After two rounds of design, we obtained 4e with an IC50 of 3

After two rounds of design, we obtained 4e with an IC50 of 3.0 0.8 M, which is one of the most potent TNF- small molecule inhibitors reported so far. TNF- inhibitor, EJMC-1 with modest activity. Here, we optimized this compound by shape screen and rational design. In the first round, we screened commercial compound library for EJMC-1 analogs based on shape similarity. Out of the 68 compounds tested, 20 compounds showed better binding affinity than EJMC-1 in the SPR competitive binding assay. These 20 compounds were tested in cell assay and the most potent compound was 2-oxo-N-phenyl-1,2-dihydrobenzo[designed small protein DS119, and then optimized their residues at the interface, which provided some small proteins that bind TNF- with sub-micromolar affinities (Zhu et al., 2016). Other than small proteins, bicyclic peptides and helical peptides were also designed as peptidic antagonists of TNF- (Lian et al., 2013; Zhang et al., 2013). In addition to peptide inhibitors, small molecular inhibitors that directly targeting TNF- have also been discovered (Leung et al., 2012; Davis and Colangelo, 2013; Shen et al., 2014). Suramin was thought to be the first small compound inhibitor that directly disrupts the interactions between TNF- and its receptor (TNFR) (Grazioli et al., 1992). But its potency was too low to be used in clinic (Alzani et al., 1993). No breakthrough was made until 2005, when SPD304 was reported as the first potent small molecule inhibitor that directly targeting TNF-, with an IC50 of 22 M by ELISA. And the co-crystal structure of SPD304 in complex with TNF- dimer was solved (He et al., 2005). However, as the 3-alkylindole moiety of SPD304 can be metabolized by cytochrome P450s to produce toxic electrophilic intermediates, its further applications is limited (Sun and Yost, 2008). After that, several novel TNF- inhibitors were discovered using structure-based virtual screening (VS) of different chemical libraries. Chan et al. identified two compounds using high-throughput ligand-docking-based VS (Figure ?(Figure1,1, quinuclidine 1 and indoloquinolizidine 2), and their experimental PF-04937319 tests showed that quinuclidine 1 is more effective than indoloquinolizidine 2 in inhibition of TNF- induced NF-B signaling in HepG2 cells, with IC50-values of 5 and 30 M, respectively (Chan et al., 2010). Choi and colleagues discovered a series of pyrimidine-2,4,6-trione derivatives from a 240,000-compound library. The best compound (Figure ?(Figure1,1, Oxole-1) showed 64% inhibition at 10 M (Choi et al., 2010). Leung et al. reported a novel iridium(III)-based direct inhibitor of TNF- (Figure ?(Figure1,1, [Ir(ppy)2(biq)]PF6; Leung et al., 2012). Mouhsine et al. used combined screening approaches to identify orally available TNF- inhibitors with IC50 of 10 M (Figure ?(Figure1,1, Benzenesulfonamide-1; PF-04937319 Mouhsine et al., 2017). Other efforts to develop TNF- inhibitors were also reported (Mancini et al., 1999; Buller et al., 2009; Leung et al., 2011; Hu et al., 2012; Alexiou et al., 2014; Ma et al., 2014; Kang et al., 2016). However, due to the low potency and high cytotoxicity, small molecule TNF- inhibitors still have a long way to go for clinical applications (Davis and Colangelo, 2013). Highly active TNF- inhibitors with novel chemical structures need to be developed. In a previous study, we have discovered a compound (Figure ?(Figure1,1, EJMC-1) that directly bound TNF- (Shen et al., 2014). The scaffold of the compound, 2-oxo-N-phenyl-1,2-dihydrobenzo[= 6.7 Hz), 8.01 (d, 1H, = 8.3 Hz), 7.75C7.70 (m, 1H), 7.53 (d, 1H, = 8.3 Hz), 7.40 (dd, 1H, = 7.5, 6.7 Hz), 6.94 (d, 1H, = 6.7 Hz). 2-oxo-1,2-dihydrobenzo[= 7.5 Hz, 1H), 6.88 (t, = 8.1 Hz, 1H), 6.92 (d, = 7.6 Hz, 1H), 7.04 (d, = 8.4 Hz, 1H), 7.11 (d, = 7.3 Hz, 1H), 7.18 (t, = 7.9 Hz, 1H), 7.87 (dd, = 7.9, 4.0 Hz, 3H), 8.07 (d, = 7.0 Hz, 1H), 8.65 (d, = 8.4 Hz, 1H), 10.18 (s, 1H), 11.07 (s, 1H). 