (4), A9 (2011)
  • Author:
    Gathe J; Andrade-Villanueva J; Santiago St; Horban A; Nelson M; Cahn P; Bogner J; Spencer D; Podzamczer D; Yong Ch-L; Thuy N; Zhang W; Drulak M; Quinson A-M
    Title:
    Efficacy and safety of nevirapine extended-release once daily versus nevirapine immediate-release twice-daily in treatment-naive HIV-1-infected patients.
    Source:
    Antiviral Ther 16 (5), 759-769 (2011)
    Abstract:
    Background: This study (VERxVE) compared the efficacy and safety of the new nevirapine extended-release (NVP XR) formulation dosed once daily with NVP immediate release (IR) twice daily in treatment-naive patients. Methods: Randomized, double-blind, double-dummy, parallel group study of HIV-1-infected adult patients with baseline viral load (VL) >= 1,000 copies/ml and CD4(+) T-cell count of >50-<400 (males) and >50-<250 cells/mm(3) (females). Patients stratified by baseline VL (<= 100,000/>100,000 copies/ml) were randomized 1:1 to NVP XR 400 mg once daily (plus placebo) or NVP IR 200 mg twice daily (plus placebo), both combined with tenofovir disoproxil fumarate (TDF) 300 mg and emtricitabine 200 mg once daily. Primary endpoint was sustained virological response (<50 copies/ml) through week 48 using the time to loss of virological response algorithm. Non-inferiority of NVP XR to NVP IR was tested using Cochran's statistic incorporating baseline VL stratum with pre-specified, non-inferiority margin of -10%. Results: Among 1,011 patients randomized and treated, virological response at week 48 was 81.0% (409/505) for NVP XR and 75.9% (384/506) for NVP IR with adjusted difference of 4.9% in favour of NVP XR (95% CI -0.1-10.0%), demonstrating non-inferiority of NVP XR to NVP IR. This finding was supported by secondary endpoints. The safety profile of NVP XR was similar to NVP IR, but showed numerically fewer treatment-related adverse events. Conclusions: NVP XR in combination with TDF and emtricitabine was shown to be non-inferior in efficacy to NVP IR with a similar safety and adverse event profile, with the potential for the added convenience of once-daily dosing. - DERWENT Abstract - This randomized, double-blind, double-dummy parallel group study (VERxVE, NCT00561925) compared the efficacy and safety of the new nevirapine extended-release (NVP XR) formulation dosed once daily with NVP immediate release (IR) b.i.d. in treatment-naive patients. Among 1011 patients randomized and treated, virological response at wk 48 was 81.0% for NVP XR and 75.9% for NVP IR with adjusted difference of 4.9% in favor of NVP XR. In conclusion, NVP XR in combination with tenofovir disoproxil fumarate (TDF) and emtricitabine was shown to be non-inferior in efficacy to NVP IR with a similar safety and adverse event profile, with the potential for the added convenience of once-daily dosing. Methods 1011 Treatment-naive HIV-1-infected patients were randomized to NVP XR 400 mg once daily (plus placebo) (n=505) or NVP IR 200 mg b.i.d. (plus placebo) (n=506), both combined with TDF 300 mg and emtricitabine 200 mg once daily. Primary endpoint was sustained virological response (less than 50 copies/ml) through wk 48 using the time to loss of virological response algorithm. Results The mean exposure time to study medication was 61.5 wk for NVP XR and 60.3 wk for NVP IR, including the lead-in period. Based on the TLOVR algorithm and Amplicorcorrected assay, the overall sustained virological response proportion at wk 48 was 81.0% (409/505) for NVP XR and 75.9% (384/506) for NVP IR, with an adjusted difference of 4.9% in favor of NVP XR. A total of 96 (19.0%) NVP XR and 122 (24.1%) NVP JR patients were classed as treatment failures. Resistance to the drugs in the treatment regimens developed with the same pattern for NVP XR and NVP IR treatments. Overall 36/86 patients (42%) did not have NVP-resistant virus at virological failure. In total, there were 55 discontinuations during this period, of which 38 (69.1%) were due to an adverse events, mostly rash.
  • Author:
    Fusaro A; Monne I; Salviato A; Valastro V; Schivo A; Capua I; Cattoli G; Amarin NM; Gonzalez C; Ismail MM; Al-Ankari A-R; Al-Blowi M-H; Khan OA; Ali ASM; Hedayati A; Garcia Garcia J; Ziay GM; Shoushtari A; Al Qahtani KN; Holmes EC
    Title:
    Phylogeography and evolutionary history of reassortant H9N2 viruses with potential human health implications.
    Source:
    J Virol 85 (16), 8413-8421 (2011)
    Abstract:
    Avian influenza viruses of the H9N2 subtype have seriously affected the poultry industry of the Far and Middle East since the mid-1990s and are considered one of the most likely candidates to cause a new influenza pandemic in humans. To understand the genesis and epidemiology of these viruses, we investigated the spatial and evolutionary dynamics of complete genome sequences of H9N2 viruses circulating in nine Middle Eastern and Central Asian countries from 1998 to 2010. We identified four distinct and cocirculating groups (A, B, C, and D), each of which has undergone widespread inter- and intrasubtype reassortments, leading to the generation of viruses with unknown biological properties. Our analysis also suggested that eastern Asia served as the major source for H9N2 gene segments in the Middle East and Central Asia and that in this geographic region within-country evolution played a more important role in shaping viral genetic diversity than migration between countries. The genetic variability identified among the H9N2 viruses was associated with specific amino acid substitutions that are believed to result in increased transmissibility in mammals, as well as resistance to antiviral drugs. Our study highlights the need to constantly monitor the evolution of H9N2 viruses in poultry to better understand the potential risk to human health posed by these viruses.
