Cucurbitacin Research Articles

Cucurbitacin I inhibits STAT3, but enhances STAT1 signaling in human cancer cells in vitro through disrupting actin filaments.

STAT1 and STAT3 are two important members of the STAT (signal transducers and activators of transcription) protein family and play opposing roles in regulating cancer cell growth. Suppressing STAT3 and/or enhancing STAT1 signaling are considered to be attractive anticancer strategies. Cucurbitacin I (CuI) isolated from the cucurbitacin family was reported to be an inhibitor of STAT3 signaling and a disruptor of actin cytoskeleton. In this study we investigated the function and mechanisms of CuI in regulating STAT signaling in human cancer cells in vitro. CuI (0.1-10 mmol/L) dose-dependently inhibited the phosphorylation of STAT3, and enhanced the phosphorylation of STAT1 in lung adenocarcinoma A549 cells possibly through disrupting actin filaments. We further demonstrated that actin filaments physically associated with JAK2 and STAT3 in A549 cells and regulated their phosphorylation through two signaling complexes, the IL-6 receptor complex and the focal adhesion complex. Actin filaments also interacted with STAT1 in A549 cells and regulated its dephosphorylation. Taken together, this study reveals the molecular mechanisms of CuI in the regulation of STAT signaling and in a possible inhibition of human cancer cell growth. More importantly, this study uncovers a novel role of actin and actin-associated signaling complexes in regulating STAT signaling.

Cucurbitacin B and SCH772984 exhibit synergistic anti-pancreatic cancer activities by suppressing EGFR, PI3K/Akt/mTOR, STAT3 and ERK signaling.

Cucurbitacin B (CuB) is a natural tetracyclic triterpene product and displays antitumor activity across a wide array of cancers. In this study, we explored the anti-pancreatic cancer activity of CuB alone and in combination with SCH772984, an ERK inhibitor, in vitro and in vivo. CuB inhibited proliferation of pancreatic cancer cells by arresting them in the G2/M cell cycle phase. This was associated with inhibition of EGFR expression and activity and downstream signaling, including PI3K/Akt/mTOR and STAT3. Interestingly, ERK activity was markedly enhanced by activating AMPK signaling after 12 h of CuB treatment. SCH772984 potentiates the cytotoxic effect of CuB on pancreatic cancer cells through complementary inhibition of EGFR, PI3K/Akt/mTOR, STAT3 and ERK signaling, followed by an increase in the pro-apoptotic protein Bim and a decrease in the anti-apoptotic proteins Mcl-1, Bcl-2, Bcl-xl and survivin. Furthermore, combined therapy with CuB and SCH772984 resulted in highly significant growth inhibition of pancreatic cancer xenografts. These results may provide a basis for further development of combining CuB and ERK inhibitors to treat pancreatic cancer.

Simultaneous determination of cucurbitacin B and cucurbitacin E in rat plasma by UHPLC-MS/MS: A pharmacokinetics study after oral administration of cucurbitacin tablets.

Cucurbitacin B (CuB) and cucurbitacin E (CuE) are tetracyclic triterpene compounds from Cucurbitaceae, and the main bioactive compounds of cucurbitacins tablets that used to treatment of chronic hepatitis. Pharmacological research has been very comprehensive, and there are few studies on pharmacokinetics, especially about CuE. An Ultra High Performance Liquid Chromatography-tandem Mass Spectrometry (UHPLC-MS/MS) method with high selectivity, simplicity and sensitivity has been used for quantitative analysis of Cucurbitacin B (CuB) and cucurbitacin E (CuE). Plasma samples were pretreatment by Liquid-liquid extraction (LLE) method with dichloromethane. The chromatographic separation was achieved on a C18 column (Agilent Eclipse Plus, 1.8μm, 50×2.1mm) using gradient elution with water - methanol at a flow rate of 0.3mL/min and the column temperature was set at 30°C. The method was validated according to FDA guidelines. Lower limit of quantification (LLOQ) was 1.60ng/mL for CuB and 1.58ng/mL for CuE. Correlation coefficients of CuB and CuE were more than 0.99 in rat plasma. All values of intra-day and inter-day precision (RSD%) were not exceeded 15%, the accuracy (RE%) were within -5.57 to 5.20% for CuB and -3.33 to 7.37% for CuE. The mean extraction recoveries were more than 80%. Pharmacokinetic parameters were also evaluated by UHPLC-MS/MS method. The results suggestion that this method was successfully applied to pharmacokinetic study of CuB and CuE in rat plasma after oral administration cucurbitacin tablets.

