title: Intrabodies targeting the Kaposi sarcoma-associated herpesvirus latency antigen inhibit viral persistence in lymphoma cells.
abstract: Kaposi sarcoma-associated herpesvirus (KSHV) latency-associated nuclear antigen-1 (LANA1) is essential for the maintenance and segregation of viral episomes in KSHV latently infected B cells. We report development of intracellular, rabbit-derived antibodies generated by phage display technology, which bind to N-terminal LANA1 epitopes and neutralize the chromosome-binding activity of LANA1. Although these cloned single-chain variable fragments (scFvs) show relatively low binding affinities for the LANA1 viral antigen in in vitro assays, they nonetheless outcompete KSHV-seropositive human sera for LANA1 epitope binding. In heterologous cells, intracellular intrabody expression inhibits LANA1-dependent plasmid maintenance of both an artificial plasmid containing KSHV LANA1 binding sequences and a bacterial artificial chromosome containing the entire KSHV genome. In KSHV naturally infected primary effusion lymphoma cells, intracellular intrabody expression causes a reduction or loss of the typical LANA1 punctate, nuclear pattern. This morphologically apparent LANA1 dispersion correlates to loss of viral episome by molecular analysis. These data suggest a novel approach to antiherpes viral therapy and confirm LANA1 is critical target for neutralization of KSHV viral latency.
authors.affiliation: URIA-Centro de Patogénese Molecular, Faculty of Pharmacy, University of Lisbon, 1649-019 Lisbon, Portugal.
authors.forename: Sofia
authors.initials: S
authors.lastname: Corte-Real
authors.forename: Chris
authors.initials: C
authors.lastname: Collins
authors.forename: Frederico
authors.initials: F
authors.lastname: Aires da Silva
authors.forename: J Pedro
authors.initials: JP
authors.lastname: Simas
authors.forename: Carlos F
authors.initials: CF
authors.lastname: Barbas
authors.forename: Yuan
authors.initials: Y
authors.lastname: Chang
authors.forename: Patrick
authors.initials: P
authors.lastname: Moore
authors.forename: Joao
authors.initials: J
authors.lastname: Goncalves
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology*; D002460:Cell Line; D019288:Herpesvirus 8, Human / Q000502:physiology; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000276:immunology / Q000494:pharmacology*; D008223:Lymphoma / Q000821:virology*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000276:immunology*; D019151:Peptide Library; D011817:Rabbits; D012514:Sarcoma, Kaposi / Q000276:immunology / Q000821:virology*; D014162:Transfection; D017735:Virus Latency / Q000187:drug effects*
chemical_list: D000956:Antigens, Viral; D007135:Immunoglobulin Variable Region; D009687:Nuclear Proteins; D019151:Peptide Library; C109540:latency-associated nuclear antigen