title: Adapting pharmacokinetic properties of a humanized anti-interleukin-8 antibody for therapeutic applications using site-specific pegylation.
abstract: A neutralizing anti-interleukin-(IL-)8 monoclonal antibody was humanized by grafting the complementary determining regions onto the human IgG framework. Subsequent alanine scanning mutagenesis and phage display enabled the production of an affinity matured antibody with a >100-fold improvement in IL-8 binding. Antibody fragments can be efficiently produced in Escherichia coli but have the limitation of rapid clearance rates in vivo. The Fab' fragment of the antibody was therefore modified with polyethylene glycol (PEG) in order to obtain a more desirable pharmacokinetic profile. PEG (5-40 kDa) was site-specifically conjugated to the Fab' via the single free cysteine residue in the hinge region. In vitro binding and bioassays showed little or no loss of activity. The pharmacokinetic profiles of the 20 kDa, 30 kDa, 40 kDa, and 40 kDa branched PEG-Fab' molecules were evaluated in rabbits. Relative to the native Fab', the clearance rates of the PEGylated molecules were decreased by 44-175-fold. In a rabbit ear model of ischemia/reperfusion injury, all PEGylated Fab' molecules were as efficacious in reducing oedema as the original monoclonal antibody. These studies demonstrate that it is possible to customize the pharmacokinetic properties of a Fab' while retaining its antigen binding activity.
authors.affiliation: Department of Immunology, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080, USA. steven.leong@maxygen.com
authors.forename: S R
authors.initials: SR
authors.lastname: Leong
authors.forename: L
authors.initials: L
authors.lastname: DeForge
authors.forename: L
authors.initials: L
authors.lastname: Presta
authors.forename: T
authors.initials: T
authors.lastname: Gonzalez
authors.forename: A
authors.initials: A
authors.lastname: Fan
authors.forename: M
authors.initials: M
authors.lastname: Reichert
authors.forename: A
authors.initials: A
authors.lastname: Chuntharapai
authors.forename: K J
authors.initials: KJ
authors.lastname: Kim
authors.forename: D B
authors.initials: DB
authors.lastname: Tumas
authors.forename: W P
authors.initials: WP
authors.lastname: Lee
authors.forename: P
authors.initials: P
authors.lastname: Gribling
authors.forename: B
authors.initials: B
authors.lastname: Snedecor
authors.forename: H
authors.initials: H
authors.lastname: Chen
authors.forename: V
authors.initials: V
authors.lastname: Hsei
authors.forename: M
authors.initials: M
authors.lastname: Schoenhoff
authors.forename: V
authors.initials: V
authors.lastname: Hale
authors.forename: J
authors.initials: J
authors.lastname: Deveney
authors.forename: I
authors.initials: I
authors.lastname: Koumenis
authors.forename: Z
authors.initials: Z
authors.lastname: Shahrokh
authors.forename: P
authors.initials: P
authors.lastname: McKay
authors.forename: W
authors.initials: W
authors.lastname: Galan
authors.forename: B
authors.initials: B
authors.lastname: Wagner
authors.forename: D
authors.initials: D
authors.lastname: Narindray
authors.forename: C
authors.initials: C
authors.lastname: Hébert
authors.forename: G
authors.initials: G
authors.lastname: Zapata
mesh_terms: D000409:Alanine / Q000378:metabolism; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000493:pharmacokinetics*; D000937:Antigen-Antibody Reactions; D018076:DNA, Complementary / Q000378:metabolism; D004487:Edema / Q000628:therapy; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology*; D020128:Inhibitory Concentration 50; D016209:Interleukin-8 / Q000276:immunology* / Q000378:metabolism / Q000493:pharmacokinetics*; D007700:Kinetics; D051379:Mice; D016296:Mutagenesis; D019151:Peptide Library; D011092:Polyethylene Glycols / Q000737:chemistry* / Q000493:pharmacokinetics*; D011485:Protein Binding; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015427:Reperfusion Injury; D013997:Time Factors; D014357:Trypsin / Q000494:pharmacology
chemical_list: D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D007074:Immunoglobulin G; D016209:Interleukin-8; D019151:Peptide Library; D011993:Recombinant Fusion Proteins; D011092:Polyethylene Glycols; D014357:Trypsin; D000409:Alanine