title: The Glycosylphosphatidylinositol-Anchored Variable Region of Llama Heavy Chain-Only Antibody JM4 Efficiently Blocks both Cell-Free and T Cell-T Cell Transmission of Human Immunodeficiency Virus Type 1.
abstract: The variable regions (VHHs) of two heavy chain-only antibodies, JM2 and JM4, from llamas that have been immunized with a trimeric gp140 bound to a CD4 mimic have been recently isolated (here referred to as VHH JM2 and VHH JM4, respectively). JM2 binds the CD4-binding site of gp120 and neutralizes HIV-1 strains from subtypes B, C, and G. JM4 binds gp120 and neutralizes HIV-1 strains from subtypes A, B, C, A/E, and G in a CD4-dependent manner. In the present study, we constructed glycosylphosphatidylinositol (GPI)-anchored VHH JM2 and JM4 along with an E4 control and transduced them into human CD4+cell lines and primary CD4 T cells. We report that by genetically linking the VHHs with a GPI attachment signal, VHHs are targeted to the lipid rafts of the plasma membranes. Expression of GPI-VHH JM4, but not GPI-VHH E4 and JM2, on the surface of transduced TZM.bl cells potently neutralizes multiple subtypes of HIV-1 isolates, including tier 2 or 3 strains, transmitted founders, quasispecies, and soluble single domain antibody (sdAb) JM4-resistant viruses. Moreover, transduction of CEMss-CCR5 cells with GPI-VHH JM4, but not with GPI-VHH E4, confers resistance to both cell-free and T cell-T cell transmission of HIV-1 and HIV-1 envelope-mediated fusion. Finally, GPI-VHH JM4-transduced human primary CD4 T cells efficiently resist both cell-free and T cell-T cell transmission of HIV-1. Thus, we conclude that VHH JM4, when targeted to the lipid rafts of the plasma membrane, efficiently neutralizes HIV-1 infection via both cell-free and T cell-T cell transmission. Our findings should have important implications for GPI-anchored antibody-based therapy against HIV-1.
  
  
  OBJECTIVE
  Lipid rafts are specialized dynamic microdomains of the plasma membrane and have been shown to be gateways for HIV-1 budding as well as entry into T cells and macrophages. In nature, many glycosylphosphatidylinositol (GPI)-anchored proteins localize in the lipid rafts. In the present study, we developed GPI-anchored variable regions (VHHs) of two heavy chain-only antibodies, JM2 and JM4, from immunized llamas. We show that by genetically linking the VHHs with a GPI attachment signal, VHHs are targeted to the lipid rafts of the plasma membranes. GPI-VHH JM4, but not GPI-VHH JM2, in transduced CD4+cell lines and human primary CD4 T cells not only efficiently blocks diverse HIV-1 strains, including tier 2 or 3 strains, transmitted founders, quasispecies, and soluble sdAb JM4-resistant strains, but also efficiently interferes T cell-T cell transmissions of HIV-1 and HIV-1 envelope-mediated fusion. Our findings should have important implications in GPI-anchored antibody-based therapy against HIV-1.
authors.affiliation: Unit of Anti-Viral Immunity and Genetic Therapy, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Lihong
authors.initials: L
authors.lastname: Liu
authors.affiliation: Unit of Anti-Viral Immunity and Genetic Therapy, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Weiming
authors.initials: W
authors.lastname: Wang
authors.affiliation: Institut Cochin-INSERM U1016-CNRS UMR 8104-University Paris Descartes, Paris, France.
authors.forename: Julie
authors.initials: J
authors.lastname: Matz
authors.affiliation: Unit of Anti-Viral Immunity and Genetic Therapy, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Chaobaihui
authors.initials: C
authors.lastname: Ye
authors.affiliation: Institut Cochin-INSERM U1016-CNRS UMR 8104-University Paris Descartes, Paris, France.
authors.forename: Lucie
authors.initials: L
authors.lastname: Bracq
authors.affiliation: Institut Cochin-INSERM U1016-CNRS UMR 8104-University Paris Descartes, Paris, France.
authors.forename: Jerome
authors.initials: J
authors.lastname: Delon
authors.affiliation: Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas, USA.
authors.forename: Jason T
authors.initials: JT
authors.lastname: Kimata
authors.affiliation: HKU-AIDS Institute, Department of Microbiology, Research Center of Infection and Immunology, The University of Hong Kong, Hong Kong Special Administrative Region, China.
authors.forename: Zhiwei
authors.initials: Z
authors.lastname: Chen
authors.affiliation: Institut Cochin-INSERM U1016-CNRS UMR 8104-University Paris Descartes, Paris, France serge.benichou@inserm.fr blzhou@sibs.ac.cn.
authors.forename: Serge
authors.initials: S
authors.lastname: Benichou
authors.affiliation: Unit of Anti-Viral Immunity and Genetic Therapy, Key Laboratory of Molecular Virology and Immunology, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China serge.benichou@inserm.fr blzhou@sibs.ac.cn.
authors.forename: Paul
authors.initials: P
authors.lastname: Zhou
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000235:genetics / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology* / Q000821:virology*; D002161:Camelids, New World / Q000235:genetics / Q000276:immunology*; D002460:Cell Line; D002474:Cell-Free System; D058851:GPI-Linked Proteins / Q000235:genetics / Q000276:immunology; D015483:HIV Antibodies / Q000235:genetics / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000276:immunology; D015658:HIV Infections / Q000276:immunology / Q000628:therapy / Q000821:virology; D015497:HIV-1 / Q000276:immunology*; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000276:immunology; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology; D021962:Membrane Microdomains / Q000276:immunology; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology
chemical_list: D057134:Antibodies, Neutralizing; D058851:GPI-Linked Proteins; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; C514793:gp120 protein, Human immunodeficiency virus 1