  • Author:
    Ertl R; Birzele F; Hildebrandt T; Klein D
    Title:
    Viral transcriptome analysis of feline immunodeficiency virus infected cells using second generation sequencing technology.
    Source:
    Vet Immunol Immunopathol 143 (3-4), 314-324 (2011)
    Abstract:
    Feline immunodeficiency virus (FIV) is a widespread pathogen causing immunodeficiency in domestic cats and related wild cat species. The virus genome includes the main structural genes common to all retroviruses as well as accessory genes displaying essential functions during the viral life cycle. Expression of viral genes involves transcription of provirus genomes into full-length transcripts, which are partially processed into several spliced mRNA variants for the translation of particular proteins. Among several FIV isolates derived from domestic cats, notable differences in pathogenicity could be observed leading to identification of low and high pathogenic virus isolates. This study investigates the viral transcriptome of two differentially virulent FIV strains using second generation sequencing (SGS) technology. The expression levels of viral genes as detected by SGS were additionally determined by reverse transcription quantitative PCR analysis in order to compare two methods of mRNA quantification. The different properties of both methods, especially regarding normalization between samples, had to be considered when comparing the resulting data. SGS turned out to be a suitable technique for comparing mRNA transcription between both FIV infected cell lines and the identification of spliced viral transcripts. In contrast to this, the quantification of these spliced isoforms using SGS data was impeded by the short length of sequencing reads. In summary, SGS analysis revealed very consistent mRNA levels for the majority of viral genes between the low pathogenic Petaluma and the more highly pathogenic Glasgow 8 isolate. Notable differences among the two FIV strains could be observed in the viral mRNA splicing where Glasgow 8 displays similarities to the transcription pattern seen in the early stages of natural lentivirus infections. Thus, divergences in the regulation of post-transcriptional RNA processing might represent an additional contributor to the diverse pathogenic effects of individual FIV isolates. Taken together, this study aims to investigate the viral transcriptome as one part of the complex network of virus-host interactions, which will contribute to gaining deeper insights into FIV pathogenesis.
  • Author:
    Kort JJ; Aslanyan S; Scherer J; Sabo JP; Kohlbrenner V; Robinson P
    Title:
    Effects of tipranavir, darunavir and ritonavir on platelet function, coagulation and fibrinolysis in healthy volunteers.
    Source:
    Curr HIV Res 9 (4), 237-246 (2011)
    Abstract:
    The use of HIV protease inhibitors (PIs) as part of antiretroviral therapy in the treatment of HIV-1 infection may be associated with an increased risk of bleeding. This prospective, randomized, open-label trial in healthy volunteers compared the effects of tipranavir/ritonavir (TPV/r), darunavir/ritonavir (DRV/r), and ritonavir (RTV) alone on platelet aggregation after a single dose and at steady-state concentrations. Subjects were selected on the basis of normal platelet aggregation and arachidonic acid (AA)-induced platelet aggregation inhibition after administration of a single 325-mg dose of aspirin. All 3 PI therapies were administered twice daily for 10 days. In some but not all subjects, TPV/r inhibited AA-induced platelet aggregation and prolonged PFA-100.RTM. closure time with collagen-epinephrine cartridge, which was of lesser magnitude and consistency compared with aspirin, but greater when compared to DRV/r and RTV. At least 2 subjects in each treatment arm showed complete inhibition of AA-induced platelet aggregation on treatment, and the magnitude of change in all platelet-function tests did not correlate with PI plasma concentrations. Effects of TPV/r on platelet aggregation were reversed 24 hours after the last TPV/r dose. None of the PI treatments tested were associated with increases in bleeding time, decreases in plasma coagulation factors, or increase in fibrinolysis. There was large inter-patient variability in antiplatelet effect for all PI treatments, ranging from no effect to complete inhibition of AA-induced platelet aggregation.
  • Author:
    Soriano V; Arasteh K; Migrone H; Lutz T; Opravil M; Andrade-Villanueva J; Antunes F; di Perri G; Podzamczer D; Taylor S; Domingo P; Gellermann H; de Rossi L
    Title:
    Nevirapine versus atazanavir/ritonavir, each combined with tenofovir disoproxil fumarate/emtricitabine, in antiretroviral-naive HIV-1 patients. The ARTEN trial.
    Source:
    Antiviral Ther 16 (3), 339-348 (2011)
    Abstract:
    Background: Selection of first-line antiretroviral therapy requires consideration of efficacy as well as effects on lipids given the increased concern about cardiovascular risk in HIV-1 patients. Methods: ARTEN is a randomized, open-label, non-inferiority trial that compares nevirapine (NVP) 200 mg twice daily or 400 mg once daily to atazanavir/ritonavir (ATZ/r) 300 mg/100 mg once daily, each combined with fixed-dose tenofovir disoproxil fumarate (TDF) 300 mg/emtricitabine (FTC) 200 mg once daily, in antiretroviralnaive HIV-1 patients with CD4+ T-cell counts <400 (men) and <250 cells/mm3 (women). The primary end point was plasma HIV RNA<50 copies/ml at two consecutive visits prior to week 48. Results: A total of 569 patients were randomized and treated. Overall, 66.8% of NVP and 65.3% of ATZ/r patients achieved the primary end point (difference 1.9%, 95% CI -5.9-9.8%). Similar rates of serious adverse events were observed (9.6% on NVP versus 8.8% on ATZ/r), although discontinuations due to adverse events were more frequent with NVP than ATZ/r (13.6% versus 3.6%, respectively). None of the 28 patients virologically failing ATZ/r selected resistance mutations, while they were selected in 29/44 patients virologically failing NVP. NVP induced a significantly greater increase in high-density lipoprotein cholesterol (HDL-c) and apolipoprotein A1 from baseline than ATZ/r, whereas triglycerides increased significantly more with ATZ/r than NVP. Mean change from baseline in TC:HDL-c ratio was -0.24 for NVP and 0.13 for ATZ/r (P=0.0001). Conclusions: NVP demonstrated at week 48 non-inferior antiviral efficacy compared with ATZ/r when given along with TDF/FTC, despite more drug-related discontinuations with NVP than ATZ/r. NVP was associated with a lower atherogenic lipid profile than ATZ/r although resistance mutations were more frequently selected with NVP than ATZ/r. - DERWENT Abstract - This ARTEN, a randomized, multicenter, open-label, non-inferiority trial compared the effects of nevirapine (NP) with atazanavir (AZ) + ritonavir (RN), each combined with tenofovir disoproxil fumarate (TD) + emtricitabine (EB), in 569 antiretroviral-naive HIV-1 patients. 66.8% Of NP-and 65.3% of AZ + RN-treated patients achieved the primary end point (plasma HIV RNA less than 50 copies/ml at 2 consecutive visits prior to wk 48). NP induced a significantly greater increase in HDL-cholesterol and apolipoprotein A1 from baseline vs. AZ + RN, while triglycerides increased significantly more with AZ + RN vs. NP. Mean change from baseline total cholesterol:HDL-cholesterol ratio was -0.24 for NP and 0.13 for AZ + RN. NP demonstrates at wk 48 non-inferior antiviral efficacy vs. AZ + RN when given along with TD + EB, despite more drug-related discontinuations with NP than AZ/RN. Methods 569 Antiretroviral-naive HIV-1 patients were randomized to receive NP at 200 mg b.i.d. + TD (300 mg/day) + EB (200 mg/day; n = 188, 163 male, mean age 40 yr; arm 1), NP at 400 mg/day + TD + EB (n = 188, 152 male, mean age 38 yr; arm 2), or AZ (300 mg/day) + RN (100 mg/day) + TD + EB (n = 193, 162 male, mean age 48 yr; arm 3). Results 62.8 And 65.8% of patients in arms 1, 2 and 3 had baseline plasma HIV RNA more than 100000 copies/ml. At wk 48, similar proportions of patients in arms 1, 2 and 3 achieved the primary end point (plasma HIV RNA less than 50 copies/ml at 2 consecutive visits prior to wk 48) of treatment response. In arm 1, 21/188 patients discontinued therapy due to lack of efficacy. In arm 2, 11/188 patients discontinued therapy due to lack of efficacy. Among patients with virological failure in arms 1 and 2, the most frequent NNRTI resistance mutation was Y181C/I/V/S, occurring in 9/20 patients in arm 2 and 15/24 patients in arm 1. In arm 3, 3/193 patients discontinued therapy due to lack of efficacy. Mean changes in fasting HDL-cholesterol and apolipoprotein A1 from baseline to wk 48 were higher with arms 1 and 2 vs. arm 3. Although a greater increase in LDL-cholesterol and total cholesterol was noted in arms 1 and 2 vs. arm 3, the mean total cholesterol:HDL-cholesterol ratio was reduced with arms 1 and 2, but increased with arm 3. Common adverse events were infection, rash, increased ALT, AST and bilirubin levels, hyperbilirubinemia, decreased GFR, renal insufficiency, increased blood creatine phosphokinase level, jaundice, pneumonia, eruptions, exanthema, pyrexia, and hypertriglyceridemia.
  • Author:
    Heidari Sh; Karim QA; Banegura A; Cahn P; Christie CDC; Dintruff R; Distel M; Hankins C; Hellmann N; Katabira E; Lehrman S; Montaner J; Rooney JF; Wood R; Purdon S
    Title:
    Asking the right questions: developing evidence-based strategies for treating HIV in women and children.
    Source:
    BMC Public Health 11 (2011)
    Abstract:
    In July 2010, the World Health Organization (WHO) issued formal revisions of its guidelines on the use of highly active antiretroviral therapy for HIV. The new guidelines greatly expand eligibility for treatment of adults and children, as well as for pregnant women seeking prophylaxis for vertical HIV transmission. WHO's new recommendations bring the guidelines closer to practices in developed countries, and its shift to earlier treatment alone will increase the number of treatment-eligible people by 50% or more. Scaling up access to HIV treatment is revealing important gaps in our understanding of how best to provide for all those in need. This knowledge gap is especially significant in developing countries, where women and children comprise a majority of those living with HIV infection. Given the magnitude and significance of these populations, the International AIDS Society, through its Industry Liaison Forum, prioritized HIV treatment and prophylaxis of women and children. In March 2010, the International AIDS Society and 15 partners launched a Consensus Statement outlining priority areas in which a relative lack of knowledge impedes delivery of optimal prevention of mother to child transmission (PMTCT) and treatment to women and children. The Consensus Statement, "Asking the Right Questions: Advancing an HIV Research Agenda for Women and Children", makes a special appeal for a more gender-sensitive approach to HIV research at all stages, from conception to design and implementation. It particularly emphasizes research to enhance the understanding of sex-based differences and paediatric needs in treatment uptake and response. In addition to clinical issues, the statement focuses on programmatic research that facilitates access and adherence to antiretroviral regimens. Better coordination of HIV management with sexual and reproductive healthcare delivery is one such approach. We discuss here our knowledge gaps concerning effective, safe PMTCT and treatment for women and children in light of the expansion envisioned by WHO's revised guidelines. The guideline's new goals present an opportunity for advancing the women and children's agenda outlined in the Consensus Statement.
