A broad-spectrum antiviral targeting entry of enveloped viruses

Wolf, Mike C.; Freiberg, Alexander N.; Zhang, Tinghu; Akyol-Ataman, Zeynep; Grock, Andrew; Hong, Patrick; Lǐ, Jiànróng; Watson, Natalya F.; Fang, Angela Q.; Aguilar, Hector C.; Porotto, Matteo; Honko, Anna N.; Damoiseaux, Robert; Miller, John P.; Woodson, Sara E.; Chantasirivisal, Steven; Fontanes, Vanessa; Negrete, Oscar; Krogstad, Paul; Dasgupta, Asim; Moscona, Anne; Hensley, Lisa E.; Whelan, Sean P. J.; Faull, Kym F.; Holbrook, Michael R.; Jung, Michael E.; Lee, Benhur

doi:10.1073/pnas.0909587107

Cited by 229 publications

(244 citation statements)

References 33 publications

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“…41,42,52,[76][77][78][79] Furthermore, cell fusion-based assays are generally not sensitive to inhibitors that inactivate diverse enveloped viruses through disrupting their membrane, while having no adverse effect on cell membranes. 39,40 Our observation that P2X1 receptor antagonists inhibit HIV-1 fusion is in agreement with the literature. Published studies implicate the P2X1 receptor in HIV-1 entry/fusion and in cell-cell transmission.…”

Section: Discussionsupporting

confidence: 81%

“…38 However, a cell-cell fusion-based screen should not detect inhibitors of virus endocytosis/trafficking steps of entry or virucidal compounds that selectively damaged the virus. 39,40 These considerations warrant the implementation of virus-cell fusion assay for HTS, which appears particularly important in the light of our finding that HIV-1 enters cells through endocytosis and fusion with endosomal compartments. 41,42 The results of such screen should reveal new targets for intervention-cellular factors involved in endocytosis and vesicular trafficking.…”

Section: Introductionmentioning

confidence: 99%

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High-Throughput HIV–Cell Fusion Assay for Discovery of Virus Entry Inhibitors

Marin

Giroud

et al. 2015

ASSAY and Drug Development Technologies

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HIV-1 initiates infection by merging its envelope membrane with the target cell membrane, a process that is mediated by the viral Env glycoprotein following its sequential binding to CD4 and coreceptors, CXCR4 or CCR5. Although HIV-1 fusion has been a target for antiviral therapy, the virus has developed resistance to drugs blocking the CCR5 binding or Env refolding steps of this process. This highlights the need for novel inhibitors. Here, we adapted and optimized an enzymatic HIV-cell fusion assay, which reports the transfer of virus-encapsulated b-lactamase into the cytoplasm, to high-throughput screening (HTS) with a 384-well format. The assay was robustly performed in HTS format and was validated by the pilot screen of a small library of pharmacologically active compounds. Several hits identified by screening included a prominent cluster of purinergic receptor antagonists. Functional studies demonstrated that P2X1 receptor antagonists selectively inhibited HIV-1 fusion without affecting the fusion activity of an unrelated virus that enters cells through an endocytic route. The inhibition of HIV-cell fusion by P2X1 antagonists was not through downmodulation of the cell surface expression of CD4 or coreceptors, thus implicating P2X1 receptor in the HIV-1 fusion step. The ability of these antagonists to inhibit viruses regardless of their coreceptor (CXCR4 or CCR5) preference indicates that fusion is blocked at a late step downstream of coreceptor binding. A future large-scale screening campaign for HIV-1 fusion inhibitors, using the above functional readout, will likely reveal novel classes of inhibitors and suggest potential targets for antiviral therapy.

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Section: Discussionsupporting

confidence: 81%

Section: Introductionmentioning

confidence: 99%

High-Throughput HIV–Cell Fusion Assay for Discovery of Virus Entry Inhibitors

Marin

Giroud

et al. 2015

ASSAY and Drug Development Technologies

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“…Interestingly, they also have a rigid and planar hydrophobic moiety attached to a hydrophilic head of larger diameter. The most potent compounds, LJ-001, -002, and -003 have a smaller hydrophobic moiety than dUY11 and an ∼200-fold higher IC 50 toward enveloped viruses (48). Such high IC 50 is consistent with the SAR data (Fig S1), and the mechanism of action herein proposed.…”

Section: Discussionsupporting

confidence: 78%

Rigid amphipathic fusion inhibitors, small molecule antiviral compounds against enveloped viruses

