RESUMO
In this study, cantharidin(CTD), a bioactive terpenoid in traditional Chinese medicine cantharidin, was selected as a model component to construct novel nano liposome delivery systems for hepatocellular carcinoma therapy. Previous studies have shown that although cantharidin has definite curative effects on primary liver cancer, it is associated with numerous toxic and side effects. Therefore, based on the glycyrrhetinic acid (GA) binding site and the asialoglycoprotein receptor (ASGPR) on the hepatocyte membrane, the surface of CTD liposomes was modified with stearyl alcohol galactoside (SA-Gal) or/and the newly synthesized 3-succinic-30-stearyl deoxyglycyrrhetinic acid (11-DGA-Suc) ligands, and the physicochemical properties, pharmacokinetics, in vivo and in vitro anti-liver tumor activity and its mechanism of modified liposomes were investigated. Compared to CTD-lip, SA-Gal-CTD-lip, and 11-DGA-Suc + SA-Gal-CTD-lip, 11-DGA-Suc-CTD-lip showed stronger cytotoxicity and increased inhibition of HepG2 cell migration had the highest apoptosis rate. The cell cycle results indicated that HepG2 cells was arrested mainly at G0/G1phase and G2/M phase. The results of in vivo pharmacokinetic experiments revealed that the distribution of modified liposomes in the liver was significantly increased compared with that of unmodified liposome. In vivo tumor inhibition experiment showed that 11-DGA-Suc-CTD-lip had excellent tumor inhibition, and the tumor inhibition rates was 80.96%. The 11-DGA-Suc-CTD-lip group also displayed the strongest proliferation inhibition with the lowest proliferation index of 7% in PCNA assay and the highest apoptotic index of 49% in TUNEL assay. Taken together, our findings provide a promising solution for improving the targeting of nano liposomes and further demonstrates the encouraging potential of poor solubility and high toxicity drugs applicable to tumor therapy.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Lipossomos , Cantaridina/farmacologia , Cantaridina/química , Ligantes , Neoplasias Hepáticas/tratamento farmacológico , Carcinoma Hepatocelular/tratamento farmacológicoRESUMO
The study focuses on a methodology providing noninvasive monitoring and evaluation of the antitumor effect of traditional Chinese medicine, cantharides complex (canth), on 4T1 breast tumor cells. Digital holographic tomography (DHT) and developed data post-processing algorithms were used for quantitative estimation of changes in optical and morphological parameters of cells. We calculated and compared data on the refractive index, thickness, and projected area of 4T1 breast tumor cells in control untreated specimens and those treated with doxorubicin hydrochloride (DOX), canth, and their combinations. Post-treatment changes in cellular morphology recorded by DHT demonstrated that the two drugs led to noticeably different morphological changes in cells that can be presumably associated with different pathways of their death, apoptosis, or necrosis. The effect of combined treatment with these two drugs strongly depended on their relative concentrations and could lead to changes characteristic either for DOX or for canth; however, being more profound than those obtained when using each drug solely. The results obtained by DHT are in a good correspondence with commonly used cell viability analysis and immunofluorescent analysis of changes in cellular cytoskeleton.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Cantaridina/farmacologia , Holografia/métodos , Tomografia/métodos , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias da Mama/diagnóstico por imagem , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/farmacologia , Feminino , Imunofluorescência , Medicina Tradicional Chinesa , Camundongos , Refratometria/métodosRESUMO
BACKGROUND: Sodium cantharidinate/vitamin B6â(SC/VB6) injection, a famous insect-derived traditional Chinese medicine preparation, has been widely applied as a promising adjunctive drug for hepatocellular carcinoma (HCC). However, its exact clinical efficacy and safety is still not well investigated. In this study, we aimed to summarize the efficacy of SC/VB6 injection on survival, liver function, immune function, and quality of life (QoL) in patients with HCC through the meta-analysis. METHODS: All available randomized controlled trials (RCTs) and high-quality prospective cohort studies that investigated the efficacy and safety of SC/VB6 for patients with HCC were searched from ten electronic databases including PubMed, Google Scholar, Cochrane Library, Excerpt Medica Database (Embase), Medline, Web of Science (WOS), Chinese Biomedical Literature Database (CBM), China National Knowledge Infrastructure (CNKI), China Scientific Journal Database (CSJ), and Wanfang Database. Papers in Chinese or English published from January 2000 to July 2020 will be included without any restrictions.Study selection and data extraction will be performed independently by 2 researchers. The clinical outcomes including overall survival (OS), QoL, liver function, immune function, and adverse events, were systematically evaluated. Review Manager 5.3 and Stata 14.0 were used for data analysis, and the quality of the clinical trials was also evaluated. RESULTS: The results of this study will be published in a peer-reviewed journal, and provide a helpful evidence for clinicians to formulate the best postoperative adjuvant treatment strategy for HCC patients. CONCLUSION: Our study will draw an objective conclusion of the efficacy of SC/VB6 on survival, liver function, immune function, and QoL in patients with HCC. INPLASY REGISTRATION NUMBER: INPLASY202070121.