  • Author:
    Seclen E; Soriano V; Gonzalez MM; Martin-Carbonero L; Gellermann H; Distel M; Kadus W; Poveda E
    Title:
    Impact of baseline HIV-1 tropism on viral response and CD4 cell count gains in HIV-infected patients receiving first-line antiretroviral therapy.
    Source:
    J Infect Dis 204 (1), 138-144 (2011)
    Abstract:
    Background. Viral tropism influences the natural history of human immunodeficiency type 1 (HIV-1) disease: X4 viruses are associated with faster decreases in CD4 cell count. There is scarce information about the influence of viral tropism on treatment outcomes. Methods. Baseline plasma samples from patients recruited to the ArTEN (Atazanavir/ritnoavir vs. Nevirapine on a background of Tenofovir and Emtricitabine) trial were retrospectively tested for HIV-1 tropism using the genotypic tool geno2phenoFPR55.75%. ArTEN compared nevirapine with atazanavir-ritonavir, both along with tenofovir-emtricitabine, in drug-naïve patients. Results. Of 569 ArTEN patients, 428 completed 48 weeks of therapy; 282 of these received nevirapine and 146 of these received atazanavir-ritonavir. Overall, non-B subtypes of HIV-1 were recognized in 96 patients (22%) and X4 viruses were detected in 55 patients (14%). At baseline, patients with X4 viruses had higher plasma HIV RNA levels (5.4 vs 5.2 log copies/mL, respectively; P 5 .044) and lower CD4 cell counts (145 vs 188 cells/?L, respectively; P &lt; .001) than those with R5 strains. At week 48, virologic responses were lower in patients with X4 viruses than in patients with R5 viruses (77% vs 92%, respectively; P = .009). Multivariate analysis confirmed HIV-1 tropism as an independent predictor of virologic response at week 24 (P = .012). This association was extended to week 48 (P = .007) in clade B viruses. Conversely, CD4 cell count recovery was not influenced by baseline HIV-1 tropism. Conclusions. HIV-1 tropism is an independent predictor of virologic response to first-line antiretroviral therapy. In contrast, it does not seem to influence CD4 cell count recovery. Clinical Trials Registration. NCT00389207. - DERWENT Abstract - This study (NCT00389207) evaluated the impact of baseline HIV-1 tropism on viral response and CD4 cell count gains in 428 HIV-infected patients receiving 1st-line antiretroviral therapy. Overall, non-B subtypes of HIV-1 were recognized in 96 patients and X4 viruses were detected in 55 patients. At baseline, patients with X4 viruses had higher plasma HIV RNA levels and lower CD4 cell counts than those with R5 strains. At wk 48, virologic responses were lower in patients with X4 viruses than in patients with R5 viruses. Multivariate analysis confirmed HIV-1 tropism as an independent predictor of virologic response at wk 24. Thus, HIV-1 tropism is an independent predictor of virologic response to 1st-line antiretroviral therapy.
  • Author:
    Squires KE; Hodder SL; Feinberg J; Bridge DA; Abrams St; Storfer StP; Aberg JA
    Title:
    Health needs of HIV-infected women in the United States: Insights from the women living positive survey.
    Source:
    AIDS Patient Care STDs 25 (5), 279-285 (2011)
    Abstract:
    The objective of this study was to describe attitudes, opinions, and perceived health needs of HIV-infected women in the United States. In this cross-sectional study, women were invited to participate in the Women Living Positive survey, a structured interview instrument with 45 questions. Collected data were deidentified and the margin of error was calculated as four percentage points. Incoming toll-free phone interviews were conducted from December 21, 2006, through March 14, 2007 among subjects recruited from a U.S. national network of AIDS counseling centers. Seven hundred HIV-infected women (43% African American, 28.5% Hispanic, 28.5% Caucasian; median age, 42.5 years) receiving combination antiretroviral therapy for 3 years or more replied to recruitment flyers. Overall, 55% of survey participants had never discussed gender-specific HIV treatment issues with their HIV care providers. Of the 45% who did discuss these issues, almost all (96%) were satisfied. On average, one-third of the women had seen three or more providers since beginning HIV treatment; 43% indicated they had switched providers because of communication issues. Among women who had been or were pregnant at the time of the survey (n=159), more than half (57%) had not had pre-pregnancy discussions with their HIV provider about the most appropriate HIV regimens for women attempting to become pregnant. Significant communication gaps exist between HIV-infected women and their providers when discussing gender-specific treatment issues. These data highlight a need for U.S. health care providers to incorporate discussion of gender-specific issues, including preconception and reproductive counseling, into management strategies for HIV-infected women.
  • Author:
    Sanford SE; Young MG; Cunningham GL
    Title:
    Circovirus vaccination in pigs with subclinical porcine circovirus type 2 infection complicated by ileits.
    Source:
    J Swine Health Prod 19 (3), 175-180 (2011)
    Abstract:
    This study demonstrated a clear benefit in vaccinating pigs subclinically infected with porcine circovirus type 2 (PCV2) with a one-dose PCV2 vaccine. At weaning (3 weeks of age), 1427 pigs were vaccinated and 1431 were sham-inoculated with a placebo (0.9% physiological saline). After 33 days in the nursery, 528 pigs from each treatment group were moved into 48 pens in a commercial finisher barn, ensuring equal age of pigs across both treatments. Each pig was weighed on day 2 in the finisher barn and on day 88 (end of the study period). The vaccinates had a 36-g-per-day advantage in daily gain and 4.6% less mortality than the unvaccinated pigs. More than twice as many vaccinates as unvaccinated pigs (40 versus 16, respectively) were marketed at the first shipment. Furthermore, carcass weight (95.0 kg versus 94.0 kg; P<.05), lean percent (60.52% versus 60.26%; P<.05) and carcass index (111.6 versus 111.1; P<.05) were all greater for the vaccinates. The mean loin depth was 65.1 mm for vaccinates and 63.3 mm for unvaccinated pigs (P<.05). The vaccinated pigs delivered a return on investment of $5.90 per pig over the unvaccinated pigs.
