Necroptosis is a type of programmed cell death that usually occurs under apoptosis-deficient conditions. Receptor-interacting protein kinase-3 (RIP3) is a central player in necroptosis, and its kinase activity is essential for downstream necroptotic signaling events. HS-1371 is a potent and ATP-competitive RIP3 inhibitor with an IC50 value of 20.8 nM. HS-1371 showed an inhibitory effect on S227 auto-phosphorylation of RIP3 at the basal level in a dose-dependent manner, indicating that HS-1371 can be used as potential preventive agent in RIP3-mediated necroptotic cell death. HT-29 cells were treated with TRAIL (stimuli leading to necroptosis) plus Smac mimetic and zVAD to induce TRAIL-mediated necroptosis; HS-1371 decreased TRAIL-induced S227 phosphorylation of RIP3 and RIP3-mediated phosphorylation of MLKL in a dose-dependent manner indicating that RIP3-mediated necroptosis could be inhibited by HS-1371. In HeLa cells that lack endogenous RIP3 expression, the ectopic expression of RIP3 resulted in basal activation of RIP3 phosphorylation; this activity was decreased by HS-1371 in a dose-dependent manner. RIP3 phosphorylation was markedly increased by TSZ treatment, but pretreatment of HS-1371 effectively blocked RIP3 phosphorylation and MLKL phosphorylation. TNF-mediated necroptosis was induced and then HS-1371 was applied 1 or 2 h later. Low concentrations of HS-1371 showed a partial inhibitory effect on RIP3 phosphorylation in both pre-treatment and post-treatment, but 5 μM HS-1371 completely inhibited RIP3 phosphorylation and TNF-induced cell death[1].
作用机制
The ability of HS-1371 to establish hydrogen bonds with the backbone groups in the hinge region of RIP3 is necessary for its tight binding in the ATP-binding site of RIP3[1].
HS-1371 细胞实验
Cell Line
Concentration
Treated Time
Description
References
MEF cells
5 μM
To test the inhibitory effect of HS-1371 on TNF plus CHX and zVAD-induced necroptosis in mouse embryonic fibroblast MEF cells. Results showed that HS-1371 effectively inhibited MLKL phosphorylation and cell death.
Exp Mol Med. 2018 Sep 20;50(9):1-15
L929 cells
5 μM
To test the inhibitory effect of HS-1371 on TNF plus zVAD-induced necroptosis in mouse fibrosarcoma L929 cells. Results showed that HS-1371 effectively inhibited necroptosis.
Exp Mol Med. 2018 Sep 20;50(9):1-15
HT-29 cells
10 μM
9 hours
To test the inhibitory effect of HS-1371 on RIP3 S227 auto-phosphorylation. Results showed that HS-1371 effectively inhibited RIP3 S227 auto-phosphorylation.
Exp Mol Med. 2018 Sep 20;50(9):1-15
H2009 cells
0-10 μM
6 hours
To test the inhibitory effect of HS-1371 on TSZ-induced necroptosis in H2009 cells with ectopic RIP3 expression. Results showed that HS-1371 effectively inhibited TSZ-induced necroptosis.
Exp Mol Med. 2018 Sep 20;50(9):1-15
HeLa cells
0-10 μM
6 hours
To test the inhibitory effect of HS-1371 on RIP3 phosphorylation in HeLa cells with ectopic RIP3 expression. Results showed that HS-1371 dose-dependently inhibited RIP3 phosphorylation.
Exp Mol Med. 2018 Sep 20;50(9):1-15
mouse intestinal organoids (MI organoids)
10 nM
48 hours
To evaluate the inhibitory effect of HS-1371 on toceranib-induced necrotic cell death. Results showed that HS-1371 significantly reduced the cell death rate in MI organoids and suppressed the expression of necrosis-related genes RIPK3 and Mlkl.
Sci Rep. 2025 Jan 2;15(1):39
HS-1371 动物实验
Species
Animal Model
Administration
Dosage
Frequency
Description
References
Mice
B16 melanoma model
In vitro treatment
2 μM
Single treatment
Evaluate the effect of HS-1371 on TNFα/IFNγ-induced cell death
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