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].
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