Effects of cucurbitacin I on in vitro proliferation of HaCaT cells and the expression of keratin 17, signal transducer and activator of transcription 3 and vascular endothelial growth factor

Objective To evaluate effects of cucurbitacin I on in vitro proliferation of HaCaT cells and the expression of keratin 17 (K17) , signal transducer and activator of transcription 3 (STAT3) and vascular endothelial growth factor (VEGF) in cultured HaCaT cells. Methods In vitro cultured HaCaT cells were divided into 6 groups to be treated with cucurbitacin I at different concentrations of 0.0125, 0.025, 0.05 and 0.1 μmol/L (0.0125, 0.025, 0.05 and 0.1 μmol/L cucurbitacin I groups) , DMEM containing the same volume of DMSO as 0.1 μmol/L cucurbitacin I (DMSO group) , DMEM (negative control group) and 10 nmol/L calcipotriol (positive control group) , respectively. Cell counting kit-8 (CCK8) assay was performed to assess in vitro cellular proliferative activity after 12-, 24-, 36-hour treatment, reverse transcription (RT) -PCR to measure the mRNA expression of K17 and VEGF in HaCaT cells after 24-hour treatment, and Western blot analysis to determine the protein expression of K17, STAT3, phosphorylated-STAT3 (p-STAT3) and VEGF after 24-hour treatment. Statistical analysis was carried out by one-way analysis of variance (ANOVA) , repeated measures ANOVA, Student-Newman-Keuls (SNK) -q test and Pearson correlation analysis. Results The proliferative activity of HaCaT cells started to decrease after 12-hour treatment with cucurbitacin I at the concentration of 0.0125 μmol/L. When the concentration of cucurbitacin I increased up to 0.1 μmol/L, the cell proliferation rates were inhibited by 43.00% ± 2.11% and 48.98% ± 2.27% after 24-and 36-hour treatment respectively. Compared with the negative control group, cucurbitacin I at different concentrations all could inhibit in vitro proliferation of HaCaT cells (all P < 0.05) , and the inhibitory effects increased gradually with the increase of cucurbitacin I concentration and treatment duration. After 24-hour treatment, the mRNA expression of K17 and VEGF and the protein expression of K17, VEGF and P-STAT3 were significantly decreased along with the increase of concentration of cucurbitacin I (all P < 0.05) . Conclusion Cucurbitacin I can inhibit in vitro proliferation of HaCaT cells, and down-regulate the mRNA expression of K17 and VEGF and protein expression of K17, VEGF and P-STAT3. Key words: Cucurbitacins; Keratin-17; STAT3 transcription factor; Vascular endothelial growth factors; HaCaT cells

Cucurbitacin E reduces obesity and related metabolic dysfunction in mice by targeting JAK-STAT5 signaling pathway.

Several members of cucurbitaceae family have been reported to regulate growth of cancer by interfering with STAT3 signaling. In the present study, we investigated the unique role and molecular mechanism of cucurbitacins (Cucs) in reducing symptoms of metabolic syndrome in mice. Cucurbitacin E (CuE) was found to reduce adipogenesis in murine adipocytes. CuE treatment diminished hypertrophy of adipocytes, visceral obesity and lipogenesis gene expression in diet induced mice model of metabolic syndrome (MetS). CuE also ameliorated adipose tissue dysfunction by reducing hyperleptinemia and TNF-alpha levels and enhancing hypoadiponectinemia. Results show that CuE mediated these effects by attenuating Jenus kinase- Signal transducer and activator of transcription 5 (JAK- STAT5) signaling in visceral fat tissue. As a result, CuE treatment also reduced PPAR gamma expression. Glucose uptake enhanced in adipocytes after stimulation with CuE and insulin resistance diminished in mice treated with CuE, as reflected by reduced glucose intolerance and glucose stimulated insulin secretion. CuE restored insulin sensitivity indirectly by inhibiting JAK phosphorylation and improving AMPK activity. Consequently, insulin signaling was up-regulated in mice muscle. As CuE positively regulated adipose tissue function and suppressed visceral obesity, dyslipedemia, hyperglycemia and insulin resistance in mice model of MetS, we suggest that CuE can be used as novel approach to treat metabolic diseases.

