title: SPR-based assays enable the full functional analysis of bispecific molecules.
abstract: The increasing complexity of novel biotherapeutics such as bispecific antibodies or fusion proteins raises new challenges for functional characterization. When compared to standard antibodies, two individual interactions and the inter-dependency of binding events need to be considered for bispecific antibodies. We have previously described an SPR-based assay setup, which enables us to assess the binding activity of a bivalent-bispecific molecule to both targets simultaneously and - in addition to one individual target - in a single setup. However, there might be some pitfalls when applying the bridging assay, e.g. change of antigen activity upon immobilization. Therefore, we have developed an alternative SPR-based assay principle, which allows the individual assessment of both targets in solution. Comparison of data between the assays showed that simultaneous binding can be calculated based on both individual readouts, and revealed a good correlation. Hence, both SPR-based assay principles allow a "full" functional analysis of a bispecific CrossMab in only one assay. The assay principles can be qualified and enable an efficient drug development.
authors.affiliation: Large Molecule Research, Pharma Research and Early Development, Roche Innovation Center Munich, Germany. Electronic address: wolfgang.meschendoerfer@roche.com.
authors.forename: W
authors.initials: W
authors.lastname: Meschendoerfer
authors.affiliation: Large Molecule Research, Pharma Research and Early Development, Roche Innovation Center Munich, Germany.
authors.forename: C
authors.initials: C
authors.lastname: Gassner
authors.affiliation: pRED Informatics, Pharma Research & Early Development, Roche Innovation Center Basel, Switzerland.
authors.forename: F
authors.initials: F
authors.lastname: Lipsmeier
authors.affiliation: Large Molecule Research, Pharma Research and Early Development, Roche Innovation Center Munich, Germany.
authors.forename: J T
authors.initials: JT
authors.lastname: Regula
authors.affiliation: Large Molecule Research, Pharma Research and Early Development, Roche Innovation Center Munich, Germany.
authors.forename: J
authors.initials: J
authors.lastname: Moelleken
mesh_terms: D042702:Angiopoietin-2 / Q000737:chemistry; D018033:Antibodies, Bispecific / Q000737:chemistry; D001681:Biological Assay / Q000379:methods*; D015374:Biosensing Techniques; D015195:Drug Design; D006801:Humans; D007128:Immunoglobulin Fragments / Q000737:chemistry; D008024:Ligands; D016014:Linear Models; D011485:Protein Binding; D012016:Reference Values; D015203:Reproducibility of Results; D020349:Surface Plasmon Resonance / Q000379:methods*; D042461:Vascular Endothelial Growth Factor A / Q000737:chemistry
chemical_list: D042702:Angiopoietin-2; D018033:Antibodies, Bispecific; D007128:Immunoglobulin Fragments; D008024:Ligands; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A
keywords: Activity assessment; Bispecific antibody; Bridging assay; Dual binding assay; Surface plasmon resonance