SPL-707是有效的，选择性的和口服可用的信号肽肽酶样2a(SPPL2a)抑制剂，IC50值为80nM。
SPL-707 is a potent, selective and orally available signal peptide peptidase-like 2a (SPPL2a) inhibitor with an IC50 of 80 nM.
SPL-707 shows 25-fold selectivity over SPP with an IC50 of 3.7 μM and some preference for SPPL2a over SPPL2b with a 3-fold selectivity, comparing IC50 values generated with comparable assay formats for the different proteases[1].
SPL-707 significantly inhibits processing of the SPPL2a substrate CD74/p8 fragment in rodents at doses ≤10 mg/kg b.i.d. po. Oral dosing of SPL-707 for 11 days at ≥10 mg/kg b.i.d. recapitulates the phenotype seen in Sppl2a knockout (ko) and ENU mutant mice (reduced number of specific B cells and myeloid dendritic cells)[1].
[1]. Velcicky J, et al. Discovery of the First Potent, Selective, and Orally Bioavailable Signal Peptide Peptidase-Like 2a (SPPL2a) Inhibitor Displaying Pronounced Immunomodulatory Effects In Vivo. J Med Chem. 2018 Feb 8;61(3):865-880.
Cell experiment: | DNA vectors encoding human Notch1-VP16-Gal4 fusion protein and a Gal4-luciferase reporter for the γ-secretase RGA, or human SPPL2a, VP16-TNFa(aa1-76)-NTF substrate, and the Gal4-luciferase reporter plasmid for the SPPL2a RGA, are transiently transfected in HEK293 cells using FuGENE. After transfection, the cell suspensions are diluted and distributed to white solid 384-well plate at 10000 cells/50 μL/well. After 3 h, 200 nL of SPL-707 in DMSO is stamped into the wells in concentration response covering final inhibitor concentrations of 10 μM to 0.3 nM in triplicate. Subsequently, plates are incubated for 24 h at 37°C, 5% CO2 in a humidified incubator before the addition of 30 μL Bright Glo. After incubation for 5 min at rt, luminescence is measured and IC50 values are determined by plotting compound concentration vs normalized luminescence values[1]. |
Animal experiment: | Rats[1]Female Lewis rats receive SPL-707 (1, 3, 10, and 30 mg/kg b.i.d.; first dose at 0 h, second dose at 8 h). Then 16 h after the second dose (24 h in total), inhibition of SPPL2a is assessed by measuring CD74/p8 accumulation in blood and splenocyte extracts by Western blot analysis[1].Mice[1] Mice receive single oral doses of 150 mg/ kg 31 or 1, 3, 10, and 30 mg/kg 40. Then 16 h after dosing, inhibition of SPPL2a is assessed by measuring CD74/p8 accumulation in splenocyte lysates by Western blot analysis[1]. |
参考文献: [1]. Velcicky J, et al. Discovery of the First Potent, Selective, and Orally Bioavailable Signal Peptide Peptidase-Like 2a (SPPL2a) Inhibitor Displaying Pronounced Immunomodulatory Effects In Vivo. J Med Chem. 2018 Feb 8;61(3):865-880. | |
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