Assuntos
Cantaridina/análogos & derivados , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/patologia , Vitamina B 6/farmacologia , Complexo Vitamínico B/farmacologia , Cantaridina/administração & dosagem , Cantaridina/farmacologia , Carcinoma Hepatocelular/mortalidade , Carcinoma Hepatocelular/psicologia , China/epidemiologia , Quimioterapia Combinada/métodos , Humanos , Sistema Imunitário/efeitos dos fármacos , Injeções/métodos , Fígado/efeitos dos fármacos , Medicina Tradicional Chinesa/métodos , Estudos Prospectivos , Qualidade de Vida , Ensaios Clínicos Controlados Aleatórios como Assunto , Segurança , Sobrevida , Resultado do Tratamento , Vitamina B 6/administração & dosagem , Complexo Vitamínico B/administração & dosagem , Metanálise como AssuntoRESUMO
Tumor necrosis factor (TNF)related apoptosisinducing ligand (TRAIL), a type II transmembrane protein, is a part of the TNF superfamily of cytokines. Cantharidin, a type of terpenoid, is extracted from the blister beetles (Mylabris genus) used in Traditional Chinese Medicine. Cantharidin elicits antibiotic, antiviral and antitumor effects, and can affect the immune response. The present study demonstrated that a cantharidin and TRAIL combination treatment regimen elicited a synergistic outcome in TRAILresistant DU145 cells. Notably, it was also identified that cantharidin treatment initiated the downregulation of cellular FLICElike inhibitory protein (cFLIP) and upregulation of death receptor 5 (DR5), and sensitized cells to TRAILmediated apoptosis by initiating autophagy flux. In addition, cantharidin treatment increased lipidmodified microtubuleassociated proteins 1A/1B light chain 3B expression and significantly attenuated sequestosome 1 expression. Attenuation of autophagy flux by a specific inhibitor such as chloroquine and genetic modification using ATG5 small interfering RNA abrogated the cantharidinmediated TRAILinduced apoptosis. Overall, the results of the present study revealed that cantharidin effectively sensitized cells to TRAILmediated apoptosis and its effects are likely to be mediated by autophagy, the downregulation of cFLIP and the upregulation of DR5. They also suggested that the combination of cantharidin and TRAIL may be a successful therapeutic strategy for TRAILresistant prostate cancer.
Assuntos
Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/metabolismo , Cantaridina/farmacologia , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/genética , Autofagia/genética , Autofagia/fisiologia , Western Blotting , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/genética , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Sinergismo Farmacológico , Imunofluorescência , Humanos , Masculino , Microscopia Eletrônica de Transmissão , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/genética , Proteína Sequestossoma-1/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/genéticaRESUMO
Cantharidin (CTD) is a traditional Chinese medicine that shows an anticancer effects in multiple types of cancer cells. However, the mechanism of CTD anti-cancer function in gastric cancer (GC) is still unclear. The aim of the present study was to investigate the underlying mechanism that CTD inhibits proliferation and migration through suppression of the PI3K/Akt signaling. CTD induced GC cell apoptosis and inhibited metastasis measured by CCK8 assays as well as wound healing assays and transwell assays. Mechanistic investigations suggested that CTD modulated the PI3K/Akt signaling via western-blot and quantitative q-PCR. In addition, we identified and confirmed CCAT1 as a novel direct target of CTD inhibited PI3K/AKt signaling expression. In conclusion, our results provide new point into the critical role of CTD in suppressing PI3K/Akt signaling via down-regulation of CCAT1, resulting in suppression GC cell growth and migration/invasion.
