Fig. 4: GSK’074 directly binds to RIP1 and RIP3. | Cell Death & Disease

Fig. 4: GSK’074 directly binds to RIP1 and RIP3.

From: Identification of a novel class of RIP1/RIP3 dual inhibitors that impede cell death and inflammation in mouse abdominal aortic aneurysm models

Fig. 4

a, b The kinase domains of recombinant human RIP1 and RIP3 were used in the competitive binding assay of GSK’074. Data were presented as mean ± S.D. n = 2. c, d In vitro kinase activity was performed with recombinant RIP3 (c) or the kinase domain of RIP1 (1–327) (d). Levels of phosphorylation in the presence of compounds indicated were determined by ADP-Glo. Data were presented as mean ± S.D. n = 3. e, f Molecular docking of GSK’074 and kinase domain of human RIP3 (e) or RIP1 (f) in a DFG-out conformation. g L929 cells were primed with IFNβ (50 units/mL) for 24 h, then treated with 10 μg/ml poly(I:C) plus 40 μM zVAD and different concentrations of compounds indicated. Cells were stained with 7-AAD and analyzed by flow cytometry. Data represent mean ± S.D. of three independent experiments. h Mouse primary aortic smooth muscle cells were pretreated with indicated compounds for 2 h, followed by 10 ng/ml TNFα for additional 2 h. Levels of mRNA were determined by Real-time PCR. Data represent mean ± S.D. of three independent experiments. *P < 0.05

Back to article page