title: Efficient method to optimize antibodies using avian leukosis virus display and eukaryotic cells.
abstract: Antibody-based therapeutics have now had success in the clinic. The affinity and specificity of the antibody for the target ligand determines the specificity of therapeutic delivery and off-target side effects. The discovery and optimization of high-affinity antibodies to important therapeutic targets could be significantly improved by the availability of a robust, eukaryotic display technology comparable to phage display that would overcome the protein translation limitations of microorganisms. The use of eukaryotic cells would improve the diversity of the displayed antibodies that can be screened and optimized as well as more seamlessly transition into a large-scale mammalian expression system for clinical production. In this study, we demonstrate that the replication and polypeptide display characteristics of a eukaryotic retrovirus, avian leukosis virus (ALV), offers a robust, eukaryotic version of bacteriophage display. The binding affinity of a model single-chain Fv antibody was optimized by using ALV display, improving affinity >2,000-fold, from micromolar to picomolar levels. We believe ALV display provides an extension to antibody display on microorganisms and offers virus and cell display platforms in a eukaryotic expression system. ALV display should enable an improvement in the diversity of properly processed and functional antibody variants that can be screened and affinity-optimized to improve promising antibody candidates.
authors.affiliation: Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905.
authors.forename: Changming
authors.initials: C
authors.lastname: Yu
authors.affiliation: Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905.
authors.forename: Gennett M
authors.initials: GM
authors.lastname: Pike
authors.affiliation: Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905.
authors.forename: Tommy A
authors.initials: TA
authors.lastname: Rinkoski
authors.affiliation: Division of Oncology Research, Mayo Clinic, Rochester, MN 55905.
authors.forename: Cristina
authors.initials: C
authors.lastname: Correia
authors.affiliation: Division of Oncology Research, Mayo Clinic, Rochester, MN 55905.
authors.forename: Scott H
authors.initials: SH
authors.lastname: Kaufmann
authors.affiliation: Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905 federspiel.mark@mayo.edu.
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Federspiel
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000378:metabolism*; D001354:Avian Leukosis Virus / Q000378:metabolism*; D002460:Cell Line; D061505:Cell Surface Display Techniques / Q000379:methods*; D002645:Chickens; D022801:Complementarity Determining Regions; D005057:Eukaryotic Cells / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D007797:Laminin / Q000378:metabolism; D008969:Molecular Sequence Data; D016296:Mutagenesis; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D057127:Single-Chain Antibodies / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000378:metabolism; D014771:Virion / Q000378:metabolism; D014779:Virus Replication
chemical_list: D000906:Antibodies; D022801:Complementarity Determining Regions; D006023:Glycoproteins; D007797:Laminin; D050505:Mutant Proteins; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; D014759:Viral Envelope Proteins
keywords: antibody binding affinity; antibody engineering; avian leukosis virus; avian leukosis virus polypeptide display; protein display technology