title: Functional analysis of the broadly neutralizing human anti-HIV-1 antibody 2F5 produced in transgenic BY-2 suspension cultures.
abstract: We report the production of an important human therapeutic antibody in plant cell suspension cultures and the functional analysis of that antibody, including a comparison with the same antibody produced in CHO cells. We established transgenic tobacco BY2 suspension cell cultures expressing the human monoclonal antibody 2F5, which shows broadly neutralizing activity against HIV-1. The antibody was directed to the endoplasmic reticulum of the plant cells and was isolated by cell disruption, followed by protein A chromatography. The plant-derived antibody was shown to be largely intact by SDS-PAGE and immunoblot. Antigen binding activity was investigated by electrophoretic mobility shift assay and quantitatively determined by ELISA and Biacore biosensor technology. Ligand binding properties were analyzed using the ectodomain of human Fc gammaRI for kinetic analysis. The plant-derived antibody showed similar kinetic properties and 89% of the binding capacity of its CHO-derived counterpart, but was only 33% as efficient in HIV-1 neutralization assays. Our results show that plant suspension cultures can be used to produce human antibodies efficiently and that the analysis methods used in this study, including biosensor technology, provide useful functional data about antibody performance. This highlights important issues raised by the use of plant systems to produce human biologics.
authors.affiliation: Institute of Molecular Biotechnology, RTWH Aachen University, Worringerweg 1, 52074 Aachen, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Sack
authors.forename: Antje
authors.initials: A
authors.lastname: Paetz
authors.forename: Renate
authors.initials: R
authors.lastname: Kunert
authors.forename: Michael
authors.initials: M
authors.lastname: Bomble
authors.forename: Friedemann
authors.initials: F
authors.lastname: Hesse
authors.forename: Gabriela
authors.initials: G
authors.lastname: Stiegler
authors.forename: Rainer
authors.initials: R
authors.lastname: Fischer
authors.forename: Hermann
authors.initials: H
authors.lastname: Katinger
authors.forename: Eva
authors.initials: E
authors.lastname: Stoeger
authors.forename: Thomas
authors.initials: T
authors.lastname: Rademacher
mesh_terms: D000937:Antigen-Antibody Reactions; D015374:Biosensing Techniques; D002478:Cells, Cultured; D003001:Cloning, Molecular / Q000379:methods*; D015483:HIV Antibodies / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D006801:Humans; D007700:Kinetics; D008722:Methods; D009500:Neutralization Tests; D059828:Plant Cells; D011994:Recombinant Proteins / Q000276:immunology; D014026:Nicotiana
chemical_list: D015483:HIV Antibodies; D011994:Recombinant Proteins