title: Development of tetravalent IgG1 dual targeting IGF-1R-EGFR antibodies with potent tumor inhibition.
abstract: In this study we present novel bispecific antibodies that simultaneously target the insulin-like growth factor receptor type I (IGF-1R) and epidermal growth factor receptor (EGFR). For this purpose disulfide stabilized scFv domains of the EGFR/ADCC antibody GA201 were fused via serine-glycine connectors to the C-terminus of the heavy (XGFR2) or light chain (XGFR4), or the N-termini of the light (XGFR5) or heavy chain (XGFR3) of the IGF-1R antibody R1507 as parental IgG1 antibody. The resulting bispecific IGF-1R-EGFR antibodies XGFR2, XGFR3 and XGFR4 were successfully generated with yields and stability comparable to conventional IgG1 antibodies. They effectively inhibited IGF-1R and EGFR phosphorylation and 3D proliferation of H322M and H460M2 tumor cells, induced strong down-modulation of IGF-1R as well as enhanced EGFR down-modulation compared to the parental EGFR antibody GA201 and were ADCC competent. The bispecific XGFR derivatives showed a strong format dependent influence of N- or C-terminal heavy and light chain scFv attachment on ADCC activity and an increase in receptor downregulation over the parental combination in vitro. XGFR2 and XGFR4 were selected for in vivo evaluation and showed potent anti-tumoral efficacy comparable to the combination of monospecific IGF-1R and EGFR antibodies in subcutaneous BxPC3 and H322M xenograft models. In summary, we have managed to overcome issues of stability and productivity of bispecific antibodies, discovered important antibody fusion protein design related differences on ADCC activity and receptor downmodulation and show that IGF-1R-EGFR antibodies represent an attractive therapeutic strategy to simultaneously target two key components de-regulated in multiple cancer types, with the ultimate goal to avoid the formation of resistance to therapy.
authors.affiliation: Large Molecule Research, Roche Pharma Research and Early Development (pRED), Roche Diagnostics GmbH, D-82372 Penzberg, Germany.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Croasdale
authors.forename: Katharina
authors.initials: K
authors.lastname: Wartha
authors.forename: Juergen M
authors.initials: JM
authors.lastname: Schanzer
authors.forename: Klaus-Peter
authors.initials: KP
authors.lastname: Kuenkele
authors.forename: Carola
authors.initials: C
authors.lastname: Ries
authors.forename: Klaus
authors.initials: K
authors.lastname: Mayer
authors.forename: Christian
authors.initials: C
authors.lastname: Gassner
authors.forename: Martina
authors.initials: M
authors.lastname: Wagner
authors.forename: Nikolaos
authors.initials: N
authors.lastname: Dimoudis
authors.forename: Sylvia
authors.initials: S
authors.lastname: Herter
authors.forename: Christiane
authors.initials: C
authors.lastname: Jaeger
authors.forename: Claudia
authors.initials: C
authors.lastname: Ferrara
authors.forename: Eike
authors.initials: E
authors.lastname: Hoffmann
authors.forename: Lothar
authors.initials: L
authors.lastname: Kling
authors.forename: Wilma
authors.initials: W
authors.lastname: Lau
authors.forename: Roland F
authors.initials: RF
authors.lastname: Staack
authors.forename: Julia
authors.initials: J
authors.lastname: Heinrich
authors.forename: Werner
authors.initials: W
authors.lastname: Scheuer
authors.forename: Jan
authors.initials: J
authors.lastname: Stracke
authors.forename: Christian
authors.initials: C
authors.lastname: Gerdes
authors.forename: Ulrich
authors.initials: U
authors.lastname: Brinkmann
authors.forename: Pablo
authors.initials: P
authors.lastname: Umana
authors.forename: Christian
authors.initials: C
authors.lastname: Klein
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use*; D000915:Antibody Affinity; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D003001:Cloning, Molecular; D066246:ErbB Receptors / Q000276:immunology* / Q000378:metabolism; D005260:Female; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics / Q000276:immunology* / Q000627:therapeutic use*; D007167:Immunotherapy; D051379:Mice; D016513:Mice, SCID; D008958:Models, Molecular; D009369:Neoplasms / Q000276:immunology / Q000378:metabolism / Q000628:therapy*; D010766:Phosphorylation / Q000187:drug effects; D015202:Protein Engineering; D017526:Receptor, IGF Type 1 / Q000276:immunology* / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000627:therapeutic use; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology / Q000627:therapeutic use
chemical_list: D018033:Antibodies, Bispecific; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D057127:Single-Chain Antibodies; D066246:ErbB Receptors; D017526:Receptor, IGF Type 1