title: Improving the solubility of anti-LINGO-1 monoclonal antibody Li33 by isotype switching and targeted mutagenesis.
abstract: Monoclonal antibodies (Mabs) are a favorite drug platform of the biopharmaceutical industry. Currently, over 20 Mabs have been approved and several hundred others are in clinical trials. The anti-LINGO-1 Mab Li33 was selected from a large panel of antibodies by Fab phage display technology based on its extraordinary biological activity in promoting oligodendrocyte differentiation and myelination in vitro and in animal models of remyelination. However, the Li33 Fab had poor solubility when converted into a full antibody in an immunoglobulin G1 framework. A detailed analysis of the biochemical and structural features of the antibody revealed several possible reasons for its propensity to aggregate. Here, we successfully applied three molecular approaches (isotype switching, targeted mutagenesis of complementarity determining region residues, and glycosylation site insertion mutagenesis) to address the solubility problem. Through these efforts we were able to improve the solubility of the Li33 Mab from 0.3 mg/mL to >50 mg/mL and reduce aggregation to an acceptable level. These strategies can be readily applied to other proteins with solubility issues.
authors.affiliation: Department of Drug Discovery, Biogen Idec, Inc., Cambridge, Massachusetts 02142, USA. blake.pepinsky@biogenidec.com
authors.forename: R Blake
authors.initials: RB
authors.lastname: Pepinsky
authors.forename: Laura
authors.initials: L
authors.lastname: Silvian
authors.forename: Steven A
authors.initials: SA
authors.lastname: Berkowitz
authors.forename: Graham
authors.initials: G
authors.lastname: Farrington
authors.forename: Alexey
authors.initials: A
authors.lastname: Lugovskoy
authors.forename: Lee
authors.initials: L
authors.lastname: Walus
authors.forename: John
authors.initials: J
authors.lastname: Eldredge
authors.forename: Allan
authors.initials: A
authors.lastname: Capili
authors.forename: Sha
authors.initials: S
authors.lastname: Mi
authors.forename: Christilyn
authors.initials: C
authors.lastname: Graff
authors.forename: Ellen
authors.initials: E
authors.lastname: Garber
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D019540:Area Under Curve; D018360:Crystallography, X-Ray; D004591:Electrophoresis, Polyacrylamide Gel; D017578:Immunoglobulin Class Switching; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007074:Immunoglobulin G / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000379:methods*; D009419:Nerve Tissue Proteins / Q000276:immunology*; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D055550:Protein Stability; D012995:Solubility; D013696:Temperature
chemical_list: D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; C482293:LINGO1 protein, human; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins