| The malignant tumor has been one of the factors that are seriously harmful to human health at present worldwide.The development of new effective drugs is an important concern.Recently,precision medicines are becoming more and more attractive due to characteristics such as high efficiency and low toxicity.Advances in precision medicine have been shifting therapeutics away from untargeted chemotherapies that damage normal cells besides cancerous cells.Targeted therapy is a significant element of precision medicine in oncology,and includes a wide range of cellular,molecular,environmental,genomic and subcellular organelle-targeted therapies.The lysosomes in cancer cells are more numerous,larger,less stable,and have greater cathepsin activity than those in the normal cells.Mitochondria are of fundamental importance in tumour growth and progression.There are differences between the mitochondria of cancer cells and non-neoplastic cells.As the Achilles heel of cancer,lysosomes and mitochondria hold promise as drug targets for improved and selective drug therapy.In this study,we investigated the underlying mechanisms of several natural products and their derivatives targeting for lysosomes and mitochondria,respectively.Lysosomes are important targets for anticancer drug discovery.Our previous study showed that Riccardin D-N(RD-N),a natural macrocylic bisbibenzyl derivative produced by Mannich reaction,induced cell death by accumulating in lysosomes.Experiments were performed on human lung squamous cell carcinoma tissue from left inferior lobar bronchus of patient xenografts and H460 cells.RD-N was administrated for 25 days.The specimens of xenografts in Balb/c athymic(nu+/nu+)male mice were removed for immunohistochemistry,subcellular fractionation,enzyme activities and Western blot analysis.mRFP-GFP-LC3 reporter was used to examine autophagy in H460 cells.Sphingomyelin assay was evaluated by thin-layer chromatography and assay kit.Lysosomal membrane permeabilization(LMP)caused by acid sphingomyelinase(ASM)inhibition and subsequent changes of sphingomyelin(SM)metabolism selectively destabilized the cancer cell lysosomes in RD-N-treated H460 cells in vitro and tumor xenograft model in vivo.The destabilized lysosomes induced the release of cathepsins from the lysosomes into the cytosol and further triggered cell death.These results explain the underlying mechanism of RD-N induced LMP.It can be concluded that a more lysosomotropic derivative was synthesized by introduction of an amine group,which could have more potential applications in cancer therapy.RDD648,a novel derivative of a natural molecule riccardin D,exhibits potent anticancer activity by targeting lysosomes in breast cancer cells in vitro and in vivo.Mechanistic studies revealed that RDD648 facilitated STAT3 translocation into the nucleus,and this activity is involved in lysosome-mediated cell death as evidenced by our finding that inhibition of STAT3 alleviated lysosomal membrane permeabilization.Further investigation indicated that nuclear STAT3 directly interacted with transcription factor TFEB,leading to the partial loss of function of TFEB which is essential for lysosome turnover.The present study first uncovered that STAT3 contributes to lysosomal-mediated cell death in RDD648-treated breast cancer cells though interacting with TFEB,and the findings might be significant in the design of treatments for breast cancers where STAT3 is constitutively expressed.Natural products play important roles in both drug discovery and chemical biology.Among them,diterpenoids exhibit considerable pharmacological activities and many of them have recently been developed into cancer therapeutic agents,such as paclitaxel,oridonin,triptolide.Our synthesis commenced with stevioside,which is naturally abundant and commercially available.Weakly basic drugs have been found to accumulate specifically in lysosomes due to the acidic lysosomal conditions.Further diverse modifications on D-ring led us to the discovery of Compd I with potent anticancer activity with IC50 value of 0.29 μM.And the mechanism research confirmed that it could damage lysosome to cause cell death.In order to enhance its anticancer activity,Compd I was combined with ADR.In our study,ADR induced autophagy flux in A549 cells,while Compd Ⅰ damaged lysosome and inhibited the degration of autophagosome,which had a synergetic effect in triggering cell death.The research provided the theory foundation for the further study about Compd I and a deeper investigation about target and mechanism was on the way.The characteristics of mitochondria in neoplasia make itself be a successful target for cancer therapy,and many mitochondrially-targeted diterpenoids represent a promising approach to eradicate chemotherapy-refractory cancer cells.Recently,compounds with triphenylphosphonium moiety have become attractive,since they might selectively accumulate within tumor mitochondria to elicit cytotoxic effect.In the present study,Compd Ⅱ and Compd Ⅲ,synthesized novel compounds bearing a triphenylphosphonium moiety,displayed excellent cytotoxicity against cancer cell lines.A series of results demonstrated that Compd Ⅱ and Compd Ⅲ could target and disrupt mitochondria,resulting in intrinsic apoptosis in A549 cells.The combination therapy using our compounds with syrosingopine,a clinical antihypertensive drug,was proved to be more targeted,less toxic and more efficient therapeutic designs.Thus,investigation provided potent anticancer agents,Compd Ⅱ and Compd Ⅲ,which might merit investigation as a potential therapeutic agent candidate as a single agent or combination with other targeted agents for treatment of non-small cell lung cancer.Natural resources have been an important supply of anticancer agents.In an attempt to detect leads especially in small amount and measure the anticancer activity more suitably and directly,the potential of TLC bioautography for the fast identification of cytotoxic natural products was studied,as there are few bioautographic assays available for the profiling of cyctotoxic natural sources.In this study,cancer cell bioautography had been established,showing advantages in the localization of toxic zones on TLC plates and coupled to TLC-MS for structural elucidation.A streamlined strategy for rapid screening and reliable characterization of anticancer constitutes inChinese liverwort was demonstrated.The innovation and significance of the project:1.We have demonstrated that RD-N induced LMP by inhibiting acid sphingomyelinase and interfering with sphingomyelin in vivo for the first time.2.The present study first uncovered that STAT3 contributes to lysosomal-mediated cell death in RDD648-treated breast cancer cells though interacting with TFEB,and the findings might be significant in the design of treatments for breast cancers where STAT3 is constitutively expressed.3.The combination research provided the theory foundation for the further study about Compd I and Compd II.4.A streamlined strategy for rapid screening and reliable characterization of anticancer constitutes in Chinese liverwort was demonstrated. |
