title: Fine tuning of receptor-selectivity for tumor necrosis factor-α using a phage display system with one-step competitive panning.
abstract: Tumor necrosis factor-α (TNF) is one of the attractive targets for the development of anti-inflammatory and anti-tumor drugs, because it is an important mediator in the pathogenesis of several inflammatory diseases and tumor progression. Thus, there is an increasing need to understand the TNF receptor (TNFR1 and TNFR2) biology for the development of TNFR-selective drugs. Nonetheless, the role of TNFRs, especially that of TNFR2, remains poorly understood. Here, using a unique competitive panning, we optimized our phage display-based screening technique for isolating receptor-selective TNF mutants, and identified several TNFR2-specific TNF mutants with high TNFR2 affinity and full bioactivity via TNFR2. Among these mutants, the R2-7 clone revealed very high TNFR2-selectivity (1.8 × 10(5) fold higher than that for the wild-type TNF), which is so far highest among the reported TNFR2-selective TNF mutants. Because of its high TNFR2-selectivity and full bioactivity, the TNF mutant R2-7 would not only help in elucidating the functional role of TNFR2 but would also help in understanding the structure-function relationship of TNF/TNFR2. In summary, our one-step competitive panning system is a simple, useful and effective technology for isolating receptor-selective mutant proteins.
authors.affiliation: Laboratory of Biopharmaceutical Research, National Institute of Biomedical Innovation, 7-6-8 Saito-Asagi, Ibaraki, Osaka, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Abe
authors.forename: Tomoaki
authors.initials: T
authors.lastname: Yoshikawa
authors.forename: Masaki
authors.initials: M
authors.lastname: Inoue
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Nomura
authors.forename: Takeshi
authors.initials: T
authors.lastname: Furuya
authors.forename: Takuya
authors.initials: T
authors.lastname: Yamashita
authors.forename: Kazuya
authors.initials: K
authors.lastname: Nagano
authors.forename: Hiromi
authors.initials: H
authors.lastname: Nabeshi
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Yoshioka
authors.forename: Yohei
authors.initials: Y
authors.lastname: Mukai
authors.forename: Shinsaku
authors.initials: S
authors.lastname: Nakagawa
authors.forename: Haruhiko
authors.initials: H
authors.lastname: Kamada
authors.forename: Yasuo
authors.initials: Y
authors.lastname: Tsutsumi
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Tsunoda
mesh_terms: D017667:Adipocytes / Q000166:cytology / Q000187:drug effects; D019943:Amino Acid Substitution / Q000502:physiology*; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D015374:Biosensing Techniques / Q000379:methods; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry / Q000235:genetics; D007700:Kinetics; D051379:Mice; D019151:Peptide Library; D011485:Protein Binding / Q000502:physiology; D018124:Receptors, Tumor Necrosis Factor / Q000235:genetics / Q000378:metabolism*; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000235:genetics / Q000378:metabolism; D047889:Receptors, Tumor Necrosis Factor, Type II / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D007141:Immunoglobulin Fc Fragments; D019151:Peptide Library; D018124:Receptors, Tumor Necrosis Factor; D047888:Receptors, Tumor Necrosis Factor, Type I; D047889:Receptors, Tumor Necrosis Factor, Type II; D011993:Recombinant Fusion Proteins; D014409:Tumor Necrosis Factor-alpha