We showed previously that eiger, the Drosophila tumor necrosis factor homolog, contributes to the pathology induced by infection with Salmonella typhimurium. We were curious whether eiger is always detrimental in the context of infection or if it plays a role in fighting some types of microbes. We challenged wild-type and eiger mutant flies with a collection of facultative intracellular and extracellular pathogens, including a fungus and Gram-positive and Gram-negative bacteria. The response of eiger mutants divided these microbes into two groups: eiger mutants are immunocompromised with respect to extracellular pathogens but show no change or reduced sensitivity to facultative intracellular pathogens. Hence, eiger helps fight infections but also can cause pathology. We propose that eiger activates the cellular immune response of the fly to aid clearance of extracellular pathogens. Intracellular pathogens, which can already defeat professional phagocytes, are unaffected by eiger.

No abstract

GS-9190 (Tegobuvir) is a novel imidazopyridine inhibitor of hepatitis C virus (HCV) RNA replication in vitroand has demonstrated potent antiviral activity in patients chronically infected with genotype 1 (GT1) HCV. GS-9190 exhibits reduced activity against GT2a (JFH1) subgenomic replicons and GT2a (J6/JFH1) infectious virus, suggesting that the compound's mechanism of action involves a genotype-specific viral component. To further investigate the GS-9190 mechanism of action, we utilized the susceptibility differences between GT1b and GT2a by constructing a series of replicon chimeras where combinations of 1b and 2a nonstructural proteins were encoded within the same replicon. The antiviral activities of GS-9190 against the chimeric replicons were reduced to levels comparable to that of the wild-type GT2a replicon in chimeras expressing GT2a NS5B. GT1b replicons in which the ␤-hairpin region (amino acids 435 to 455) was replaced by the corresponding sequence of GT2a were markedly less susceptible to GS-9190, indicating the importance of the thumb subdomain of the polymerase in this effect. Resistance selection in GT1b replicon cells identified several mutations in NS5B (C316Y, Y448H, Y452H, and C445F) that contributed to the drug resistance phenotype. Reintroduction of these mutations into wild-type replicons conferred resistance to GS-9190, with the number of NS5B mutations correlating with the degree of resistance. Analysis of GS-9190 cross-resistance against previously reported NS5B drug-selected mutations showed that the resistance pattern of GS-9190 is different from other nonnucleoside inhibitors. Collectively, these data demonstrate that GS-9190 represents a novel class of nonnucleoside polymerase inhibitors that interact with NS5B likely through involvement of the ␤-hairpin in the thumb subdomain.Hepatitis C virus (HCV) is a major cause of morbidity, affecting approximately 170 million people worldwide with an estimated 3 to 4 million additional new infections occurring each year (36). HCV is a positive-strand RNA virus with six major genotypes that are further divided into multiple subtypes. Due to the error-prone nature of its replication enzyme, a myriad of different viral quasispecies exists within an infected individual (32). With this high degree of viral variability, the current treatment regimen, which consists of weekly injections of pegylated alpha interferon (PEG-IFN) and twice-daily oral doses of ribavirin (RBV), is of limited efficacy and, in addition, carries significant side effects (8, 23). Although the HCV NS3/4A protease inhibitors telaprevir and boceprevir for treatment of chronic HCV infection will soon be available, these compounds will still need to be combined with the current standard of care (PEG-IFN/RBV) to be efficacious and will not cure all infected individuals (10,14,30). Therefore, the development of additional direct antiviral agents with diverse resistance profiles is necessary, with the ultimate goal of developing all-oral antiviral combinations that can achieve superior ...

A series of ethynyl-or (4-boronophenyl)bipyridines and -phenanthrolines were prepared as versatile building blocks for attachment of bidentate N-ligands to other molecules via cross-coupling reactions. Their complexation with Ru(bpy) 2 -Cl 2 gave the corresponding Ru II complexes. 9-Benzyladenine derivatives bearing the bipyridine or phenanthroline complexes in position 8, attached via a conjugate acetylene or phenylene linker were prepared by cross-coupling reactions of the ethynyl-or 4-boronophenylbipyridines and -phenanthrolines with 9-benzyl-8-bromoadenine. Their complexation with Ru(bpy) 2 Cl 2 afforded the corresponding Ru complexes as model compounds for electrochemical DNA labeling. The same compounds were also prepared directly by crosscoupling of 9-benzyl-8-bromoadenine with Ru complexes of the alkynes and boronic acids. Both approaches are compared in terms of potential applications for labeling of nucleic

GS-9669 is a highly optimized thumb site II nonnucleoside inhibitor of the hepatitis C virus (HCV) RNA polymerase, with a binding affinity of 1.35 nM for the genotype (GT) 1b protein. It is a selective inhibitor of HCV RNA replication, with a mean 50% effective concentration (EC 50 ) of <11 nM in genotype 1 and 5 replicon assays, but lacks useful activity against genotypes 2 to 4. The M423T mutation is readily generated clinically upon monotherapy with the thumb site II inhibitors filibuvir and lomibuvir, and it is notable that GS-9669 exhibited only a 3-fold loss in potency against this variant in the genotype 1b replicon. Rather than M423T, resistance predominantly tracks to residues R422K and L419M and residue I482L in GT 1b and 1a replicons, respectively. GS-9669 exhibited at least additive activity in combination with agents encompassing four other direct modes of action (NS3 protease, NS5A, NS5B via an alternative allosteric binding site, and NS5B nucleotide) as well as with alpha interferon or ribavirin in replicon assays. It exhibited high metabolic stability in in vitro human liver microsomal assays, which, in combination with its pharmacokinetic profiles in rat, dog, and two monkey species, is predictive of good human pharmacokinetics. GS-9669 is well suited for combination with other orally active, direct-acting antiviral agents in the treatment of genotype 1 chronic HCV infection. (This study has been registered at ClinicalTrials.gov under registration number NCT01431898.)

Eiger is the sole TNF family member found in Drosophila melanogaster. This signaling molecule is induced during infection and is required for an appropriate immune response to many microbes; however, little is known about where eiger is produced. Here, we show that eiger is made in the fly’s fat body during a Salmonella typhimurium infection. Using tissue-specific knockdown, we found that eiger expression in the fat body is required for all of the phenotypes we observed in eiger null mutant flies. This includes reduced melanization, altered antimicrobial peptide expression and reduced feeding rates. The effect of eiger on feeding rates alone may account for the entire phenotype seen in eiger mutants infected with S. typhimurium.

Bacterial protein synthesis is the target for several classes of established antibiotics. This report describes the characterization of a novel translation inhibitor produced by the soil bacterium Flexibacter. The dipeptide antibiotic TAN1057 A/B was synthesized and designated GS7128. As reported previously, TAN1057 inhibits protein synthesis in both Escherichia coli and Staphylococcus aureus, leaving transcription unaffected. Cell-free translation systems from E. coli were used to further dissect the mechanism of translational inhibition. Binding of mRNA to ribosomes was unaffected by the drug, whereas the initiation reaction was reduced. Elongation of translation was completely inhibited by GS7128. Detailed analysis showed that the peptidyl transferase reaction was strongly inhibited, whereas tRNA binding to both A- and P-site was unaffected. Selection and analysis of drug-resistant mutants of S. aureus suggests that drug uptake may be mediated by a dipeptide transport mechanism.

GS-327073 is a potent in vitro inhibitor of HCV replication either alone or in combination with other selective HCV inhibitors.