Creation of an improved mutant TNF with TNFR1-selectivity and antagonistic activity by phage display technology.
Tumor necrosis factor-alpha (TNF), which binds two types of TNF receptors (TNFR1 and TNFR2), regulates the onset and exacerbation of autoimmune diseases such as rheumatoid arthritis and Crohn's disease. In particular, TNFR1-mediated signals are predominantly related to the induction of inflammatory responses. We have previously generated a TNFR1-selective antagonistic TNF-mutant (mutTNF) and shown that mutTNF efficiently inhibits TNFR1-mediated bioactivity in vitro and attenuates inflammatory conditions in vivo. In this study, we aimed to improve the TNFR1-selectivity of mutTNF This was achieved by constructing a phage library displaying mutTNF-based variants, in which the amino acid residues at the predicted receptor binding sites were substituted to other amino acids. From this mutant TNF library, 20 candidate TNFR1-selective antagonists were isolated. Like mutTNF, all 20 candidates were found to have an inhibitory effect on TNFR1-mediated bioactivity. However, one of the mutants, N7, displayed significantly more than 40-fold greater TNFR1-selectivty than mutTNF. Therefore, N7 could be a promising anti-autoimmune agent that does not interfere with TNFR2-mediated signaling pathways.
Nomura
T
T
National Institute of Biomedical Innovation (NiBio), Osaka University, Osaka, Japan.
Abe
Y
Y
Kamada
H
H
Inoue
M
M
Kawara
T
T
Arita
S
S
Furuya
T
T
Minowa
K
K
Yoshioka
Y
Y
Shibata
H
H
Kayamuro
H
H
Yamashita
T
T
Nagano
K
K
Yoshikawa
T
T
Mukai
Y
Y
Nakagawa
S
S
Tsunoda
S
S
Tsutsumi
Y
Y
D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D005347:Fibroblasts / Q000187:drug effects; D014644:Genetic Variation; D006801:Humans; D009154:Mutation; D019151:Peptide Library; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000037:antagonists & inhibitors* / Q000235:genetics*; D047889:Receptors, Tumor Necrosis Factor, Type II / Q000187:drug effects; D020349:Surface Plasmon Resonance; D048069:Tumor Necrosis Factors / Q000235:genetics* / Q000494:pharmacology*
D019151:Peptide Library; D047888:Receptors, Tumor Necrosis Factor, Type I; D047889:Receptors, Tumor Necrosis Factor, Type II; D048069:Tumor Necrosis Factors