Synthesis and biological evaluation of celastrol derivatives as potential anti-glioma agents by activating RIP1/RIP3/MLKL pathway to induce necroptosis.

Celastrol, a quinone methide triterpenoid, possesses potential anti-glioma activity. However, its relatively low activity limit its application as an effective agent for glioma treatment. In search for effective anti-glioma agents, this work designed and synthesized two series of celastrol C-3 OH and C-20 COOH derivatives 4a-4o and 6a-6o containing 1, 2, 3-triazole moiety. Their anti-glioma activities against four human glioma cell lines (A172, LN229, U87, and U251) were then evaluated using MTT assay in vitro. Results showed that compound 6i (IC50 = 0.94 μM) exhibited substantial antiproliferative activity against U251 cell line, that was 4.7-fold more potent than that of celastrol (IC50 = 4.43 μM). In addition, compound 6i remarkably inhibited the colony formation and migration of U251 cells. Further transmission electron microscopy and mitochondrial depolarization assays in U251 cells indicated that the potent anti-glioma activity of 6i was attributed to necroptosis. Mechanism investigation revealed that compound 6i induced necroptosis mainly by activating the RIP1/RIP3/MLKL pathway. Additionally, compound 6i exerted acceptable BBB permeability in mice and inhibited U251 cell proliferation in an in vivo zebrafish xenograft model, obviously. In summary, compound 6i might be a promising lead compound for potent celastrol derivatives as anti-glioma agents.

Feng Y ，Wang W ，Zhang Y ，Fu X ，Ping K ，Zhao J ，Lei Y ，Mou Y ，Wang S ... -  《-》

Emodin induced necroptosis in the glioma cell line U251 via the TNF-α/RIP1/RIP3 pathway.

Zhou J ，Li G ，Han G ，Feng S ，Liu Y ，Chen J ，Liu C ，Zhao L ，Jin F ... -  《-》

Shikonin induces glioma cell necroptosis in vitro by ROS overproduction and promoting RIP1/RIP3 necrosome formation.

Lu B ，Gong X ，Wang ZQ ，Ding Y ，Wang C ，Luo TF ，Piao MH ，Meng FK ，Chi GF ，Luo YN ，Ge PF ... -  《-》

RIP1, RIP3, and MLKL Contribute to Cell Death Caused by Clostridium perfringens Enterotoxin.

Clostridium perfringens type F strains cause gastrointestinal disease when they produce a pore-forming toxin named C. perfringens enterotoxin (CPE). In human enterocyte-like Caco-2 cells, low CPE concentrations cause caspase-3-dependent apoptosis, while high CPE concentrations cause necrosis. Since necrosis or apoptosis sometimes involves receptor-interacting serine/threonine-protein kinase-1 or 3 (RIP1 or RIP3), this study examined whether those kinases are important for CPE-induced apoptosis or necrosis. Highly specific RIP1 or RIP3 inhibitors reduced both CPE-induced apoptosis and necrosis in Caco-2 cells. Those findings suggested that the form of necrosis induced by treating Caco-2 cells with high CPE concentrations involves necroptosis, which was confirmed when high, but not low, CPE concentrations were shown to induce oligomerization of mixed-lineage kinase domain-like pseudokinase (MLKL), a key late step in necroptosis. Furthermore, an MLKL oligomerization inhibitor reduced cell death caused by high, but not low, CPE concentrations. Supporting RIP1 and RIP3 involvement in CPE-induced necroptosis, inhibitors of those kinases also reduced MLKL oligomerization during treatment with high CPE concentrations. Calpain inhibitors similarly blocked MLKL oligomerization induced by high CPE concentrations, implicating calpain activation as a key intermediate in initiating CPE-induced necroptosis. In two other CPE-sensitive cell lines, i.e., Vero cells and human enterocyte-like T84 cells, low CPE concentrations also caused primarily apoptosis/late apoptosis, while high CPE concentrations mainly induced necroptosis. Collectively, these results establish that high, but not low, CPE concentrations cause necroptosis and suggest that RIP1, RIP3, MLKL, or calpain inhibitors can be explored as potential therapeutics against CPE effects in vivoIMPORTANCEC. perfringens type F strains are a common cause of food poisoning and antibiotic-associated diarrhea. Type F strain virulence requires production of C. perfringens enterotoxin (CPE). In Caco-2 cells, high CPE concentrations cause necrosis while low enterotoxin concentrations induce apoptosis. The current study determined that receptor-interacting serine/threonine-protein kinases 1 and 3 are involved in both CPE-induced apoptosis and necrosis in Caco-2 cells, while mixed-lineage kinase domain-like pseudokinase (MLKL) oligomerization is involved in CPE-induced necrosis, thereby indicating that this form of CPE-induced cell death involves necroptosis. High CPE concentrations also caused necroptosis in T84 and Vero cells. Calpain activation was identified as a key intermediate for CPE-induced necroptosis. These results suggest inhibitors of RIP1, RIP3, MLKL oligomerization, or calpain are useful therapeutics against CPE-mediated diseases.

Shrestha A ，Mehdizadeh Gohari I ，McClane BA 《mBio》

Necroptosis regulates tumor repopulation after radiotherapy via RIP1/RIP3/MLKL/JNK/IL8 pathway.

Wang Y ，Zhao M ，He S ，Luo Y ，Zhao Y ，Cheng J ，Gong Y ，Xie J ，Wang Y ，Hu B ，Tian L ，Liu X ，Li C ，Huang Q ... -  《JOURNAL OF EXPERIMENTAL & CLINICAL CANCER RESEARCH》