Assuntos
Cantaridina/farmacologia , Movimento Celular/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Longo não Codificante/metabolismo , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , RNA Longo não Codificante/genética , Transdução de Sinais/efeitos dos fármacos , Neoplasias GástricasRESUMO
Cantharidin (CTD) is a potent anticancer small molecule produced by several species of blister beetle. It has been a traditional medicine for the management of warts and tumors for many decades. CTD suppresses tumor growth by inducing apoptosis, cell cycle arrest, and DNA damage and inhibits protein phosphatase 2 phosphatase activator (PP2A) and protein phosphatase 1 (PP1). CTD also alters lipid homeostasis, cell wall integrity, endocytosis, adhesion, and invasion in yeast cells. In this study, we identified additional molecular targets of CTD using a Saccharomyces cerevisiae strain that expresses a cantharidin resistance gene (CRG1), encoding a SAM-dependent methyltransferase that methylates and inactivates CTD. We found that CTD specifically affects phosphatidylethanolamine (PE)-associated functions that can be rescued by supplementing the growth media with ethanolamine (ETA). CTD also perturbed endoplasmic reticulum (ER) homeostasis and cell wall integrity by altering the sorting of glycosylphosphatidylinositol (GPI)-anchored proteins. A CTD-dependent genetic interaction profile of CRG1 revealed that the activity of the lipid phosphatase cell division control protein 1 (Cdc1) in GPI-anchor remodeling is the key target of CTD, independently of PP2A and PP1 activities. Moreover, experiments with human cells further suggested that CTD functions through a conserved mechanism in higher eukaryotes. Altogether, we conclude that CTD induces cytotoxicity by targeting Cdc1 activity in GPI-anchor remodeling in the ER.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Cantaridina/farmacologia , Proteínas de Ciclo Celular/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Glicosilfosfatidilinositóis/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Ciclo Celular/genética , Morte Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Células HeLa , Células Hep G2 , Humanos , Modelos Biológicos , Transporte Proteico/efeitos dos fármacos , Proteínas de Saccharomyces cerevisiae/genéticaRESUMO
PURPOSE: Plasmonic nanostructure-mediated photothermal therapy (PTT) is a promising alternative therapy for the treatment of skin cancer and other diseases. However, the insufficient efficiency of PTT at irradiation levels tolerable to tissues and the limited biodegradability of nanomaterials are still crucial challenges. In this study, a novel nanosystem for PTT based on liposome-nanoparticle assemblies (LNAs) was established. MATERIALS AND METHODS: Thermal-sensitive liposomes (TSLs) encapsulating cantharidin (CTD) were coated with gold nanoparticles (GNPs) and used in near-infrared (NIR) illumination-triggered PTT and thermally induced disruption on A431 cells. RESULTS: The coated GNPs disintegrated into small particles of 5-6 nm after disruption of TSLs, allowing their clearance by the liver and kidneys. CTD encapsulated in the TSLs was released into cytoplasm after PTT. The released CTD increased the apoptosis of PTT-treated tumor cells by blocking the heat shock response (HSR) and inhibiting the expression of HSP70 and BAG3 inhibiting the expression of HSP70 and BAG3 with the synergistic enhancement of CTD, the new nanosystem CTD-encapsulated TSLs coated with GNPs (CTD-TSL@GNPs) had an efficient PTT effect using clinically acceptable irradiation power (200 mW//cm2) on A431 cells. CONCLUSION: The developed CTD-TSL@GNPs may be a promising PTT agent for clinical skin cancer therapy.