  • Author:
    Podzamczer D; Andrade-Villanueva J; Clotet B; Taylor S; Rockstroh J; Reiss P; Domingo P; Gellermann H; de Rossi L; Cairns V; Soriano V
    Title:
    Lipid profiles for nevirapine vs. atazanavir/ritonavir, both combined with tenofovir disoproxil fumarate and emtricitabine over 48 weeks, in treatment naive HIV-1-infected patients (the ARTEN study).
    Source:
    HIV Med 12 (6), 374-382 (2011)
    Abstract:
    The Authors evaluated lipid profiles for nevirapine (NVP) vs. atazanavir/ritonavir (ATZ/r), both combined with tenofovir disoproxil fumarate and emtricitabine over 48 wk, in an ongoing multinational, multicenter, randomized, open-label, prospective study of 569 antiretroviral (ARV) treatment-naive HIV-1-infected patients. NVP showed a potentially less atherogenic lipid profile compared with ATZ/r in ARV-naive patients with low cardiovascular risk (CR) at the outset. Methods 569 ARV treatment-naive HIV-1-infected patients received ritonavir-boosted atazanavir (ATZ/r) 300 mg/100 mg once/day (qd) with immediate-release NVP 200 mg b.i.d. or 400 mg/day, each combined with fixed-dose tenofovir 300 mg/emtricitabine 200 mg qd. Lipid profiles and CR from baseline to wk 48 were reported. Changes from baseline to wk 48 in fasting plasma levels of total cholesterol (TC), HDL cholesterol, LDL cholesterol, TC:HDL cholesterol ratio, apolipoprotein A1 (ApoA1), apolipoprotein B (ApoB), and total triglycerides (TG) were determined. Results At wk 48, NVP treatment resulted in significantly greater mean increases from baseline in TC (24.4 mg/dl vs. 19.6 mg/dl), HDL cholesterol (9.7 m/dl vs. 3.9 mg/dl), LDL cholesterol (15.0 mg/dl vs. 10.4 mg/dl) and ApoA1 (0.18 g/l vs. 0.08 g/l) but not ApoB (0.02 g/l vs. 0.02 g/l) compared with ATZ/r treatment. ATZ/r use was associated with higher mean TG increases (27.80 mg/dl vs. 0.02 mg/dl). Significantly greater mean decreases in TC:HDL cholesterol and ApoB/ApoA ratios were observed with NVP vs. ATZ/r. Framingham CR scores were low and comparable between the arms, with only a slight mean increase from baseline to wk 48 of 0.70 for NVP and 0.80 for ATZ/r (difference -0.069).
  • Author:
    Stoltz JH; Stern JO; Huang Q; Seidler RW; Pack FD; Knight BL
    Title:
    A twenty-eight-day mechanistic time course study in the rhesus monkey with hepatitis C virus protease inhibitor BILN 2061.
    Source:
    Toxicol Pathol 39 (3), 496-501 (2011)
    Abstract:
    BILN 2061 is a potent, reversible inhibitor of hepatitis C virus NS3/NS4A serine protease. Early clinical proof of principle with the drug was offset by the results of subsequent safety studies in Rhesus monkeys revealing cardiotoxicity that featured myocardial vacuolation corresponding to mitochondrial swelling. Here we describe an investigation into the nature, onset, and reversibility of the lesion, and an assessment of potentially predictive biomarkers for the change. Rhesus monkeys were orally administered 1,000 mg/kg/day BILN 2061 and either necropsied after one, three, fourteen, or twenty-eight doses or afforded a ten-week recovery period. The results of electrocardiographic and plasma troponin I and T measurements were unaffected by BILN 2061, but cardiac myocytic vacuolation, correlated with mitochondrial swelling, was observed after three or more doses. Echocardiographic traces obtained after twenty-eight consecutive days of dosing revealed two animals with diminished left ventricular cardiac ejection fraction. One animal was immediately necropsied and exhibited marked cardiotoxicity. The other was afforded a ten-week treatment-free period during which the left ventricular ejection fraction returned to normal. All recovery animal hearts were microscopically and ultrastructurally normal. High-dose BILN 2061 cardiotoxicity in Rhesus monkeys appeared early in the treatment regimen and exhibited reversibility. A reliable biomarker has yet to be identified.
  • Author:
    Huang YM; Harrall KK; Dryman BA; Beach NM; Kenney SP; Opriessnig T; Vaughn EM; Roof MB; Meng XJ
    Title:
    Expression of the putative ORF1 capsid protein of torque teno sus virus 2 (TTSuV2) and development of Western blot and ELISA serodiagnostic assays: Correlation between TTSuV2 viral load and IgG antibody level in pigs.
