| Ursane-type derivatives are a major group of natural pentacyclic triterpenes that have been proven to have chemopreventive properties against various cancers.However,the anti-cancer effects of these derivatives against glioblastoma multiforme(GBM)are not known and the underlying mechanisms of their antineoplastic activity are not fully understood.In the present study,we isolated 14 ursane-type triterpene analogues from the root of Rosa cymosa Tratt and determined the anti-glioma activities of these derivatives in four established temozolomide-resistant GBM cell lines.Based on cell viability analysis,we evaluated the initial structure-activity relationship(SAR)of these 14 compounds and found that compound12 exhibited the strongest anti-glioma activity.Furthermore,we identified the pro-apoptotic effect of compound 12 and explored its potential molecular mechanisms and targets.We demonstrated that Drp1-dependent mitochondrial fission was involved in the induction of apoptosis by compound 12 and that its in-depth mechanism might contribute to the attenuation of Mia40 and Erv1/ALR,which blocked the mitochondrial intermembrane space assembly and retro-translocation in temozolomide-resistant GBM cells.Results1.The viability(% of control)after a 48 h treatment with the tested compounds at various concentrations was determined,and the half maximal inhibitory concentration(IC50)values for growth inhibition were summarized to evaluate the anti-glioma activity.we evaluated the anti-glioma activity of 14 tested compounds in four established TMZ-resistant cell lines,most tested compounds showed dose-dependent activity and had an IC50 value of <100 μM(excepted 2,5,10,13 and 14)against the TMZ-resistant GBM cell lines.However,only five showed better inhibitory action(compounds 3,4 and 7,IC50 values of <50 μM;compounds11 and 12,IC50 values of <20 μM).Comparing with these naturally occurring and synthetic anti-cancer agents,compound 12 exhibited the strongest anti-proliferative activity in multiple cell lines.2.The results from the current study provided data on the initial structure–activity relationship(SAR)of ursane-type derivatives of this plant.These data identified the electron-deficient moieties at C-2 or C-3 of the A ring moiety as critical for the anticancer activity in this family of natural products.3.C-12 caused a dramatic increase in caspase-3 activity,triggered mitochondrial dysfunction and pro-apoptotic signaling in TR/U118 and TR/U87 GBM cells.The present observation proposed that Mia40 might be a target of ursane-type triterpenes derivatives C-12 to encourage mitochondrial apoptotic signalling via obstructing MIA pathway.4.The automated docking simulation was performed using Autodock tools(ADTs)to assess the possible binding orientations and conformations of the C-12 with the Mia40 protein.For docking studies,the active CPC(Cys53-Pro54-Cys55)redox site of Mia40 was considered as the most likely region for C-12 binding among six conserved cysteine residues of Mia40(-CPC-CX9C-CX9C-),since the CPC motif participates in the formation of a mixed disulfide bonded intermediate that is crucial for the substrates protein folding.ConclusionIn conclusion,the present study first determined the cytotoxicity and pro-apoptotic effect of pomonic acid derivatives in GBM cells and evaluated the initial SAR.Furthermore,we identified the pro-apoptotic mechanism by which C-12 induced apoptosis through Drp1-dependent mitochondrial fission via decreasing Mia40 and ALR.Although an in-depth understanding of the molecular mechanisms both in vitro and in vivo should be further explored,the pomonic acid derivatives might be considered to be leading compounds for the chemotherapy of malignant glioma,and structural modification studies should also be encouraged to obtain additional valuable analogues. |
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