Comparison between the effects of potassium phosphite and chitosan on changes in the concentration of Cucurbitacin E and on antibacterial property of Cucumis sativus

BackgroundCucurbitacins are mostly found in the members of the family Cucurbitaceae and are responsible for the bitter taste of cucumber. Pharmacological activities such as anti-bacterial and anti-tumor effects have been attributed to these structurally divers triterpens. The aim of this study was to investigate the effect of potassium phosphite (KPhi) and chitosan on Cucurbitacin E (CuE) concentration in different tissues of Cucumis sativus. The antibacterial effect of plant ethanolic extracts was also examined against E.coli PTCC 1399 and Pseudomonas aeruginosa PTCC 1430 bacterial strains.MethodsAfter emergence of secondary leaves, cucumber plants were divided into 4 groups (each group consisted of 6 pots and each pot contained one plant) and different treatments performed as follows: group1. Leaves were sprayed with distilled water (Control), group 2. The leaves were solely treated with potassium phosphite (KPhi), group 3. Leaves were solely sprayed with chitosan (Chitosan), group 4. Leaves were treated with KPhi and chitosan (KPhi + chitosan). The KPhi (2 g L−1) and chitosan (0.2 g L−1) were applied twice every 12 h for one day. Fruits, roots and leaves were harvested 24 h later. The ethanolic extract of plant organs was used for determination of CuE concentration using HPLC approach. The antimicrobial activity was evaluated by the agar well diffusion method. The experiments were arranged in a completely randomized design (CRD) and performed in six biological replications for each treatment. Analysis of variance was performed by one-way ANOVA and Dunnette multiple comparison using SPSS.ResultsThe highest level of CuE was recorded in fruit (2.2 g L−1) of plants under concomitant applications of KPhi and chitosan. Result of antibacterial activity evaluation showed that under concomitant treatments of KPhi and chitosan, fruit extract exhibited the highest potential for activity against E. coli PTCC 1399 (with mean zone of inhibition equal to 36 mm) and Pseudomonas aeruginosa PTCC 1430 (with mean zone of inhibition equal to 33 mm).ConclusionsKPhi and chitosan can induce production of CuE compound and increase antibacterial potential of cucumber plant extract. The application of KPhi and chitosan may be considered as promising prospect in the biotechnological production of CuE.

Inhibition of paclitaxel resistance and apoptosis induction by cucurbitacin B in ovarian carcinoma cells.

Ovarian cancer is the leading cause of mortality among all gynecological malignancies. Drug resistance is a cause of ovarian cancer recurrence and low rate of overall survival. There is a requirement for more effective treatment approaches. Cucurbitacin B (CuB) is an antineoplastic agent derived from traditional Chinese medicinal herbs. Its activity against paclitaxel-resistant human ovarian cancer cells has, however, not yet been established. The purpose of the present study was to investigate the effect and mechanism of CuB on human paclitaxel-resistant ovarian cancer A2780/Taxol cells. Cell viability was evaluated by a cell counting assay, while cell cycle arrest and apoptosis were assessed by microscopy and flow cytometry, and proteins associated with apoptotic pathways and drug resistance were evaluated by western blotting. The present results demonstrated that CuB exerts dose- and time-dependent cytotoxicity against the ovarian cancer A2780 cell line, with half-maximal inhibitory concentration (IC50) values 0.48, 0.25 and 0.21 µM following 24, 48 and 72 h of incubation, respectively. Compared with its sensitive counterpart, A2780, paclitaxel-resistant A2780/Taxol cells had almost identical IC50 values. Cell cycle analysis demonstrated that treatment with CuB may induce cell cycle arrest at the G2/M phase of the cell cycle in the two cell lines. As revealed by Annexin V/propidium iodide-labeled flow cytometry and Hoechst 33258 staining, CuB-induced apoptosis was accompanied by activation of caspase-3 and downregulation of B-cell lymphoma-2. Western blotting demonstrated that CuB may enhance the expression of p53 and p21 in the two cell lines. CuB may also downregulate the expression of P-glycoprotein. These results indicate that CuB may exert a therapeutic effect on paclitaxel-resistant human ovarian cancer.

Protective Effects of Cucurbitacin B on Acute Lung Injury Induced by Sepsis in Rats.

