title: A high throughput Cre-lox activated viral membrane fusion assay identifies pharmacological inhibitors of HIV entry.
abstract: Enveloped virus entry occurs when viral and cellular membranes fuse releasing particle contents into the target cell. Human immunodeficiency virus (HIV) entry occurs by cell-free virus or virus transferred between infected and uninfected cells through structures called virological synapses. We developed a high-throughput cell-based assay to identify small molecule inhibitors of cell-free or virological synapse-mediated entry. An HIV clone carrying Cre recombinase as a Gag-internal gene fusion releases active Cre into cells upon viral entry activating a recombinatorial gene switch changing dsRed to GFP-expression. A screen of a 1998 known-biological profile small molecule library identified pharmacological HIV entry inhibitors that block both cell-free and cell-to-cell infection. Many top hits were noted as HIV inhibitors in prior studies, but not previously recognized as entry antagonists. Modest therapeutic indices for simvastatin and nigericin were observed in confirmatory HIV infection assays. This robust assay is adaptable to study HIV and heterologous viral pseudotypes.
authors.affiliation: Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, Immunology Institute, New York, NY, United States.
authors.forename: Anthony M
authors.initials: AM
authors.lastname: Esposito
authors.affiliation: Integrated Screening Core, Icahn School of Medicine at Mount Sinai, New York, NY, United States.
authors.forename: Pamela
authors.initials: P
authors.lastname: Cheung
authors.affiliation: Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, Immunology Institute, New York, NY, United States.
authors.forename: Talia H
authors.initials: TH
authors.lastname: Swartz
authors.affiliation: Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, Immunology Institute, New York, NY, United States.
authors.forename: Hongru
authors.initials: H
authors.lastname: Li
authors.affiliation: Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY, United States.
authors.forename: Tshidi
authors.initials: T
authors.lastname: Tsibane
authors.affiliation: Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, Immunology Institute, New York, NY, United States.
authors.forename: Natasha D
authors.initials: ND
authors.lastname: Durham
authors.affiliation: Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY, United States.
authors.forename: Christopher F
authors.initials: CF
authors.lastname: Basler
authors.affiliation: Integrated Screening Core, Icahn School of Medicine at Mount Sinai, New York, NY, United States.
authors.forename: Dan P
authors.initials: DP
authors.lastname: Felsenfeld
authors.affiliation: Division of Infectious Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, Immunology Institute, New York, NY, United States. Electronic address: benjamin.chen@mssm.edu.
authors.forename: Benjamin K
authors.initials: BK
authors.lastname: Chen
mesh_terms: D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000821:virology; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D023581:HIV Fusion Inhibitors / Q000494:pharmacology*; D015658:HIV Infections / Q000821:virology*; D015497:HIV-1 / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D014773:Virology / Q000379:methods*; D053586:Virus Internalization / Q000187:drug effects; D014779:Virus Replication / Q000187:drug effects
chemical_list: D023581:HIV Fusion Inhibitors
keywords: Cre recombinase; Drug screening; Env; Gag; HIV-1; Viral membrane fusion; Virus entry