Assuntos
Cantaridina/farmacologia , Lipossomos/química , Nanopartículas Metálicas/uso terapêutico , Fototerapia/métodos , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Proteínas Reguladoras de Apoptose/metabolismo , Cantaridina/administração & dosagem , Cantaridina/química , Linhagem Celular Tumoral , Liberação Controlada de Fármacos , Ouro/química , Ouro/farmacologia , Proteínas de Choque Térmico HSP70/metabolismo , Humanos , Nanopartículas Metálicas/químicaRESUMO
As an active constituent of the beetle Mylabris used in traditional Chinese medicine, cantharidin is a potent and selective inhibitor of protein phosphatase 2A (PP2A) that plays a crucial role in cell cycle progression, apoptosis, and cell fate. The role and possible mechanisms exerted by cantharidin in cell growth and metastasis of breast cancer were investigated in this study. Cantharidin was found to inhibit cell viability and clonogenic potential in a time- and dose-dependent manner. Cell cycle analysis revealed that cell percentage in G2/M phase decreased, whereas cells in S and G1 phases progressively accumulated with the increasing doses of cantharidin treatment. In a xenograft model of breast cancer, cantharidin inhibited tumor growth in a dose-dependent manner. Moreover, high doses of cantharidin treatment inhibited cell migration in wound and healing assay and downregulated protein levels of major matrix metalloproteinases (MMP)-2 and MMP-9. MDA-MB-231 cell migration and invasion were dose-dependently inhibited by cantharidin treatment. Interestingly, the members of the mitogen-activated protein kinase (MAPK) signaling family were less phosphorylated as the cantharidin dose increased. Cantharidin was hypothesized to exert its anticancer effect through the MAPK signaling pathway. The data of this study also highlighted the possibility of using PP2A as a therapeutic target for breast cancer treatment.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Cantaridina/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , CamundongosRESUMO
Cantharidin, a monoterpene isolated from the insect blister beetle, has long been used as a medicinal agent in the traditional Chinese medicine. Cantharidin inhibits a subgroup of serine/threonine phosphatases, thus inducing cell growth inhibition and cytotoxicity. Cantharidin has anticancer activity in vitro, since it is able of inducing p53-dependent apoptosis and double-strand breakage of DNA in cancer cells. Although the toxicity of cantharidin to the gastrointestinal and urinary tracts prevents its medical use, it is a promising lead compound for chemical modification to develop new anticancer therapeutics. In fact, cantharidin does not cause myelosuppression and displays anticancer activity against cells with a multidrug resistance phenotype. Here, the competitive inhibitory effect of cantharidin on heme-Fe(III) binding to the fatty acid site 1 (FA1) of human serum albumin (HSA) is reported. Docking and molecular dynamics simulations support functional data indicating the preferential binding of cantharidin to the FA1 site of HSA. Present results may be relevant in vivo as HSA could transport cantharidin, which in turn could affect heme-Fe(III) scavenging by HSA.
Assuntos
Ligação Competitiva , Cantaridina/farmacologia , Ácidos Graxos/metabolismo , Heme/metabolismo , Albumina Sérica Humana/metabolismo , Cantaridina/química , Compostos de Dansil/química , Compostos de Dansil/metabolismo , Humanos , Ligantes , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Ligação Proteica/efeitos dos fármacos , Sarcosina/análogos & derivados , Sarcosina/química , Sarcosina/metabolismo , Albumina Sérica Humana/química , TermodinâmicaRESUMO
As an active constituent of the beetle Mylabris used in traditional Chinese medicine, cantharidin is a potent and selective inhibitor of protein phosphatase 2A (PP2A) that plays a crucial role in cell cycle progression, apoptosis, and cell fate. The role and possible mechanisms exerted by cantharidin in cell growth and metastasis of breast cancer were investigated in this study. Cantharidin was found to inhibit cell viability and clonogenic potential in a time- and dose-dependent manner. Cell cycle analysis revealed that cell percentage in G2/M phase decreased, whereas cells in S and G1 phases progressively accumulated with the increasing doses of cantharidin treatment. In a xenograft model of breast cancer, cantharidin inhibited tumor growth in a dose-dependent manner. Moreover, high doses of cantharidin treatment inhibited cell migration in wound and healing assay and downregulated protein levels of major matrix metalloproteinases (MMP)-2 and MMP-9. MDA-MB-231 cell migration and invasion were dose-dependently inhibited by cantharidin treatment. Interestingly, the members of the mitogen-activated protein kinase (MAPK) signaling family were less phosphorylated as the cantharidin dose increased. Cantharidin was hypothesized to exert its anticancer effect through the MAPK signaling pathway. The data of this study also highlighted the possibility of using PP2A as a therapeutic target for breast cancer treatment.