    Source:
    Virus Res 158 (1-2), 79-88 (2011)
    Abstract:
    Porcine Torque teno virus (TTV) has a single-stranded circular DNA genome and is currently classified into a new genus Iotatorquevirus with two species in a newly established family Anelloviridae. Viral DNA of both porcine TTV species (TTSuV1 and TTSuV2) has a high prevalence in both healthy and diseased pigs worldwide and multiple infections of TTSuV with distinct genotypes or subtypes of the same species has been documented in the United States and in Europe. However, the prevalence of specific TTSuV antibodies in pigs remains unknown. In this study, the putative ORF1 capsid protein from TTSuV2 isolate PTTV2c-VA was expressed in Escherichia coli. The purified recombinant ORF1 protein was used as the antigen for the development of Western blot and indirect ELISA to detect TTSuV2-specific IgG antibodies in pig sera. The results revealed a relatively high rate of seropositivity to TTSuV2 in conventional pigs from different sources but not in gnotobiotic pigs. Overall, pigs with undetectable TTSuV2 viral load were more likely to have a lower anti-TTSuV2 antibody level. An analysis of 10 conventional pigs during a 2-month period showed that decreased viral loads or presumed virus clearance were associated with elevated anti-ORF1 IgG antibody levels. Interestingly, porcine circovirus associated disease (PCVAD)-affected pigs had a significantly lower level of TTSuV2 antibody than PCVAD-unaffected pigs (p < 0.01). This is the first study to establish essential serodiagnostic tools for investigation of TTSuV seroprevalence and infection dynamics, which will help elucidate the potential pathogenicity of TTSuV infection in pigs.
  • Author:
    Lemke ChT; Goudreau N; Zhao S; Hucke O; Thibeault D; Llinas-Brunet M; White PW
    Title:
    Combined X-ray, NMR, and kinetic analyses reveal uncommon binding characteristics of the hepatitis C virus NS3-NS4A protease inhibitor BI 201335.
    Source:
    J Biol Chem 286 (13), 11434-11443 (2011)
    Abstract:
    Hepatitis C virus infection, a major cause of liver disease worldwide, is curable, but currently approved therapies have suboptimal efficacy. Supplementing these therapies with direct-acting antiviral agents has the potential to considerably improve treatment prospects for hepatitis C virus-infected patients. The critical role played by the viral NS3 protease makes it an attractive target, and despite its shallow, solvent-exposed active site, several potent NS3 protease inhibitors are currently in the clinic. BI 201335, which is progressing through Phase IIb trials, contains a unique C-terminal carboxylic acid that binds noncovalently to the active site and a bromo-quinoline substitution on its proline residue that provides significant potency. In this work we have used stopped flow kinetics, x-ray crystallography, and NMR to characterize these distinctive features. Key findings include: slow association and dissociation rates within a single-step binding mechanism; the critical involvement of water molecules in acid binding; and protein side chain rearrangements, a bromine-oxygen halogen bond, and profound pK(a) changes within the catalytic triad associated with binding of the bromo-quinoline moiety.
  • Author:
    Kwong AD; Najera I; Bechtel J; Bowden S; Fitzgibbon J; Harrington P; Kempf D; Kieffer TL; Koletzki D; Kukolj G; Lim S; Pilotmatias T; Lin K; Mani N; Mo H; O'Rear J; Otto M; Parkin N; Pawlotsky J; Petropoulos C; Picchio G; Ralston R; Reeves JD; Schooley RT; Seiwert S; Standring D; Stuyver L; Sullivan J; Miller V
    Title:
    Sequence and phenotypic analysis for resistance monitoring in hepatitis C virus drug development recommendations from the HCV DRAG.
    Source:
    Gastroenterology 140 (3), 755-760.e12 (2011)
  • Author:
    Opriessnig T; Madson DM; Roof M; Layton SM; Ramamoorthy S; Meng XJ; Halbur PG
    Title:
    Experimental reproduction of porcine circovirus type 2 (PCV2)-associated enteritis in pigs infected with PCV2 alone or concurrently with lawsonia intracellularis or salmonella typhimurium.
    Source:
    J Comp Pathol 145 (2-3), 261-270 (2011)
    Abstract:
    Porcine circovirus (PCV)-associated disease (PCVAD) has emerged to become one of the most economically important pig diseases globally. One of the less commonly recognized clinical manifestations of PCVAD is PCV2 type 2 (PCV2)-associated enteritis in growing pigs; however, experimental confirmation of the ability of PCV2 alone or PCV2 coinfection with other agent(s) to induce enteritis is lacking. In this study, 120 specific-pathogen-free (SPF) pigs were divided randomly into six groups: controls (negative control pigs), PCV2 (inoculated with PCV2), LAW (inoculated with Lawsonia intracellularis), SALM (inoculated with Salmonella typhimurium), PCV2-LAW (concurrently inoculated with PCV2 and Lawsonia intracellularis) and PCV2-SALM (concurrently inoculated with PCV2 and Salmonella typhimurium). One half of the pigs in each group were subject to necropsy examination 14 days postinoculation (dpi) and the remaining pigs were examined at 28 dpi. The average daily weight gain was not different (P > 0.05) between groups. Individual pigs inoculated orally with PCV2 regardless of coinfection status (2/10 PCV2, 1/10 PCV2-LAW, 3/10 PCV2-SALM) developed PCVAD with diarrhoea and reduced weight gain or weight loss between 14 and 28 dpi. Those pigs had characteristic microscopic lesions in lymphoid and enteric tissues associated with abundant PCV2 antigen. Enteric lesions were characterized by necrosuppurative and proliferative enteritis with crypt elongation and epithelial hyperplasia in LAW and PCV2-LAW pigs by 14 dpi, ulcerative and necrosuppurative colitis in SALM and PCV2-SALM pigs by 14 dpi, and lymphohistiocytic enteritis with depletion of Peyer's patches in PCV2, PCV2-SALM and PCV2-LAW pigs by 28 dpi. To the authors' knowledge, this is the first report documenting that under experimental conditions, PCV2 can induce enteritis independently from other enteric pathogens and that oral challenge is a potentially important route and perhaps the natural route of PCV2 transmission in growing pigs.
