Objectives:Targeting epigenetic modifications in anticancer therapy is a promising approach to overcoming cancer cell chemoresistance.The histone deacetylase/DNA methyltransferase inhibitor,parthenolide(PTL),has antit...Objectives:Targeting epigenetic modifications in anticancer therapy is a promising approach to overcoming cancer cell chemoresistance.The histone deacetylase/DNA methyltransferase inhibitor,parthenolide(PTL),has antitumor activity,but contrasting findings exist on its effect in normal cells.This study aims to examine the nongenomic toxic mechanisms of PTL in human erythrocytes.Methods:Cell death as stimulated by 20–200μMof PTL for 24 h at 37℃ was assessed using fluorescence-assorted cell sorting and spectrophotometric assays.Canonical markers of cell death,including membrane scrambling,oxidative stress,and Ca^(2+)mobilization,were captured by annexin V-fluorescein isothiocyanate,2′,7′-dichlorodihydrofluorescein diacetate,and Fluo4/AM labeling,respectively.Rescue experiments usingawide arrayof inhibitorswere also conducted.Results:PTL stimulated significantmembrane scrambling and blebbing,and showed potent hemolytic activity coupled with significant elevations in dichlorofluorescein and Fluo4 fluorescence.While hemolysis was ameliorated by glutathione,caffeine,acetylsalicylic acid,staurosporin,necrosulfonamide,guanosine,and Ca^(2+)deprivation,it was rather exacerbated by necrostatin-2,tumor necrosis factor α(TNFα)or Fas ligand(FasL)neutralization,and concurrent Ca^(2+)deprivation and membrane depolarization.In contrast,eryptosis was attenuated by N-acetyl-cysteine,TNFα,or FasL blockade,and simultaneous Ca^(2+)elimination and KCl enrichment;and augmented by necrostatin-2,necrosulfonamide,and adenosine triphosphate.Interestingly,loss of volume was only prevented by melatonin,acetylsalicylic acid,and N(gamma)-nitro-L-arginine methyl ester.Conclusion:PTL stimulates oxidative hemolysis and eryptosis through Ca^(2+)mobilization and death ligand signaling involving the cyclooxygenase/protein kinase C/mixed lineage kinase domain-like pseudokinase axis.This research highlights the non-genomic toxic mechanisms of PTL and presents potential pharmacological targets for mitigating its adverse off-target effects.展开更多
基金funded by the Ongoing Research Funding Program at King Saud University,Riyadh,Saudi Arabia through grant number ORF-2025-554.
文摘Objectives:Targeting epigenetic modifications in anticancer therapy is a promising approach to overcoming cancer cell chemoresistance.The histone deacetylase/DNA methyltransferase inhibitor,parthenolide(PTL),has antitumor activity,but contrasting findings exist on its effect in normal cells.This study aims to examine the nongenomic toxic mechanisms of PTL in human erythrocytes.Methods:Cell death as stimulated by 20–200μMof PTL for 24 h at 37℃ was assessed using fluorescence-assorted cell sorting and spectrophotometric assays.Canonical markers of cell death,including membrane scrambling,oxidative stress,and Ca^(2+)mobilization,were captured by annexin V-fluorescein isothiocyanate,2′,7′-dichlorodihydrofluorescein diacetate,and Fluo4/AM labeling,respectively.Rescue experiments usingawide arrayof inhibitorswere also conducted.Results:PTL stimulated significantmembrane scrambling and blebbing,and showed potent hemolytic activity coupled with significant elevations in dichlorofluorescein and Fluo4 fluorescence.While hemolysis was ameliorated by glutathione,caffeine,acetylsalicylic acid,staurosporin,necrosulfonamide,guanosine,and Ca^(2+)deprivation,it was rather exacerbated by necrostatin-2,tumor necrosis factor α(TNFα)or Fas ligand(FasL)neutralization,and concurrent Ca^(2+)deprivation and membrane depolarization.In contrast,eryptosis was attenuated by N-acetyl-cysteine,TNFα,or FasL blockade,and simultaneous Ca^(2+)elimination and KCl enrichment;and augmented by necrostatin-2,necrosulfonamide,and adenosine triphosphate.Interestingly,loss of volume was only prevented by melatonin,acetylsalicylic acid,and N(gamma)-nitro-L-arginine methyl ester.Conclusion:PTL stimulates oxidative hemolysis and eryptosis through Ca^(2+)mobilization and death ligand signaling involving the cyclooxygenase/protein kinase C/mixed lineage kinase domain-like pseudokinase axis.This research highlights the non-genomic toxic mechanisms of PTL and presents potential pharmacological targets for mitigating its adverse off-target effects.
