Hepatitis C virus (HCV) infection is a serious cause of chronic liver disease worldwide with more than 170 million infected individuals at risk of developing significant morbidity and mortality. Current interferon-based therapies are suboptimal especially in patients infected with HCV genotype 1, and they are poorly tolerated, highlighting the unmet medical need for new therapeutics. The HCV-encoded NS3 protease is essential for viral replication and has long been considered an attractive target for therapeutic intervention in HCV-infected patients. Here we identify a class of specific and potent NS3 protease inhibitors and report the evaluation of BILN 2061, a small molecule inhibitor biologically available through oral ingestion and the first of its class in human trials. Administration of BILN 2061 to patients infected with HCV genotype 1 for 2 days resulted in an impressive reduction of HCV RNA plasma levels, and established proof-of-concept in humans for an HCV NS3 protease inhibitor. Our results further illustrate the potential of the viral-enzyme-targeted drug discovery approach for the development of new HCV therapeutics.

IL-10 is a potent anti-inflammatory cytokine interfering with antigen presentation by inducing the intracellular sequestration of MHC class II (MHC-II) molecules. Here we studied the contribution of membrane-associated RING-CH (MARCH) ubiquitin ligase family members to the IL-10-induced down-regulation of MHC-II molecules. We found that MARCH1 and MARCH8 proteins are the most potent family members for the downregulation of MHC-II surface expression in transfected cells, but only MARCH1 mRNA expression is strongly induced by IL-10 in human primary monocytes. We detected monoand poly-ubiquitinated forms of MHC-II molecules both in IL-10-treated monocytes and in cells transfected with MARCH1. We also show direct interaction between MHC-II and MARCH1 molecules in co-immunoprecipitation assays. Finally, we found that siRNAmediated knockdown of MARCH1 reverses IL-10-induced MHC-II down-regulation in primary monocytes. Thus, the immunosuppressive effect of IL-10 on antigen presentation is mediated through induced expression of MARCH1.

CommunicationsPotent and selective macrocyclic inhibitors of the hepatitis C virus NS3 serine protease based on the conformation of a enzyme-bound substratelike hexapeptide demonstrate many of the desirable properties of a druglike archetype, which could lead to an antiviral agent for the treatment of hepatitis C in man. For more details see the following communication by Tsantrizos et al.

The mitochondrial antiviral signaling (MAVS) protein plays a central role in innate antiviral immunity. Upon recognition of a virus, intracellular receptors of the RIG-I-like helicase family interact with MAVS to trigger a signaling cascade. In this study, we investigate the requirement of the MAVS structure for enabling its signaling by structure-function analyses and resonance energy transfer approaches in live cells. We now report the essential role of the MAVS oligomer in signal transduction and map the transmembrane domain as the main determinant of dimerization. A combination of mutagenesis and computational methods identified a cluster of residues making favorable van der Waals interactions at the MAVS dimer interface. We also correlated the activation of IRF3 and NF-B with MAVS oligomerization rather than its mitochondrial localization. Finally, we demonstrated that MAVS oligomerization is disrupted upon expression of HCV NS3/4A protease, suggesting a mechanism for the loss of antiviral signaling. Altogether, our data suggest that the MAVS oligomer is essential in the formation of a multiprotein membrane-associated signaling complex and enables downstream activation of IRF3 and NF-B in antiviral innate immunity.Viral nucleic acids, potent inducers of the antiviral innate immune response, are recognized at the extracellular level by a subset of endosomal Toll-like receptors and, upon permissive virus infection, at the cytoplasmic level by a family of DexD/H box RNA helicases including RIG-I (retinoic acid inducible gene I) and MDA5 (melanoma differentiation-associated gene 5) (26).The sensor protein RIG-I is believed to be maintained in an auto-inhibited state in resting cells and to undergo a conformational change upon viral RNA binding. This conformational change exposes two N-terminal caspase activation and recruitment domains (CARDs) (36), induces cytoplasmic oligomerization (31), and promotes interaction with mitochondrial antiviral signaling (MAVS) protein (also known as IPS-1, Cardif, and VISA). The interaction between RIG-I and MAVS occurs through the protein-interacting CARDs of both proteins (17,25,32,34) and initiates formation of an as-yet-undefined macromolecular signaling complex to the mitochondrial membrane. Formation of this complex involves the recruitment of multiple signaling components to activate interferon (IFN) regulatory factor 3 (IRF3) and nuclear factor-B (NF-B) transcription factors that are required for production of type-I IFNs (reviewed in reference 26). The RIG-I/MAVS pathway plays an important role in the antiviral host response to hepatitis C virus (HCV) infection, where the uncapped viral RNA of HCV triggers RIG-I signaling (33). However, HCV can counteract the antiviral response; its NS3/4A serine protease cleaves MAVS at cysteine 508, resulting in the loss of mitochondrial localization and the abrogation of signaling function (9,19,20,25). This mechanism has been confirmed to occur in infected cells (22).Here, we applied fluorescence and bioluminescence resonance energ...