title: Anti-tumor activity of stability-engineered IgG-like bispecific antibodies targeting TRAIL-R2 and LTbetaR.
abstract: Bispecific antibodies (BsAbs) represent an emerging class of biologics that achieve dual targeting with a single agent. Recombinant DNA technologies have facilitated a variety of creative bispecific designs with many promising therapeutic applications; however, practical methods for producing high quality BsAbs that have good product stability, long serum half-life, straightforward purification, and scalable production have largely been limiting. Here we describe a protein-engineering approach for producing stable, scalable tetravalent IgG-like BsAbs. The stability-engineered IgG-like BsAb was envisioned to target and crosslink two TNF family member receptors, TRAIL-R2 (TNF-Related Apoptosis Inducing Ligand Receptor-2) and LTbetaR (Lymphotoxin-beta Receptor), expressed on the surface of epithelial tumor cells with the goal of triggering an enhanced anti-tumor effect. Our IgG-like BsAbs consists of a stability-engineered anti-LTbetaR single chain Fv (scFv) genetically fused to either the N- or C-terminus of the heavy chain of a fulllength anti-TRAIL-R2 IgG1 monoclonal antibody. Both N- or C-terminal BsAbs were active in inhibiting tumor cell growth in vitro, and with some cell lines demonstrated enhanced activity relative to the combination of parental Abs. Pharmacokinetic studies in mice revealed long serum half-lives for the BsAbs. In murine tumor xenograft models, therapeutic treatment with the BsAbs resulted in reduction in tumor volume either comparable to or greater than the combination of parental antibodies, indicating that simultaneously targeting and cross-linking receptor pairs is an effective strategy for treating tumor cells. These studies support that stability-engineering is an enabling step for producing scalable IgG-like BsAbs with properties desirable for biopharmaceutical development.
authors.affiliation: Biogen Idec, Inc, Cambridge, MA 02142, USA. jennifer.michaelson@biogenidec.com
authors.forename: Jennifer S
authors.initials: JS
authors.lastname: Michaelson
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Demarest
authors.forename: Brian
authors.initials: B
authors.lastname: Miller
authors.forename: Aldo
authors.initials: A
authors.lastname: Amatucci
authors.forename: William B
authors.initials: WB
authors.lastname: Snyder
authors.forename: Xiufeng
authors.initials: X
authors.lastname: Wu
authors.forename: Flora
authors.initials: F
authors.lastname: Huang
authors.forename: Samantha
authors.initials: S
authors.lastname: Phan
authors.forename: Sharon
authors.initials: S
authors.lastname: Gao
authors.forename: Adam
authors.initials: A
authors.lastname: Doern
authors.forename: Graham K
authors.initials: GK
authors.lastname: Farrington
authors.forename: Alexey
authors.initials: A
authors.lastname: Lugovskoy
authors.forename: Ingrid
authors.initials: I
authors.lastname: Joseph
authors.forename: Veronique
authors.initials: V
authors.lastname: Bailly
authors.forename: Xin
authors.initials: X
authors.lastname: Wang
authors.forename: Ellen
authors.initials: E
authors.lastname: Garber
authors.forename: Jeff
authors.initials: J
authors.lastname: Browning
authors.forename: Scott M
authors.initials: SM
authors.lastname: Glaser
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D018033:Antibodies, Bispecific / Q000737:chemistry / Q000235:genetics / Q000627:therapeutic use*; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000235:genetics / Q000627:therapeutic use*; D000918:Antibody Specificity; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D006801:Humans; D053259:Lymphotoxin beta Receptor / Q000235:genetics / Q000276:immunology*; D051379:Mice; D016513:Mice, SCID; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000276:immunology / Q000628:therapy*; D015202:Protein Engineering / Q000379:methods*; D055550:Protein Stability; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand / Q000235:genetics / Q000276:immunology*; D057127:Single-Chain Antibodies / Q000737:chemistry; D023041:Xenograft Model Antitumor Assays
chemical_list: D018033:Antibodies, Bispecific; D000911:Antibodies, Monoclonal; D053259:Lymphotoxin beta Receptor; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D057127:Single-Chain Antibodies