title: Production, purification and biological characterization of mono-PEGylated anti-IL-17A antibody fragments.
abstract: The aim of this study was to maximize the yield of the production of mono-PEGylated anti-interleukin-17A (anti-IL-17A) antibody fragments using large (≥ 20 kDa) polyethylene glycol (PEG) chains. Particular attention was paid to selectively yield mono-PEGylated species to maintain the maximum possible functionality and to simplify the purification. Neutralization of IL-17A by antibody constructs might find application for the treatment of bronchial hyperreactivity. Amino-directed and sulfhydryl-directed PEGylation of the native antibody fragments were compared. The former was selected as it produced the most interesting construct in terms of yield and preservation of biological activity. In particular, the F(ab')2-PEG conjugate with one 40 kDa branched PEG prepared in this study was produced at a 42% yield. The conjugate presented only a slight decrease in its binding activity and in its in vitro inhibitory potency offering interesting perspectives for in vivo studies.
authors.affiliation: Pharmaceutics and Drug Delivery Group, Louvain Drug Research Institute, Université catholique de Louvain, Brussels 1200, Belgium.
authors.forename: Salome-Juliette
authors.initials: SJ
authors.lastname: Koussoroplis
authors.forename: Sam
authors.initials: S
authors.lastname: Heywood
authors.forename: Catherine
authors.initials: C
authors.lastname: Uyttenhove
authors.forename: Céline
authors.initials: C
authors.lastname: Barilly
authors.forename: Jacques
authors.initials: J
authors.lastname: Van Snick
authors.forename: Rita
authors.initials: R
authors.lastname: Vanbever
mesh_terms: D000818:Animals; D000918:Antibody Specificity; D001667:Binding, Competitive; D002460:Cell Line; D005347:Fibroblasts / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D006825:Hybridomas; D007128:Immunoglobulin Fragments / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D020381:Interleukin-17 / Q000008:administration & dosage / Q000037:antagonists & inhibitors / Q000276:immunology* / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D051379:Mice; D015394:Molecular Structure; D011092:Polyethylene Glycols / Q000737:chemistry*; D055550:Protein Stability; D013678:Technology, Pharmaceutical / Q000379:methods; D014611:Vaccination
chemical_list: C098680:Il17a protein, mouse; D007128:Immunoglobulin Fragments; D020381:Interleukin-17; D015850:Interleukin-6; D011092:Polyethylene Glycols
keywords: 2-MEA; 2-mercaptoethylamine·HCl; AUC; Anti-interleukin-17A; Antibody fragment; BHR; Bronchial hyperreactivity; CEC; FCS; FPLC; GBS; GCSF; GFC; HSFM; LPS; N-hydroxysuccinimide; NHS; PBS; PEG; PEGylation; PVDF; area under the curve; bronchial hyperreactivity; cation exchange chromatography; fast protein liquid chromatography; fetal calf serum; gel filtration chromatography; glycine buffered saline; granulocyte colony-stimulating factor; hybridoma serum free medium; lipopolysaccharide; mAbs; monoclonal antibodies; phosphate buffered saline; polyethylene glycol; polyvinylidene fluoride