Assuntos
Humanos , Animais , Feminino , Coelhos , Neoplasias da Mama/tratamento farmacológico , Cantaridina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Antineoplásicos/farmacologia , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Ensaios de Seleção de Medicamentos Antitumorais , Movimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacosRESUMO
Cantharidin (CTD) is an active compound isolated from the traditional Chinese medicine blister beetle and displayed anticancer properties against various types of cancer cells. However, little is known about its effect on human chronic myeloid leukemia (CML) cells, including imatinib-resistant CML cells. The objective of this study was to investigate whether CTD could overcome imatinib resistance in imatinib-resistant CML cells and to explore the possible underlying mechanisms associated with the effect. Our results showed that CTD strongly inhibited the growth of both imatinib-sensitive and imatinib-resistant CML cells. CTD induced cell cycle arrest at mitotic phase and triggered DNA damage in CML cells. The ATM/ATR inhibitor CGK733 abrogated CTD-induced mitotic arrest but promoted the cytotoxic effects of CTD. In addition, we demonstrated that CTD downregulated the expression of the BCR-ABL protein and suppressed its downstream signal transduction. Real-time quantitative PCR revealed that CTD inhibited BCR-ABL at transcriptional level. Knockdown of BCR-ABL increased the cell-killing effects of CTD in K562 cells. These findings indicated that CTD overcomes imatinib resistance through depletion of BCR-ABL. Taken together, CTD is an important new candidate agent for CML therapy.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Cantaridina/farmacologia , Proteínas de Fusão bcr-abl/deficiência , Mesilato de Imatinib/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Cantaridina/administração & dosagem , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Dano ao DNA , Sinergismo Farmacológico , Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Mesilato de Imatinib/administração & dosagem , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologiaRESUMO
The aim of this study was to assess the influence of cantharidin on the activities of the drug-metabolizing enzymes CYP1A2, CYP2C9, CYP2C19, CYP2D6 and CYP3A4 in rats. The activities of CYP1A2, CYP2C9, CYP2C19, CYP2D6 and CYP3A4 were measured using specific probe drugs. After pretreatment for 1week with cantharidin or physiological saline (control group) by intraperitoneal injection, probe drugs phenacetin (5.0mg/kg; CYP1A2 activity), tolbutamide (1.0mg/kg; CYP2C9 activity), omeprazole (10mg/kg; CYP2C19 activity), metoprolol (20mg/kg; CYP2D6 activity) and midazolam (10mg/kg; CYP3A4 activity) were administered to rats by oral administration. The blood was then collected at different times for ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) analysis. The data showed that cantharidin exhibits an inhibitory effect on CYP2D6 and CYP3A4 by increasing t1/2, Cmax and AUC(0-∞), and decreasing CL/F compared with those of the control group. In addition, cantharidin has induction effect on CYP2C9 activity. However, no significant changes in CYP1A2 and CYP2C19 activities were observed. In conclusion, the results indicated that cantharidin could inhibit CYP2D6 and CYP3A4, while induce CYP2C9, which may affect the disposition of medicines primarily dependent on these pathways. Our work may be the basis of related herb-drug interactions in the clinic.
Assuntos
Cantaridina/farmacologia , Indutores das Enzimas do Citocromo P-450/farmacologia , Inibidores das Enzimas do Citocromo P-450/farmacologia , Sistema Enzimático do Citocromo P-450/efeitos dos fármacos , Animais , Cromatografia Líquida de Alta Pressão , Indutores do Citocromo P-450 CYP2C9 , Inibidores do Citocromo P-450 CYP2D6 , Inibidores do Citocromo P-450 CYP3A , Interações Ervas-Drogas , Masculino , Ratos , Ratos Sprague-Dawley , Espectrometria de Massas em TandemRESUMO
Cantharidin is an active constituent of mylabris, a traditional Chinese medicine, and is a potent and selective inhibitor of protein phosphatase 2A (PP2A) that plays an important role in cell cycle control, apoptosis, and cell-fate determination. In the present study, we found that cantharidin repressed the invasive ability of pancreatic cancer cells and downregulated matrix metalloproteinase 2 (MMP2) expression through multiple pathways, including ERK, JNK, PKC, NF-κB, and ß-catenin. Interestingly, transcriptional activity of the MMP2 promoter increased after treatment with PP2A inhibitors, suggesting the involvement of a posttranscriptional mechanism. By using an mRNA stability assay, we found accelerated degradation of MMP2 mRNA after treatment of cantharidin. Microarray analyses revealed that multiple genes involved in the 3' â 5' decay pathway were upregulated, especially genes participating in cytoplasmic deadenylation. The elevation of these genes were further demonstrated to be executed through ERK, JNK, PKC, NF-κB, and ß-catenin pathways. Knockdown of PARN, RHAU, and CNOT7, three critical members involved in cytoplasmic deadenylation, attenuated the downregulation of MMP2. Hence, we present the mechanism of repressed invasion by cantharidin and other PP2A inhibitors through increased degradation of MMP2 mRNA by elevated cytoplasmic deadenylation.
