title: Structural basis for the dual recognition of IL-12 and IL-23 by ustekinumab.
abstract: Interleukin (IL)-12 and IL-23 are heterodimeric proinflammatory cytokines that share a common p40 subunit, paired with p35 and p19 subunits, respectively. They represent an attractive class of therapeutic targets for the treatment of psoriasis and other immune-mediated diseases. Ustekinumab is a fully human monoclonal antibody (mAb) that binds specifically to IL-12/IL-23p40 and neutralizes human IL-12 and IL-23 bioactivity. The crystal structure of ustekinumab Fab (antigen binding fragment of mAb), in complex with human IL-12, has been determined by X-ray crystallography at 3.0 Å resolution. Ustekinumab Fab binds the D1 domain of the p40 subunit in a 1:1 ratio in the crystal, consistent with a 2 cytokines:1 mAb stoichiometry, as measured by isothermal titration calorimetry. The structure indicates that ustekinumab binds to the same epitope on p40 in both IL-12 and IL-23 with identical interactions. Mutational analyses confirm that several residues identified in the IL-12/IL-23p40 epitope provide important molecular binding interactions with ustekinumab. The electrostatic complementarity between the mAb antigen binding site and the p40 D1 domain epitope appears to play a key role in antibody/antigen recognition specificity. Interestingly, this structure also reveals significant structural differences in the p35 subunit and p35/p40 interface, compared with the published crystal structure of human IL-12, suggesting unusual and potentially functionally relevant structural flexibility of p35, as well as p40/p35 recognition. Collectively, these data describe unique observations about IL-12p35 and ustekinumab interactions with p40 that account for its dual binding and neutralization of IL-12 and IL-23.
authors.affiliation: Centocor Research and Development, Inc., 145 King of Prussia Road, Radnor, PA 19087, USA. jluo@its.jnj.com
authors.forename: Jinquan
authors.initials: J
authors.lastname: Luo
authors.forename: Sheng-Jiun
authors.initials: SJ
authors.lastname: Wu
authors.forename: Eilyn R
authors.initials: ER
authors.lastname: Lacy
authors.forename: Yevgeniya
authors.initials: Y
authors.lastname: Orlovsky
authors.forename: Audrey
authors.initials: A
authors.lastname: Baker
authors.forename: Alexey
authors.initials: A
authors.lastname: Teplyakov
authors.forename: Galina
authors.initials: G
authors.lastname: Obmolova
authors.forename: George A
authors.initials: GA
authors.lastname: Heavner
authors.forename: Hans-Thomas
authors.initials: HT
authors.lastname: Richter
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Benson
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000378:metabolism*; D061067:Antibodies, Monoclonal, Humanized; D002151:Calorimetry; D018360:Crystallography, X-Ray; D000939:Epitopes / Q000235:genetics / Q000276:immunology; D006801:Humans; D007155:Immunologic Factors / Q000737:chemistry* / Q000378:metabolism*; D018664:Interleukin-12 / Q000235:genetics / Q000378:metabolism*; D053759:Interleukin-23 / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D000069549:Ustekinumab
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000939:Epitopes; D007155:Immunologic Factors; D053759:Interleukin-23; D018664:Interleukin-12; D000069549:Ustekinumab