  • Author:
    Sabo JP; Cong XJ; Kraft M-F; Wallace L; Castles MA; Mauss St; Macgregor ThR
    Title:
    Lack of a pharmacokinetic interaction between steady-state tipranavir/ritonavir and single-dose valacyclovir in healthy volunteers.
    Source:
    Eur J Clin Pharmacol 67 (3), 277-281 (2011)
    Abstract:
    OBJECTIVE: This study assessed the single-dose pharmacokinetics of the herpes antiviral acyclovir (administered as the pro-drug valacyclovir) alone and in combination with twice-daily 200 mg ritonavir-boosted tipranavir (500 mg) at steady state. METHODS: The study was an open label, one-sequence cross-over pharmacokinetic study in HIV-negative adults. Plasma drug concentrations were measured by validated LC/MS/MS assays; pharmacokinetics (AUC, C(max)) were determined using noncompartmental methods. The geometric mean ratio and 90% confidence interval [GMR, 90% CI] were used to evaluate the drug interaction. RESULTS: Twenty-six of 29 subjects completed the trial. With steady-state tipranavir/ritonavir, acyclovir C(max) decreased 4.9% [0.95, 0.88-1.02] and AUC increased 6.6% [1.07, 1.04-1.09]. The majority of subjects experienced at least one adverse event, most of which were mild gastrointestinal disorders. Three subjects discontinued tipranavir/ritonavir treatment as a result of drug-related increases in ALT/AST, including one subject who experienced mild upper abdominal pain. All subjects recovered without sequelae. CONCLUSIONS: When administered as a single dose of valacyclovir with steady-state tipranavir/ritonavir, there were no clinically important changes in acyclovir pharmacokinetics. This result indicates that valacyclovir can be co-administered safely with no dose adjustments. - DERWENT Abstract - This open-label study evaluated the single-dose pharmacokinetics of acyclovir along and in combination with ritonavir (RTV)-boosted tipranavir (TPV) in 29 healthy subjects. With steady-state tipranavir/ritonavir, acyclovir Cmax decreased 4.9% and AUC increased 6.6%. The majority of subjects experienced at least 1 adverse event (AE), most of which were mild GI disorders. 3 Subjects discontinued tipranavir/ritonavir treatment as a result of drug-related increases in ALT/AST, including 1 subject who experienced mild upper abdominal pain. All subjects recovered without sequelae. Thus, when administered as a single dose of valacyclovir with steady-state tipranavir (TPV)/ritonavir (RTV), there were no clinically important changes in acyclovir pharmacokinetics. Methods 29 Healthy subjects (aged 28-58 yr, mean 43.1 yr, 25 male) received valacyclovir (500 mg) alone or in combination with RTV (100 mg b.i.d.)/TPV (250 mg b.i.d.). Results For safety reasons, 3 subjects prematurely discontinued the study during treatment with TPV and RTV as a result of AE. These patients developed DAIDS grade 3 or 4 with rapidly increasing ALT and/or AST elevations which occurred between days 7 and 8 of the study. A total of 70 AE in 24/29 subjects treated were reported. The most common AE observed were increased ALT, headache, increased AST, diarrhea, flatulence and nausea, and renal pain. Steady-state TPV and RTV had no effect on acyclovir Cmax or AUC. Although some individuals showed an observable increase or decrease in maximum plasma acyclovir concentrations or systemic exposure with concomitant TPV and RTV exposure, the increases or decreases were balanced in the sample group and an overall trend was not observed. A single dose of valacyclovir had no effect on TPV Cmax, plasma concentration 12 hr post-dose or AUC0-12hr.
  • Author:
    White PW; Faucher A-M; Goudreau N
    Title:
    Small molecule inhibitors of the human papillomavirus e1-e2 interaction.
    Source:
    Curr Top Microbiol Immunol 348, 61-88 (2011)
    Abstract:
    Human papillomaviruses are responsible for multiple human diseases, including cervical cancer caused by multiple high-risk types and genital warts caused by the low-risk types 6 and 11. Based on the research indicating that low-risk HPV could be successfully targeted by inhibitors of viral DNA replication, we carried out several high-throughput screens for inhibitors of DNA replication activities. Two series were identified in screens for inhibitors of the interaction between the viral proteins E1 and E2. The two series were demonstrated to bind to overlapping sites on the transactivation domain of E2, at the E1-binding interface, by a series of biochemical and biophysical experiments. A member of the first series was also cocrystallized with the E2 transactivation domain. For both series, structure-activity investigations are described, which resulted in several hundred fold improvements in activity. The best compounds in each series had low nanomolar activity against the HPV11 E1-E2 interaction, and EC(50) values in cellular DNA replication assays of approximately 1 .mu.M. Binding modes for the two series are compared, and some general conclusions about the discovery of protein-protein interaction inhibitors are drawn from the work described.
  • Author:
    Khan AG; Pickl-Herk A; Gajdzik L; Marlovitis TC; Fuchs R; Blaas D
    Title:
    Entry of a heparan sulphate-binding HRV8 variant strictly depends on dynamin but not on clathrin, caveolin and flotillin.
    Source:
    Virology 412 (1), 55-67 (2011)
    Abstract:
    The major group human rhinovirus type 8 can enter cells via heparan sulphate. When internalized into ICAM-1 negative rhabdomyosarcoma (RD) cells, HRV8 accumulated in the cells but caused CPE only after 3 days when used at high MOI. Adaptation by three blind passages alternating between RD and HeLa cells resulted in variant HRV8v with decreased stability at acidic pH allowing for productive infection in the absence of ICAM-1. HRV8v produced CPE at 10 times lower MOI within 1 day. Confocal fluorescence microscopy colocalization and the use of pharmacological and dominant negative inhibitors revealed that viral uptake is clathrin, caveolin, and flotillin independent. However, it is blocked by dynasore, amiloride, and EIPA. Furthermore, HRV8v induced FITC-dextran uptake and colocalized with this fluid phase marker. Except for the complete inhibition by dynasore, the entry pathway of HRV8v via HS is similar to that of HRV14 in RD cells that overexpress ICAM-1.
