title: Functional and structural characterization of four mouse monoclonal antibodies to complement C3 with potential therapeutic and diagnostic applications.
abstract: C3 is the central component of the complement system. Upon activation, C3 sequentially generates various proteolytic fragments, C3a, C3b, iC3b, C3dg, each of them exposing novel surfaces, which are sites of interaction with other proteins. C3 and its fragments are therapeutic targets and markers of complement activation. We report the structural and functional characterization of four monoclonal antibodies (mAbs) generated by immunizing C3-deficient mice with a mixture of human C3b, iC3b and C3dg fragments, and discuss their potential applications. This collection includes three mAbs interacting with native C3 and inhibiting AP complement activation; two of them by blocking the cleavage of C3 by the AP C3-converase and one by impeding formation of the AP C3-convertase. The interaction sites of these mAbs in the target molecules were determined by resolving the structures of Fab fragments bound to C3b and/or iC3b using electron microscopy. A fourth mAb specifically recognizes the iC3b, C3dg, and C3d fragments. It binds to an evolutionary-conserved neoepitope generated after C3b cleavage by FI, detecting iC3b/C3dg deposition over opsonized surfaces by flow cytometry and immunohistochemistry in human and other species. Because well-characterized anti-complement mAbs are uncommon, the mAbs reported here may offer interesting therapeutic and diagnostic opportunities.
authors.affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.|Centro de Investigación Biomédica en Enfermedades Raras, Madrid, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Subías Hidalgo
authors.affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.
authors.forename: Hugo
authors.initials: H
authors.lastname: Yébenes
authors.affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.
authors.forename: César
authors.initials: C
authors.lastname: Rodríguez-Gallego
authors.affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.|Centro de Investigación Biomédica en Enfermedades Raras, Madrid, Spain.
authors.forename: Adrián
authors.initials: A
authors.lastname: Martín-Ambrosio
authors.affiliation: Servicio de Inmunología Microbiana, Centro Nacional de Microbiología, Instituto de Investigación Carlos III Madrid, Spain.
authors.forename: Mercedes
authors.initials: M
authors.lastname: Domínguez
authors.affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.|Centro de Investigación Biomédica en Enfermedades Raras, Madrid, Spain.
authors.forename: Agustin
authors.initials: A
authors.lastname: Tortajada
authors.affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.|Centro de Investigación Biomédica en Enfermedades Raras, Madrid, Spain.
authors.forename: Santiago
authors.initials: S
authors.lastname: Rodríguez de Córdoba
authors.affiliation: Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, Madrid, Spain.
authors.forename: Oscar
authors.initials: O
authors.lastname: Llorca
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000378:metabolism*; D000936:Antigen-Antibody Complex / Q000378:metabolism*; D003176:Complement C3 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D050577:Complement C3-C5 Convertases / Q000378:metabolism*; D003170:Complement Pathway, Alternative; D005818:Genetic Engineering; D006462:Hemolytic Plaque Technique; D006801:Humans; D006825:Hybridomas; D007140:Immunoglobulin Fab Fragments / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D011485:Protein Binding; D011487:Protein Conformation
chemical_list: D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D003176:Complement C3; D007140:Immunoglobulin Fab Fragments; D050577:Complement C3-C5 Convertases
keywords: C3; C3b; C3bBb convertase; Complement inhibition; Monoclonal antibody