BackgroundThe aim of this study was to investigate the protective effects of cucurbitacin B (CuB) on sepsis-induced acute lung injury (ALI) in rats.Material/MethodsAn ALI model was made by cecal ligation and puncture (CLP) in SD rats. Rats were randomly divided into 5 groups (n=15 per group): animals undergoing a sham CLP (sham group); animals undergoing CLP (CLP control group); animals undergoing CLP and treated with CuB at 1 mg/kg of body weight (bw) (low-dose CuB [L-CuB] group), animals undergoing CuB at 2 mg/kg of bw (mid-dose CuB [M-CuB] group); and animals undergoing CuB at 5 mg/kg of bw (high-dose CuB [H-CuB] group). Samples of blood and lung tissue were harvested at different time points (6, 12, and 24 hour post-CLP surgery) for the detection of indicators which represented ALI. Five rats were respectively sacrificed at each time point. Pathological changes of lung tissue were observed by H&E staining. Another 50 rats were distributed into the same five groups to record the 72 hour survival rates.ResultsTreatment with CuB significantly increased the blood gas PaO2 levels and decreased lung wet/dry (W/D) ratio (p<0.05). It significantly reduced protein concentration, accumulation of the inflammatory cells, and levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), (p<0.05), in the bronchoalveolar lavage fluid (BALF). Pulmonary pathological damage and survival rates at 72 hours were found to be effectively improved by CuB. In addition, CuB performed its pulmonary protection effects in a dose-depended manner.ConclusionsCuB can effectively improve the pulmonary gas exchange function, reduce pulmonary edema, and inhibit the inflammatory response in the lung, revealing that CuB may serve as a potential therapeutic strategy for sepsis-induced ALI.

CucurbitacinB induces autophagy and apoptosis by suppressing CIP2A/PP2A/mTORC1 signaling axis in human cisplatin resistant gastric cancer cells.

Cancerous inhibitor of protein phosphatase2A (CIP2A) is a human oncoprotein that is overexpressed in multiple kinds of tumors including gastric cancer(GC). Mammalian target of rapamycin complex1 (mTORC1) over-activation is detected in GC and many other cancers. Previous study found that CIP2A/mTORC1 controls cell growth and autophagy through direct association. CIP2A plays an 'oncogenic nexus' in several cancer types to participate in the tumorigenic transformation and chemoresistance. In the present study, we investigated whether CucurbitacinB(CuB), a natural compound found in Cucurbitaceae, can be used in cisplatin (DDP)-resistant human GC cell line SGC7901/DDP. Results demonstrated that CuB treatment significantly suppressed SGC7901/DDP cell proliferation, induced caspase-dependent apoptosis, and autophagy. The activation of autophagy was mediated through CuB-induced inhibition of mTORC1. Furthermore, CuB inhibited mTORC1 via the activation of protein phosphatase2A(PP2A) which is mediated by CIP2A inhibition. These findings indicated that CuB can inhibit the proliferation, induce caspase-dependent apoptosis, and autophagy of SGC7901/DDP cells by suppressing CIP2A/PP2A/mTORC1 signaling axis. Thus, CuB may be a novel effective candidate to treat DDP-resistant human GC cells.

Cucurbitacin E inhibits osteosarcoma cells proliferation and invasion through attenuation of PI3K/AKT/mTOR signalling pathway.

Cucurbitacin E (CuE), a potent member of triterpenoid family isolated from plants, has been confirmed as an antitumour agent by inhibiting proliferation, migration and metastasis in diverse cancer. However, the effects and mechanisms of CuE on osteosarcoma (OS) have not been well understood. The present study aimed to test whether CuE could inhibit growth and invasion of OS cells and reveal its underlying molecular mechanism. After various concentrations of CuE treatment, the anti-proliferative effect of CuE was assessed using the cell counting Kit-8 assay. Flow cytometry analysis was employed to measure apoptosis of OS cells. Cell cycle distribution was analysed by propidium iodide staining. Transwell assay was performed to evaluate the effect of CuE on invasion potential of OS cells. The protein levels were measured by western blot. In addition, the potency of CuE on OS cells growth inhibition was assessed in vivo. Our results showed that CuE inhibited cell growth and invasion, induced a cell cycle arrest and triggered apoptosis and modulated the expression of cell growth, cell cycle and cell apoptosis regulators. Moreover, CuE inhibited the PI3K/Akt/mTOR pathway and epithelial–mesenchymal transition (EMT), which suppressed the invasion and metastasis of OS. In addition, we also found that CuE inhibited OS cell growth in vivo. Taken together, our study demonstrated that CuE could inhibit OS tumour growth and invasion through inhibiting the PI3K/Akt/mTOR signalling pathway. Our findings suggest that CuE can be considered to be a promising anti-cancer agent for OS.