St.Vincent¹,

Colpitts

Устинов

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

141

130

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Antiviral drugs targeting viral proteins often result in prompt selection for resistance. Moreover, the number of viral targets is limited. Novel antiviral targets are therefore needed. The unique characteristics of fusion between virion envelopes and cell membranes may provide such targets. Like all fusing bilayers, viral envelopes locally adopt hourglass-shaped stalks during the initial stages of fusion, a process that requires local negative membrane curvature. Unlike cellular vesicles, however, viral envelopes do not redistribute lipids between leaflets, can only use the energy released by virion proteins, and fuse to the extracellular leaflets of cell membranes. Enrichment in phospholipids with hydrophilic heads larger than their hydrophobic tails in the convex outer leaflet of vesicles favors positive curvature, therefore increasing the activation energy barrier for fusion. Such phospholipids can increase the activation barrier beyond the energy provided by virion proteins, thereby inhibiting viral fusion. However, phospholipids are not pharmacologically useful. We show here that a family of synthetic rigid amphiphiles of shape similar to such phospholipids, RAFIs (rigid amphipathic fusion inhibitors), inhibit the infectivity of several otherwise unrelated enveloped viruses, including hepatitis C and HSV-1 and -2 (lowest apparent IC 50 48 nM), with no cytotoxic or cytostatic effects (selectivity index > 3,000) by inhibiting the increased negative curvature required for the initial stages of fusion.lipid bilayer fusion | DNA virus | RNA virus | Sindbis virus | nucleosides

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“…Beyond lipid envelope-rupturing peptides, a small molecule compound has been reported to physically disrupt lipid membranes [109]. Interestingly, this compound displays a general property of membrane disruption but only permanently damages the membranes of viruses and not host cells.…”

Section: Outlook On Engineering Strategies For Antiviral Drug Developmentioning

confidence: 99%

Model Membrane Platforms for Biomedicine: Case Study on Antiviral Drug Development

Jackman

Cho

2012

Biointerphases

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As one of the most important interfaces in cellular systems, biological membranes have essential functions in many activities such as cellular protection and signaling. Beyond their direct functions, they also serve as scaffolds to support the association of proteins involved in structural support, adhesion, and transport. Unfortunately, biological processes sometimes malfunction and require therapeutic intervention. For those processes which occur within or upon membranes, it is oftentimes difficult to study the mechanism in a biologically relevant, membranous environment. Therefore, the identification of direct therapeutic targets is challenging. In order to overcome this barrier, engineering strategies offer a new approach to interrogate biological activities at membrane interfaces by analyzing them through the principles of the interfacial sciences. Since membranes are complex biological interfaces, the development of simplified model systems which mimic important properties of membranes can enable fundamental characterization of interaction parameters for such processes. We have selected the hepatitis C virus (HCV) as a model viral pathogen to demonstrate how model membrane platforms can aid antiviral drug discovery and development. Responsible for generating the genomic diversity that makes treating HCV infection so difficult, viral replication represents an ideal step in the virus life cycle for therapeutic intervention. To target HCV genome replication, the interaction of viral proteins with model membrane platforms has served as a useful strategy for target identification and characterization. In this review article, we demonstrate how engineering approaches have led to the discovery of a new functional activity encoded within the HCV nonstructural 5A protein. Specifically, its N-terminal amphipathic, α-helix (AH) can rupture lipid vesicles in a size-dependent manner. While this activity has a number of exciting biotechnology and biomedical applications, arguably the most promising one is in antiviral medicine. Based on the similarities between lipid vesicles and the lipid envelopes of virus particles, experimental findings from model membrane platforms led to the prediction that a range of medically important viruses might be susceptible to rupturing treatment with synthetic AH peptide. This hypothesis was tested and validated by molecular virology studies. Broad-spectrum antiviral activity of the AH peptide has been identified against HCV, HIV, herpes simplex virus, and dengue virus, and many more deadly pathogens. As a result, the AH peptide is the first in class of broad-spectrum, lipid envelope-rupturing antiviral agents, and has entered the drug pipeline. In summary, engineering strategies break down complex biological systems into simplified biomimetic models that recapitulate the most important parameters. This approach is particularly advantageous for membrane-associated biological processes because model membrane platforms provide more direct characterization of target interactions than is possible with other methods. Consequently, model membrane platforms hold great promise for solving important biomedical problems and speeding up the translation of biological knowledge into clinical applications.

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A broad-spectrum antiviral targeting entry of enveloped viruses

Cited by 229 publications

References 33 publications

High-Throughput HIV–Cell Fusion Assay for Discovery of Virus Entry Inhibitors

High-Throughput HIV–Cell Fusion Assay for Discovery of Virus Entry Inhibitors

Rigid amphipathic fusion inhibitors, small molecule antiviral compounds against enveloped viruses

Model Membrane Platforms for Biomedicine: Case Study on Antiviral Drug Development

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