title: Cross-species analysis of Fc engineered anti-Lewis-Y human IgG1 variants in human neonatal receptor transgenic mice reveal importance of S254 and Y436 in binding human neonatal Fc receptor.
abstract: IgG has a long half-life through engagement of its Fc region with the neonatal Fc receptor (FcRn). The FcRn binding site on IgG1 has been shown to contain I253 and H310 in the CH2 domain and H435 in the CH3 domain. Altering the half-life of IgG has been pursued with the aim to prolong or reduce the half-life of therapeutic IgGs. More recent studies have shown that IgGs bind differently to mouse and human FcRn. In this study we characterize a set of hu3S193 IgG1 variants with mutations in the FcRn binding site. A double mutation in the binding site is necessary to abrogate binding to murine FcRn, whereas a single mutation in the FcRn binding site is sufficient to no longer detect binding to human FcRn and create hu3S193 IgG1 variants with a half-life similar to previously studied hu3S193 F(ab')2 (t1/2β, I253A, 12.23 h; H310A, 12.94; H435A, 12.57; F(ab')2, 12.6 h). Alanine substitutions in S254 in the CH2 domain and Y436 in the CH3 domain showed reduced binding in vitro to human FcRn and reduced elimination half-lives in huFcRn transgenic mice (t1/2β, S254A, 37.43 h; Y436A, 39.53 h; wild-type, 83.15 h). These variants had minimal effect on half-life in BALB/c nu/nu mice (t1/2β, S254A, 119.9 h; Y436A, 162.1 h; wild-type, 163.1 h). These results provide insight into the interaction of human Fc by human FcRn, and are important for antibody-based therapeutics with optimal pharmacokinetics for payload strategies used in the clinic.
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.|b School of Cancer Medicine, La Trobe University , Melbourne , VIC , Australia.
authors.forename: Ingrid J G
authors.initials: IJ
authors.lastname: Burvenich
authors.affiliation: c Centre for Biomedical Research, Burnet Institute , Melbourne , VIC , Australia.
authors.forename: William
authors.initials: W
authors.lastname: Farrugia
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.
authors.forename: Fook T
authors.initials: FT
authors.lastname: Lee
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.
authors.forename: Bruno
authors.initials: B
authors.lastname: Catimel
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.
authors.forename: Zhanqi
authors.initials: Z
authors.lastname: Liu
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.
authors.forename: Dahna
authors.initials: D
authors.lastname: Makris
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.
authors.forename: Diana
authors.initials: D
authors.lastname: Cao
authors.affiliation: b School of Cancer Medicine, La Trobe University , Melbourne , VIC , Australia.|d Department of Molecular Imaging and Therapy, Austin Health , Melbourne , Australia.
authors.forename: Graeme J
authors.initials: GJ
authors.lastname: O'Keefe
authors.affiliation: e Radioimmune Inorganic Chemistry Section, Radiation Oncology Branch, Center for Cancer Research, National Cancer Institute , Bethesda , MD , USA.
authors.forename: Martin W
authors.initials: MW
authors.lastname: Brechbiel
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.
authors.forename: Dylan
authors.initials: D
authors.lastname: King
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.
authors.forename: Violeta
authors.initials: V
authors.lastname: Spirkoska
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.
authors.forename: Laura C
authors.initials: LC
authors.lastname: Allan
authors.affiliation: c Centre for Biomedical Research, Burnet Institute , Melbourne , VIC , Australia.|f School of Science, RMIT University , Bundoora , VIC , Australia.|g Department of Immunology , Monash University , Melbourne , VIC , Australia.|h Department of Surgery Austin Health , University of Melbourne , Heidelberg , VIC , Australia.
authors.forename: Paul A
authors.initials: PA
authors.lastname: Ramsland
authors.affiliation: a Tumour Targeting Laboratory, Ludwig Institute for Cancer Research and Olivia Newton-John Cancer Research Institute , Melbourne , VIC , Australia.|b School of Cancer Medicine, La Trobe University , Melbourne , VIC , Australia.|d Department of Molecular Imaging and Therapy, Austin Health , Melbourne , Australia.|i Faculty of Medicine, University of Melbourne , Melbourne , VIC , Australia.
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Scott
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology / Q000494:pharmacology; D006207:Half-Life; D015395:Histocompatibility Antigens Class I / Q000276:immunology*; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry* / Q000276:immunology / Q000494:pharmacology; D007983:Lewis Blood Group Antigens / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic; D015202:Protein Engineering; D055550:Protein Stability; D011961:Receptors, Fc / Q000737:chemistry* / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D015395:Histocompatibility Antigens Class I; D007074:Immunoglobulin G; D007983:Lewis Blood Group Antigens; C094162:Lewis Y antigen; D011961:Receptors, Fc; C100688:Fc receptor, neonatal
keywords: Antibody engineering; Fc receptors; molecular biology; neonatal Fc receptor; transgenic mice