Assuntos
Antineoplásicos/farmacologia , Cantaridina/farmacologia , Metaloproteinase 2 da Matriz/metabolismo , Neoplasias Pancreáticas/tratamento farmacológico , Estabilidade de RNA/efeitos dos fármacos , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Repressão Enzimática/efeitos dos fármacos , Humanos , Metaloproteinase 2 da Matriz/genética , Invasividade Neoplásica , Proteína Fosfatase 2/antagonistas & inibidores , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de SinaisRESUMO
Bladder cancer is a common malignancy worldwide. However, there is still no effective therapy for bladder cancer. In this study, we investigated the cytotoxic effects of cantharidin [a natural toxin produced (pure compound) from Chinese blister beetles (Mylabrisphalerata or Mylabriscichorii) and Spanish flies (Cantharis vesicatoria)] in human bladder cancer cell lines (including: T24 and RT4 cells). Treatment of human bladder cancer cells with cantharidin significantly decreased cell viability. The increase in the expressions of caspase-3 activity and cleaved form of caspase-9/-7/-3 were also increased in cantharidin-treated T24 cells. Furthermore, cantharidin increased the levels of phospho-eIF2α and Grp78 and decreased the protein expression of procaspase-12, which was accompanied by the increase in calpain activity in T24 cells. Cantharidin was capable of increasing the intracellular Ca (2+) and the phosphorylation of protein kinase C (PKC) in T24 cells. The addition of BAPTA/AM (a Ca (2+) chelator) and RO320432 (a selective cell-permeable PKC inhibitor) effectively reversed the increase in caspase-3 and calpain activity, the phosphorylation levels of PKC and eIF2α and Grp78 protein expression, and the decrease in procaspase-12 expression induced by cantharidin. Importantly, cantharidin significantly decreased the tumor volume (a dramatic 71% reduction after 21 days of treatment) in nude mice xenografted with T24 cells. Taken together, these results indicate cantharidin induced human bladder cancer cell apoptosis through a calcium/PKC-regulated ER stress pathway. These findings suggest that cantharidin may be a novel and potential anticancer agent targeting on bladder cancer cells.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Cantaridina/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Papiloma/genética , Transdução de Sinais/efeitos dos fármacos , Neoplasias da Bexiga Urinária/genética , Animais , Cálcio/fisiologia , Caspase 3/metabolismo , Linhagem Celular Tumoral , Chaperona BiP do Retículo Endoplasmático , Humanos , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Papiloma/patologia , Proteína Quinase C/fisiologia , Regulação para Cima/efeitos dos fármacos , Neoplasias da Bexiga Urinária/patologiaRESUMO
Cantharidin is an active component of mylabris, which has been used as a traditional Chinese medicine. Cantharidin has been shown to have antitumor activity against several types of human cancers in vitro and in animal models in vivo. We investigated whether cantharidin induces DNA damage and affects DNA damage repair-associated protein levels in TSGH8301 human bladder cancer cells. Using flow cytometry to measure viable cells, cantharidin was found to reduce the number of viable cells in a dose-dependent manner. Comet assay, 4',6-diamidino-2-phenylindole (DAPI) staining and DNA gel electrophoresis were used to measure DNA damage and condensation; the results indicated that cantharidin induced DNA damage (comet tail), DNA condensation (white DAPI staining) and DNA damage (DNA smear). Results from western blotting showed that cantharidin inhibited the expression of DNA-dependent serine/threonine protein kinase, poly-ADP ribose polymerase, phosphate-ataxia-telangiectasia and RAD3-related, O-6-methylguanine-DNA methyltransferase, breast cancer susceptibility protein 1, mediator of DNA damage checkpoint protein 1, phospho-histone H2A.X, but increased that of phosphorylated p53 following 6 and 24 h treatment. Confocal laser microscopy was used to examine the protein translocation; cantharidin suppressed the levels of p-H2A.X and MDC1 but increased the levels of p-p53 in TSGH8301 cells. In conclusion, we found that cantharidin-induced cell death may occur through the induction of DNA damage and suppression of DNA repair-associated protein expression in TSGH8301 cells.