  • Author:
    Geitmann M; Dahl G; Danielson UH
    Title:
    Mechanistic and kinetic characterization of hepatitis C virus NS3 protein interactions with NS4A and protease inhibitors.
    Source:
    J Mol Recognit 24 (1), 60-70 (2011)
    Abstract:
    The mechanism and kinetics of the interactions between ligands and immobilized full-length hepatitis C virus (HCV) genotype 1a NS3 have been characterized by SPR biosensor technology. The NS3 interactions for a series of NS3 protease inhibitors as well as for the NS4A cofactor, represented by a peptide corresponding to the sequence interacting with the enzyme, were found to be heterogeneous. It may represent interactions with two stable conformations of the protein. The NS3-NS4A interaction consisted of a high-affinity (K <sub>D</sub> = 50 nM) and a low-affinity (K<sub>D</sub> = 2 Â?M) interaction, contributing equally to the overall binding. By immobilizing NS3 alone or together with NS4A it was shown that all inhibitors had a higher affinity for NS3 in the presence of NS4A. NS4A thus has a direct effect on the binding of inhibitors to NS3 and not only on catalysis. As predicted, the mechanism-based inhibitor VX 950 exhibited a time-dependent interaction with a slow formation of a stable complex. BILN 2061 or ITMN-191 showed no signs of time-dependent interactions, but ITMN-191 had the highest affinity of the tested compounds, with both the slowest dissociation (k<sub>off</sub>) and fastest association rate, closely followed by BILN 2061. The k<sub>off</sub> for the inhibitors correlated strongly with their NS3 protease inhibitory effect as well as with their effect on replication of viral proteins in replicon cell cultures, confirming the relevance of the kinetic data. This approach for obtaining kinetic and mechanistic data for NS3 protease inhibitor and cofactor interactions is expected to be of importance for understanding the characteristics of HCV NS3 functionality as well as for anti-HCV lead discovery and optimization.
  • Author:
    Bin L; Howell MD; Kim BE; Streib JE; Hall CF; Leung DYM
    Title:
    Specificity protein 1 is pivotal in the skin's antiviral response.
    Source:
    J Allergy Clin Immunol 127 (2), 430-438 (2011)
    Abstract:
    Background: Previous studies have found specificity protein (Sp) 1 transcription factor in the viral replication machinery and postulated that Sp1 was required for viral replication in host cells. Objectives: We investigated the role of Sp1 in the skin's antiviral responses from the perspective of host defense and its biological relevance in patients with atopic dermatitis and a history of eczema herpeticum (ADEH<sup>+</sup>). Methods: Small interfering RNA duplexes were used to knock down Sp1 in keratinocytes. The expression of vaccinia virus (VV), herpes simplex virus 1, and other genes were evaluated by real-time PCR, or combined with Western blot and immunohistofluorescence staining. A total of 106 human subjects participated in this study. Results: Both VV and herpes simplex virus 1 replication were enhanced in Sp1 knocked-down keratinocytes. Sp1 gene expression was significantly decreased in ADEH <sup>+</sup> subjects compared with patients with atopic dermatitis without a history of eczema herpeticum and nonatopic subjects (P &lt; .0001) and inversely correlated with VV DNA copy number in human skin explants incubated with VV in vitro (partial correlation r = -0.256; P = .009). Gene profiling revealed that the antiviral genes, double-stranded RNA-dependent protein kinase (PKR) and 2'5'-oligoadenylate synthetase 2 (OAS2), were significantly downregulated in Sp1-silenced keratinocytes. Gene expression of PKR and OAS2 was also significantly decreased in skin biopsies from ADEH<sup>+</sup> subjects compared with patients with atopic dermatitis without a history of eczema herpeticum and nonatopic subjects. IFN-? augmented the antiviral capacity of Sp1-silenced keratinocytes. Conclusion: Specificity protein 1 knockdown enhances viral replication in keratinocytes by downregulating gene expression of PKR and OAS2. Sp1 deficiency in ADEH<sup>+</sup> patients may contribute to their increased propensity to disseminated skin viral infections. IFN-? augmentation may be a potential treatment for ADEH<sup>+</sup> patients.
  • Author:
    Wagner J; Kneucker A; Liebler-Tenorlo E; Fachinger V; Glaser M; Pesch S; Murtaugh MP; Reinhold P
    Title:
    Respiratory function and pulmonary lesions in pigs infected with porcine reproductive and respiratory syndrome virus.
    Source:
    Vet J (Lond) 187 (3), 310-319 (2011)
    Abstract:
    Pulmonary dysfunction was evaluated in pigs infected with porcine reproductive and respiratory syndrome virus (PRRSV, isolate VR-2332) and compared to clinical and pathological findings. Infected pigs developed fever, reduced appetite, respiratory distress and dullness at 9. days post-inoculation (dpi). Non-invasive pulmonary function tests using impulse oscillometry and rebreathing of test gases (He, CO) revealed peripheral airway obstruction, reduced lung compliance and reduced lung CO-transfer factor. PRRSV-induced pulmonary dysfunction was most marked at 9-18 dpi and was accompanied by a significantly increased respiratory rate and decreased tidal volume. Expiration was affected more than inspiration. On histopathological examination, multifocal areas of interstitial pneumonia (more severe and extensive at 10 dpi than 21 dpi) were identified as a possible structural basis for reduced lung compliance and gas exchange disturbances.
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