Cucurbitacin B exerts anti-cancer activities in human multiple myeloma cellsin vitroandin vivoby modulating multiple cellular pathways

Cucurbitacin B (CuB), a triterpenoid compound isolated from the stems of Cucumis melo, has long been used to treat hepatitis and hepatoma in China. Although its remarkable anti-cancer activities have been reported, the mechanism by which it achieves this therapeutic activity remains unclear. This study was designed to investigate the molecular mechanisms by which CuB inhibits cancer cell proliferation. Our results indicate that CuB is a novel inhibitor of Aurora A in multiple myeloma (MM) cells, arresting cells in the G2/M phase. CuB also inhibited IL-10-induced STAT3 phosphorylation, synergistically increasing the anti-tumor activity of Adriamycin in vitro. CuB induced dephosphorylation of cofilin, resulting in the loss of mitochondrial membrane potential, release of cytochrome c, and activation of caspase-8. CuB inhibited MM tumor growth in a murine MM model, without host toxicity. In conclusion, these results indicate that CuB interferes with multiple cellular pathways in MM cells. CuB thus represents a promising therapeutic tool for the treatment of MM.

Cucurbitacin B reverses multidrug resistance by targeting CIP2A to reactivate protein phosphatase 2A in MCF-7/adriamycin cells.

Cancerous inhibitor of protein phosphatase 2A (CIP2A) is a human oncoprotein that is overexpressed in various tumors. A previous study found that CIP2A expression is associated with doxorubicin (Dox) resistance. In the present study, we investigated whether cucurbitacin B (CuB), a natural anticancer compound found in Cucurbitaceae, reversed multidrug resistance (MDR) and downregulated CIP2A expression in MCF-7/Adriamycin (MCF-7/Adr) cells, a human breast multidrug-resistant cancer cell line. CuB treatment significantly suppressed MCF-7/Adr cell proliferation, and reversed Dox resistance. CuB treatment also induced caspase-dependent apoptosis, decreased phosphorylation of Akt (pAkt). The suppression of pAkt was mediated through CuB-induced activation of protein phosphatase 2A (PP2A). Furthermore, CuB activated PP2A through the suppression of CIP2A. Silencing CIP2A enhanced CuB-induced growth inhibition, apoptosis and MDR inhibition in MCF-7/Adr cells. In conclusion, we found that enhancement of PP2A activity by inhibition of CIP2A promotes the reversal of MDR induced by CuB.

Cucurbitacin B inhibits breast cancer metastasis and angiogenesis through VEGF-mediated suppression of FAK/MMP-9 signaling axis

Available breast cancer therapeutic strategies largely target the primary tumor but are ineffective against tumor metastasis and angiogenesis. In our current study, we determined the effect of Cucurbitacin B (CuB), a plant triterpenoid, on the metastatic and angiogenic potential of breast cancer cells. CuB was found to inhibit cellular proliferation and induce apoptosis in breast cancer cells in a time- and dose-dependent manner. Further, CuB-treatment significantly inhibited the migratory and invasive potential of highly metastatic breast cancer MDA-MB-231 and 4T1 cells at sub-IC50 concentrations, where no significant apoptosis was observed. CuB was also found to inhibit migratory, invasive and tube-forming capacities of HUVECs in vitro. In addition, inhibition of pre-existing vasculature in chick embryo chorioallantoic membrane ex vivo further supports the anti-angiogenic effect of CuB. CuB-mediated anti-metastatic and anti-angiogenic effects were associated with the downregulation of VEGF/FAK/MMP-9 signaling, which has been validated by using FAK-inhibitor (FI-14). CuB-treatment resulted in a significant inhibition of VEGF-induced phosphorylation of FAK and MMP-9 expressions similar to the action of FI-14. CuB was also found to decrease the micro-vessel density as evidenced by the decreased expression of CD31, a marker for neovasculature. Further, CuB-treatment inhibited tumor growth, lung metastasis and angiogenesis in a highly metastatic 4T1-syngeneic mouse mammary cancer. Collectively, our findings suggest that CuB inhibited breast cancer metastasis and angiogenesis, at least in part, through the downregulation of VEGF/FAK/MMP-9 signaling.

