title: Secretory expression of a bispecific antibody targeting tumor necrosis factor and ED-B fibronectin in Pichia pastoris and its functional analysis.
abstract: Specific targeting of tumor necrosis factor (TNF)-α antagonist to the inflamed site could increase its efficacy and reduce side-effects. Here, we constructed a bispecific diabody (BsDb) that targets TNF-α and ED-B-containing fibronectin, a fibronectin isoform specifically expressed in the pannus of the inflamed synovium in rheumatoid arthritis. BsDb was secreted from Pichia pastoris as functional protein and was purified to homogeneity. BsDb could simultaneously bind to human TNF-α and B-FN and neutralize TNF-α action. Additionally, BsDb showed a significant gain both in the antigen-binding affinity and in TNF-α-neutralizing ability as compared to its original antibodies, L19 and anti-TNF-α scFv, which were produced in E. coli. BsDb was constructed and was endowed with enhanced bioactivities and improved production processing. Therefore, it holds great potential for in vivo applications.
authors.affiliation: Center for Infection and Immunity Research, School of Life Sciences, Hubei University, Youyi Road 368, Wuhan, 430062, China, liumengy@tom.com.
authors.forename: Meng-yuan
authors.initials: MY
authors.lastname: Liu
authors.forename: Xue-ping
authors.initials: XP
authors.lastname: Hu
authors.forename: Mian
authors.initials: M
authors.lastname: Xie
authors.forename: Si-jing
authors.initials: SJ
authors.lastname: Jiang
authors.forename: Lu-jun
authors.initials: LJ
authors.lastname: Li
authors.forename: Dong-xu
authors.initials: DX
authors.lastname: Liu
authors.forename: Xiao-song
authors.initials: XS
authors.lastname: Yang
mesh_terms: D018033:Antibodies, Bispecific / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D057134:Antibodies, Neutralizing / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D005353:Fibronectins / Q000276:immunology*; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D009500:Neutralization Tests; D010843:Pichia / Q000235:genetics*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology*
chemical_list: D018033:Antibodies, Bispecific; D057134:Antibodies, Neutralizing; C067625:FN1 protein, human; D005353:Fibronectins; D011994:Recombinant Proteins; C439524:TNF protein, human; D014409:Tumor Necrosis Factor-alpha