13C NMR (101 MHz, DMSO-= 7.6 Hz, 1H), 7.05 (ddd, = 8.1, 6.7, 1.2 Hz, 1H), 7.22 (ddd, = 8.2, 6.8, 1.3 Hz, 1H), 7.35 (s, 1H), 7.41 (d, = 8.2 Hz, 1H), 7.46 (d, = 8.2 Hz, 1H), 7.85 (dd, = 8.4, 7.0 Hz, 1H), 7.95 (d, = 7.6 Hz, 1H), 8.08 (d, = 7.0 Hz, 1H), 8.65 (d, = 8.4 Hz, 1H), 11.12 (s, 1H). 13C NMR (101 MHz, DMSO-= 9.7, 5.3 Hz, 1H),.These differences might due to the flexibility of SPD304, which adopted a U shape conformation, and the conformational sampling preference of the docking software. residues at the interface, which provided some small proteins that PF-04937319 bind TNF- with sub-micromolar affinities (Zhu et al., 2016). Other than small proteins, bicyclic peptides and helical peptides were also designed as peptidic antagonists of TNF- (Lian et al., 2013; Zhang et al., 2013). In addition to peptide inhibitors, small molecular inhibitors that directly targeting TNF- have also been discovered (Leung et al., 2012; Davis and Colangelo, 2013; Shen et al., 2014). Suramin was thought to be the first small compound inhibitor that directly disrupts the interactions between TNF- and its receptor (TNFR) (Grazioli et al., 1992). But its potency was too low to be used in clinic (Alzani et al., 1993). No breakthrough was made until 2005, when SPD304 was reported as the first potent small molecule inhibitor that directly targeting TNF-, with an IC50 of 22 M by ELISA. And the co-crystal structure of SPD304 in complex with TNF- dimer was solved (He et al., 2005). However, as the 3-alkylindole moiety of SPD304 can be metabolized by cytochrome P450s to produce toxic electrophilic intermediates, its further applications is limited (Sun and Yost, 2008). After that, several novel TNF- inhibitors were discovered using structure-based virtual screening (VS) of different chemical libraries. Chan et al. identified two compounds using high-throughput ligand-docking-based VS (Figure ?(Figure1,1, quinuclidine 1 and indoloquinolizidine 2), and their experimental tests showed that quinuclidine 1 is more effective than indoloquinolizidine 2 in inhibition of TNF- induced NF-B signaling in HepG2 cells, with IC50-values of 5 and 30 M, respectively (Chan et al., 2010). Choi and colleagues discovered a series of pyrimidine-2,4,6-trione derivatives from a 240,000-compound library. The best compound (Figure ?(Figure1,1, Oxole-1) showed 64% inhibition PF-04937319 at 10 M (Choi et al., 2010). Leung et al. reported a novel iridium(III)-based direct inhibitor of TNF- (Figure ?(Figure1,1, [Ir(ppy)2(biq)]PF6; Leung et al., 2012). Mouhsine et al. used combined screening approaches to identify orally available TNF- inhibitors with IC50 of 10 M (Figure ?(Figure1,1, Benzenesulfonamide-1; Mouhsine et al., 2017). Other efforts to develop TNF- inhibitors were also reported (Mancini et al., 1999; Buller et al., 2009; Leung et al., 2011; Hu et al., 2012; Alexiou et al., 2014; Ma et al., 2014; Kang et al., 2016). However, due to the low potency and high cytotoxicity, small molecule TNF- inhibitors still have a long way to go for clinical applications (Davis and Colangelo, 2013). Highly active TNF- inhibitors with novel chemical structures need to be developed. In a previous study, we have discovered a compound (Figure ?(Figure1,1, EJMC-1) that directly bound TNF- (Shen et al., 2014). The scaffold of the compound, 2-oxo-N-phenyl-1,2-dihydrobenzo[= 6.7 Hz), 8.01 (d, 1H, = 8.3 Hz), 7.75C7.70 (m, 1H), 7.53 (d, 1H, = PF-04937319 8.3 Hz), 7.40 (dd, 1H, = 7.5, 6.7 Hz), 6.94 (d, 1H, = 6.7 Hz). 2-oxo-1,2-dihydrobenzo[= 7.5 Hz, 1H), 6.88 (t, = 8.1 Hz, 1H), 6.92 (d, = 7.6 Hz, 1H), 7.04 (d, = 8.4 Hz, 1H), 7.11 (d, = 7.3 Hz, 1H), 7.18 (t, = 7.9 Hz, Mouse monoclonal to MBP Tag 1H), 7.87 (dd, = 7.9, 4.0 Hz, 3H), 8.07 (d, = 7.0 Hz, 1H), 8.65 (d, = 8.4 Hz, 1H), 10.18 (s, 1H), 11.07 (s, 1H). 13C NMR (101 MHz, DMSO-= 7.6 Hz, 1H), 7.05 (ddd, = 8.1, 6.7, 1.2 Hz, 1H), 7.22 (ddd, = 8.2, 6.8, 1.3 Hz, 1H), 7.35 (s, 1H), 7.41 (d, = 8.2 Hz, 1H), 7.46 (d, = 8.2 Hz, 1H), 7.85 (dd, = 8.4, 7.0 Hz, 1H), 7.95 (d, =.