A biotransformation process for the production of cucurbitacin B from its glycoside using a selected Streptomyces sp.

Cucurbitacin B (CuB) and its glycoside, cucurbitacin B 2-o-β-D-glucoside (CuBg), abundantly occur in the pedicels of Cucumis melo. Compared with CuB, CuBg is not efficiently extracted from the pedicels. Furthermore, the anticancer activity of CuBg is lower than that of the aglycone. A process for CuBg biotransformation to CuB was developed for the first time. A strain of Streptomyces species that converts CuBg into CuB was isolated from an enrichment culture of C. melo pedicels. After optimization of conditions for enzyme production and biotransformation, a maximum conversion rate of 92.6% was obtained at a CuBg concentration of 0.25g/L. When biotransformation was performed on C. melo pedicel extracts, the CuB concentration in the extracts increased from 1.50 to 3.27g/L. The conversion rate was almost 100%. The developed process may be an effective biotransformation method for industrial production CuB from C. melo pedicels for pharmaceuticals.

Cucurbitacin E induces caspase-dependent apoptosis and protective autophagy mediated by ROS in lung cancer cells

Cucurbitacin E (CuE) is a triterpenoid with potent anticancer activities while the underlying mechanisms remain elusive. In the present study, the anticancer effects of CuE on 95D lung cancer cells were investigated. CuE decreased cell viability, inhibited colony formation, and increased reactive oxygen species (ROS) in a concentration-dependent manner, which were reversed by N-acetyl-l-cysteine (NAC). CuE induced apoptosis as determined by JC-1 staining, expression of Bcl-2 family proteins, cleavage of caspases, and TUNEL staining. NAC and Ac-DEVD-CHO partially reversed CuE-induced cleavage of caspase-3, caspase-7, and PARP. Furthermore, CuE caused accumulation of autophagic vacuoles and concentration- and time-dependent expression of LC3II protein. Autophagy inhibitors chloroquine and bafilomycin A1 enhanced CuE-induced LC3II expression and cell death. CuE-triggered protein expression of p-AKT, p-mTOR, Beclin-1, and p-ULK1 was partially reversed by NAC pretreatment. In addition, CuE treatment damaged F-actin without affecting β-tubulin as confirmed by immunofluorescence. In conclusion, CuE induced ROS-dependent apoptosis through Bcl-2 family and caspases in 95D lung cancer cells. Furthermore, CuE induced protective autophagy mediated by ROS through AKT/mTOR pathway. This study provides novel roles of ROS in the anticancer effect of CuE.

Cucurbitacin B Enhances the Anticancer Effect of Imatinib Mesylate Through Inhibition of MMP-2 Expression in MCF-7 and SW480 Tumor Cell Lines.

The combination of medicinal plant extracts with known chemotherapeutics offers significant potential for the development of novel therapies in cancer disease. Cucurbitacin B (CuB) is one of the most potent and widely used members of cucurbitacin family and it is known to have important effects on several diseases including cancer. To determine whether CuB can enhance chemosensitivity to imatinib mesylate (IM), in the present study, the combined effects of CuB with IM on MCF-7 and SW480 cells were investigated. The cells were treated with CuB alone or in combination with IM and the results showed that the combination treatment synergistically inhibited cell proliferation and induced apoptosis. Furthermore, the combined effect of CuB and IM on matrix metalloproteinase-2 (MMP-2) gene expression, a member of MMP family which is responsible for the degradation of extracellular matrix was also evaluated. CuB increased the inhibitory effect of IM on MMP-2 expression synergistically in a dose dependent manner. The results suggest that CuB in combination with IM may serve as a potentially useful therapeutic strategy for patients with breast and colorectal cancer.

Diffusivity of Cucurbitacin B in water swollen polyethylene oxide-b-polycaprolactone matrices with different PCL/PEO weight ratios

We used isobaric–isothermal molecular dynamics simulation to study the diffusivity of a hydrophobic drug Cucurbitacin B (CuB) in polymer matrices made up of a single polyethylene oxide-b-polycaprolactone (PEO-b-PCL) chain and different numbers of water molecules at body temperature (310K) and ambient pressure (1bar). Four PEO-b-PCL models were used. Three of them had the same total molecular weight of 3750g/mol but different PCL/PEO weight ratios at 0.5, 1 and 2. The remaining model had a total molecular weight of 5000g/mol and a PCL/PEO weight ratio of 1. The data showed that self-diffusion coefficients of CuB decreased with increasing PCL/PEO weight ratio and was insensitive to the water concentration at fixed PCL/PEO weight ratios. The first observation was attributed to a high number of hydrogen bonds formed between the oxygen atoms on CuB and the activated hydrogen atoms on the PCL block as the PCL/PEO weight ratio increased. It was found that degree of swelling and effects of the mobility of the PEO and PCL blocks on the diffusivity of CuB were minor and were overshadowed by the PCL block-CuB hydrogen bonds.