Assuntos
Cantaridina/farmacologia , Dano ao DNA , Reparo do DNA , Proteínas de Neoplasias/antagonistas & inibidores , Neoplasias da Bexiga Urinária/patologia , Linhagem Celular Tumoral , Humanos , Microscopia Confocal , Proteínas de Neoplasias/metabolismo , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/metabolismoRESUMO
Cantharidin is an active constituent of mylabris, a traditional Chinese medicine, and presents strong anticancer activity in various cell lines. Cantharidin is a potent and selective inhibitor of serine/threonine protein phosphatase 2A (PP2A). Our previous studies revealed the prospect of application of cantharidin, as well as other PP2A inhibitors, in the treatment of pancreatic cancer. However, the mechanisms involved in the anticancer effect of PP2A inhibitors have not been fully explored. The Wnt/ßcatenin pathway is involved in cell migration and proliferation and participates in the progression of pancreatic cancer. If ßcatenin is phosphorylated and degraded, the Wnt/ßcatenin pathway is blocked. PP2A dephosphorylates ßcatenin and keeps the Wnt/ßcatenin pathway active. In the present study, we found that PP2A inhibitor treatment induced phosphorylation and degradation of ßcatenin. The suppression on the migration and growth of PANC1 pancreatic cancer cells could be attenuated by pretreatment with FH535, a ßcatenin pathway inhibitor. Microarray showed that PP2A inhibitor treatment induced expression changes in 13 of 138 genes downstream of the ßcatenin pathway. Realtime PCR further confirmed that FH535 attenuated the expression changes induced by PP2A inhibitors in 6 of these 13 candidate genes. These 6 genes, VEGFB, Dkk3, KRT8, NRP1, Cacnalg and WISP2, have been confirmed to participate in the migration and/or growth regulation in previous studies. Thus, the phosphorylation- and degradation-mediated suppression on ßcatenin participates in the cytotoxicity of PP2A inhibitors. Our findings may provide insight into the treatment of pancreatic cancer using a targeting PP2A strategy.
Assuntos
Inibidores Enzimáticos/farmacologia , Neoplasias Pancreáticas/patologia , Proteína Fosfatase 2/antagonistas & inibidores , beta Catenina/metabolismo , Antracenos/farmacologia , Cantaridina/farmacologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Pancreáticas/tratamento farmacológico , Fosforilação , Sulfonamidas/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , beta Catenina/genéticaRESUMO
BACKGROUND: [corrected] Cantharidin, and its derivatives can not only inhibit the proliferation of tumor cells, but can also induce tumor cell apoptosis. It shows cantharidin exhibits a wide range of reactivity in anticancer. The objective of this paper was to study the inhibitory effect of sodium cantharidinate on human hepatoma HepG2 cells. MATERIALS AND METHODS: MTT assay was used to detect the proliferation of HepG2 cells, and immunohisto-chemical method was used to detect the change in VEGF, protein level, and to determine the inhibitory effect of sodium cantharidinate on human hepatoma HepG2 cells. RESULTS: As results, sodium cantharidinate significantly inhibited the growth of HepG2 cells in a time-and dose-dependent manner. CONCLUSION: We conclude that sodium cantharidinate has an inhibitory effect on human hepatoma HepG2 cells.
Assuntos
Antineoplásicos/uso terapêutico , Cantaridina/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Besouros/química , Neoplasias Hepáticas/tratamento farmacológico , Animais , Antineoplásicos/farmacologia , Apoptose , Cantaridina/análogos & derivados , Cantaridina/farmacologia , Proliferação de Células , Relação Dose-Resposta a Droga , Células Hep G2 , Humanos , SódioRESUMO
Heat shock factor 1 (HSF1) enhances the survival of cancer cells under various stresses. The knock-out of HSF1 impairs cancer formation and progression, suggesting that HSF1 is a promising therapeutic target. To identify inhibitors of HSF1 activity, we performed cell-based screening with a library of marketed and experimental drugs and identified cantharidin as an HSF1 inhibitor. Cantharidin is a potent antitumor agent from traditional Chinese medicine. Cantharidin inhibited heat shock-induced luciferase activity with an IC50 of 4.2 µm. In contrast, cantharidin did not inhibit NF-κB luciferase reporter activity, demonstrating that cantharidin is not a general transcription inhibitor. When the HCT-116 colorectal cancer cells were exposed to heat shock in the presence of cantharidin, the induction of HSF1 downstream target proteins, such as HSP70 and BAG3 (Bcl-2-associated athanogene domain 3), was suppressed. HSP70 and its co-chaperone BAG3 have been reported to protect cells from apoptosis by stabilizing anti-apoptotic Bcl-2 family proteins. As expected, treating HCT-116 cancer cells with cantharidin significantly decreased the amounts of BCL-2, BCL-xL, and MCL-1 protein and induced apoptotic cell death. Chromatin immunoprecipitation analysis showed that cantharidin inhibited the binding of HSF1 to the HSP70 promoter and subsequently blocked HSF1-dependent p-TEFb recruitment. Therefore, the p-TEFb-dependent phosphorylation of the C-terminal domain of RNA polymerase II was blocked, arresting transcription at the elongation step. Protein phosphatase 2A inhibition with PP2CA siRNA or okadaic acid did not block HSF1 activity, suggesting that cantharidin inhibits HSF1 in a protein phosphatase 2A-independent manner. We show for the first time that cantharidin inhibits HSF1 transcriptional activity.