Ethanol enhances cucurbitacin B-induced apoptosis by inhibiting cucurbitacin B-induced autophagy in LO2 hepatocytes

Ethanol is a common risk factor for liver injury. Cucurbitacin B (CuB) is a natural product with potent cytotoxic activities mediated by inducing apoptosis. This study investigated the effect of ethanol on CuB-induced cytotoxicity in LO2 hepatocytes. Low concentration of ethanol alone showed no significant cytotoxic effect on LO2 cells. CuB dose-dependently decreased cell viability. However, ethanol co-treatment significantly enhanced CuB-induced cytotoxicity. CuB-induced mitochondria membrane potential (ΔΨ) depolarization was further decreased by ethanol. Furthermore, CuB-induced apoptosis was augmented by ethanol, as evidenced by DNA fragmentation, Annexin V staining, and apoptotic protein expression. Ethanol inhibited CuB-induced autophagy, as determined by MDC staining, autophagic protein expression, and transmission electron microscopy. Therefore, the present data suggested that ethanol enhanced CuB cytotoxicity in LO2 hepatocytes, which was mediated by inhibiting autophagy and augmenting apoptosis.

Cucurbitacin B inhibits the stemness and metastatic abilities of NSCLC via downregulation of canonical Wnt/β-catenin signaling axis.

Lack of effective anti-metastatic drugs creates a major hurdle for metastatic lung cancer therapy. For successful lung cancer treatment, there is a strong need of newer therapeutics with metastasis-inhibitory potential. In the present study, we determined the anti-metastatic and anti-angiogenic potential of a natural plant triterpenoid, Cucurbitacin B (CuB) against non-small cell lung cancer (NSCLC) both in vitro and in vivo. CuB demonstrated a strong anti-migratory and anti-invasive ability against metastatic NSCLC at nanomolar concentrations. CuB also showed significant tumor angiogenesis-inhibitory effects as evidenced by the inhibition of migratory, invasive and tube-forming capacities of human umbilical vein endothelial cells. CuB-mediated inhibition of angiogenesis was validated by the inhibition of pre-existing vasculature in chick embryo chorio-allantoic membrane and matrigel plugs. Similarly, CuB inhibited the migratory behavior of TGF-β1-induced experimental EMT model. The CuB-mediated inhibition of metastasis and angiogenesis was attributable to the downregulation of Wnt/β-catenin signaling axis, validated by siRNA-knockdown of Wnt3 and Wnt3a. The CuB-mediated downregulation of Wnt/β-catenin signaling was also validated using 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced lung tumorigenesis model in vivo. Collectively, our findings suggest that CuB inhibited the metastatic abilities of NSCLC through the inhibition of Wnt/β-catenin signaling axis.

Cytotoxic effects of natural and semisynthetic cucurbitacins on lung cancer cell line A549.

Cucurbitacins and their derivatives are triterpenoids that are found in various plant families, and are known for their pharmacological and biological activities, including anti-cancer effects. Lung cancer represents a major public health problem, with non-small-cell lung cancer (NSCLC) being the most frequent and aggressive type of lung cancer. The objective of this work was to evaluate four cucurbitacins (CUCs) for their cytotoxic activity, effects on apoptosis induction, cell cycle progression, anti-migratory, and anti-invasive effects on the human NSCLC cell line (A549 cells). Our findings showed that these CUCs could suppress human NSCLC cell growth in vitro through their effects on the PI3Kinase and MAPK pathways, which lead to programmed cell death induction, as well as inhibition of cell migration and cell invasion. Additionally, these effects culminate in apoptosis induction and G2/M cell cycle arrest by modulating cyclin B1 expression, and in the mitigation of strategic steps of lung cancer metastasis, including migration and invasion of A549 cells. These results suggest that two natural (DDCB and CB) and two novel semisynthetic derivatives of cucurbitacin B (ACB and DBCB) could be considered as promising compounds with antitumor potential.