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Cantaridina/farmacologia , Proteínas de Ligação a DNA/metabolismo , Proteínas de Choque Térmico HSP70/antagonistas & inibidores , Neoplasias/patologia , Regiões Promotoras Genéticas/genética , Fatores de Transcrição/metabolismo , Acetilação/efeitos dos fármacos , Proteínas Reguladoras de Apoptose , Cantaridina/química , Cantaridina/uso terapêutico , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Fatores de Transcrição de Choque Térmico , Resposta ao Choque Térmico/efeitos dos fármacos , Humanos , Mitose/efeitos dos fármacos , Mitose/genética , Modelos Biológicos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/genética , Fator B de Elongação Transcricional Positiva , Inibidores de Proteases/farmacologia , Ligação Proteica/efeitos dos fármacos , Proteína Fosfatase 2/antagonistas & inibidores , Proteína Fosfatase 2/metabolismo , Transporte Proteico/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteína bcl-X/metabolismoRESUMO
Cancer metastasis is a highly coordinated and dynamic multistep process in which cancer cells interact with a variety of host cells. Morphological studies have documented the association of circulating tumor cells with host platelets, where a surface coating of platelets protects tumor cells from mechanical trauma and the immune system. Cantharidin is an active constituent of mylabris, a traditional Chinese medicine. Cantharidin and norcantharidin are potent protein phosphatase 2A (PP2A) inhibitors that exhibit in vitro and in vivo antitumor activity against several types of cancer, including breast cancer. We investigated whether cantharidin and norcantharidin could repress the ability of MCF-7 breast cancer cells to adhere to platelets. Using MTT, clone formation, apoptosis, adhesion and wound-healing assays, we found that cantharidin and norcantharidin induced apoptosis and repressed MCF-7 cell growth, adhesion and migration. Moreover, we developed a flow cytometry-based analysis of tumor cell adhesion to platelets. We proved that cantharidin and norcantharidin repressed MCF-7 cell adhesion to platelets through downregulation of α2 integrin, an adhesion molecule present on the surface of cancer cells. The repression of α2 integrin expression was found to be executed through the protein kinase C pathway, the activation of which could have been due to PP2A inhibition.
Assuntos
Plaquetas/metabolismo , Neoplasias da Mama/tratamento farmacológico , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Cantaridina/farmacologia , Integrina alfa2/química , Adesividade Plaquetária/efeitos dos fármacos , Proteína Quinase C/metabolismo , Apoptose/efeitos dos fármacos , Plaquetas/efeitos dos fármacos , Western Blotting , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Adesão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo , Inibidores Enzimáticos/farmacologia , Feminino , Citometria de Fluxo , Humanos , Integrina alfa2/genética , Integrina alfa2/metabolismo , Células MCF-7 , Proteína Quinase C/genética , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Transcrição RelA/antagonistas & inibidores , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/metabolismo , Cicatrização/efeitos dos fármacosRESUMO
Norcantharidin (NCTD) is currently used for anticancer therapy but the exact mechanism of action remains unknown. Pre-replicative complexes (pre-RCs) are essential for cell DNA replication and highly related to malignant proliferation. Here, we examined the inhibitory effect of NCTD on pre-RC components in HepG2 cells. We showed that NCTD induced degradation of Cdc6 and Mcm2 in a dose-dependent manner. Under 100 µM NCTD concentration, about 70% of Cdc6 and 50% of Mcm2 were degraded. In addition, the nuclear translocation of Mcm6 was inhibited by NCTD. Further studies aiming at G1 synchronous cells showed that, NCTD reduced the chromatin-bound Cdc6, Mcm2 and Mcm6. Moreover, the cells were blocked from entering the S phase and accumulated at the G1 phase when released synchronously into the cell cycle. Consistently, the DNA replication was inhibited by NCTD. Finally, the combination NCTD with Cdc6 depletion lead to more severe cytotoxicity (88%) than NCTD (52%) and Cdc6 depletion (39%) alone. A synergic cytotoxicity was observed between Cdc6 depletion and NCTD. In conclusion, our results demonstrate that NCTD inhibits pre-RC assembly; subsequently blocks the G1 to S transition; and inhibits DNA replication in HepG2 cells. Pre-RCs are an intriguing target for cancer therapy, which merits further investigations for anticancer development.