Dox In Cells Research Articles

The translation of fundamental research discoveries into cell and gene based medicinal products

Efficient intracellular delivery of exogenous materials remains a critical issue in fundamental biological researches and clinical applications. Here, we developed a novel chemical-free method for intracellular delivery enhancement using a designed gigahertz ultrasonic electromechanical resonator. When excited by a sinusoidal electric signal, the propagation and attenuation of acoustic wave in liquid will generate high-speed acoustic streaming. The liquid above the device working area will be accelerated and strike the substrate surface, thus generates pressure on cells, induces deformation and membrane poration, and finally realizes delivery of exogenous materials. To verify the intracellular delivery ability, DOX was selected as an example, and an enhanced fluorescence of DOX in cells exposed to resonator stimulation can be seen. We also realized the delivery of fluorescent-labeled DNA strains and plasmids. Besides, different power applied to the resonator can induce different fluid velocity, thus generate different force intensity and control the deliver efficiency. Pores on membranes induced by acoustic streaming treatment were observed by SEM. Disrupted cell membranes and porous structures can be seen after treatment, and resealed after 10 min recovery, indicating a strong fluid force exerted on cells and the influence is temporary and reversible.

Long Non-Coding RNA <i>ENST00000508435</i> Promotes Breast Cancer Chemoresistance Through Pyroptosis Inhibition

Chemotherapeutic resistance is a significant issue in the management of breast cancer patients, and the involvement of pyroptosis, a type of inflammatory cell necrosis, in cancer has recently been proposed. Here we explored the role of lncRNA ENS0508435 in pyroptosis and tumor chemoresistance. Our results showed that the ENS0508435 expression was significantly upregulated in doxorubicin-resistant MCF-7 cells (MCF-7/DOX) and that ENS0508435 knockdown decreased the resistance of MCF-7/DOX cells to doxorubicin. Moreover, ENS0508435 overexpression could inhibit doxorubicin-induced pyroptosis by reducing the expression of NLRP3, ASC, caspase-1, and GSDMD. Further evaluation of the mechanism demonstrated that ENS0508435 directly bound to hnRNPU, causing it to recruit EZH2, which repressed TRAF6 gene expression. Thus, TRAF6 was unavailable to combine with MALT1 and caspase-1, which attenuated pyroptosis. Altogether, the results indicated that ENS0508435 promoted breast cancer chemoresistance and was involved in pyroptosis inhibition through the hnRNPU-EZH2-TARF6-MALT1-caspase 1 pathway.

A nano-complex system to overcome antagonistic photo-chemo combination cancer therapy

Photo-Chemo combination therapy has been intensively investigated for treatment of cancers, especially multidrug resistance cancer. However, antagonistic interactions between chemo-drugs and photosensitizers are frequently reported, and drugs doses and treatment sequences have been changed to overcome the problems. We observed the antagonistic effect by a decrease in singlet oxygen generation from the photosensitizer when Dox was in close physical proximity. To control the distance between Dox and the photosensitizer, we developed a novel pH-sensitive poly ionomer complex system composed of PEG-PLL(-g-Ce6) [Chlorin e6 grafted poly(ethylene glycol)-poly(l-lysine)] and PEG-PLL(-g-DMA)-PLA [2,3-dimethylmaleic anhydride grafted poly(ethylene glycol)-poly(l-lysine)-poly(lactic acid)] and evaluated this system with regard to singlet oxygen generation and antiproliferative activity against MCF-7/Dox cells. Enhanced singlet oxygen generation and antiproliferative activities were observed in vitro and in vivo for the poly ionomer complex system compared to PEG-PLL(-g-Ce6)-PLA/Dox due to the change in distance between Dox and Ce6 in the PIC system under acidic conditions. Our results highlight the importance of interactions between co-loaded drugs in combination therapy, and provide new insights into design principles for tailor-made nanomedicine platforms.

Down-Regulation of MiR-125b Reverses Drug Resistance of Doxorubicin-Resistant Leukemia Cells

To investigate whether the down-regulation of miR-125b can reverse the drug-resistence of doxorubicine-resistant leukemia cell lines or not, so as to explore a new method for treatment of drug-resistant leukemia patients. The expression levels of miR125b in doxorubicine drug-sensitive and doxorubicine drug-resistant leukemia cell lines.HL-60, K562 and HL-60/Dox, the K562/Dox were detected by using RT-qPCR; the up-regulation or inhibition of miR-1256 expression in HL-60/Dox were performed by electroporation transfection, then the viability of cells treated with doxorubicine of different concentration was detected by CCK-8 method, the proliferation inhibition curve of cells was drawed, and the IC50 was calculated. The miR-125b expression was obviously up-regulated in drug-resistant cell lines HL-60/DOX and K562/DOX, as compared with HL-60 and K562 cell lines. The miR-125b expression level in HL-60/DOX and K562/DOX cells was 15 times and 5 times higher than that in HL-60 and K562 cells, respectively. The up-regulating or inhibiting expression of miR-125b in HL-60/DOX cells found that the proliferation inhibition rate in cells transfected with miR-125b mimic significantly decreased, compared with control group (P<0.01), while the proliferation inhibition rate in cells transfected with miR-125b inhibitor significantly increased, compared with control group(P<0.01). The miR-125b expression in HL-60/Dox and K562/Dox cells has been up-regulated, down-regulation of miR-125b expression can reverse the drug resistance of leukemia cells to doxorubicine.

Rhein reverses doxorubicin resistance in SMMC-7721 liver cancer cells by inhibiting energy metabolism and inducing mitochondrial permeability transition pore opening.

Rhein, a monomeric anthraquinone obtained from the plant herb species Polygonum multiflorum and P. cuspidatum, has been proposed to have anticancer activity. This activity has been suggested to be associated with mitochondrial injury due to the induction of mitochondrial permeability transition pore (mPTP) opening. In this study, the effects of 5-80 μM rhein on cell viability, half-maximal inhibitory concentration (IC50 value), resistance index, and apoptosis were assessed in the liver cancer cell lines SMMC-7721 and SMMC-7721/DOX (doxorubicin-resistant cells). Rhein (10-80 μM) significantly reduced the viability of both cell lines; 20 μM rhein significantly increased sensitivity to DOX and increased apoptosis in SMMC-7721 cells, but reversed resistance to DOX by 7.24-fold in SMMC-7721/DOX cells. Treatment with rhein increased accumulation of DOX in SMMC-7721/DOX cells, inhibited mitochondrial energy metabolism, decreased cellular ATP, and ADP levels, and altered the ratio of ATP to ADP. These effects may result from the binding of rhein with voltage-dependent ion channels (VDACs), adenine nucleotide translocase (ANT), and cyclophilin D, affecting their function and leading to the inhibition of ATP transport by VDACs and ANT. ATP synthesis was greatly reduced and mitochondrial inner membrane potential decreased. Together, these results indicate that rhein could reverse drug resistance in SMMC-7721/DOX cells by inhibiting energy metabolism and inducing mPTP opening. © 2018 BioFactors, 45(1):85-96, 2019.

Synthesis and In Vitro Anti‐Tumor, Anti‐Mycobacterial and Anti‐HIV Activities of Diethylene‐Glycol‐Tethered Bis‐Isatin Derivatives

AbstractA series of diethylene glycol tethered bis‐istain derivatives 4 a‐o were designed, synthesized and screened for their biological activities in this paper. Eight bis‐isatin derivatives displayed broad‐spectrum activities against all tested drug‐sensitive and drug‐resistant cancer cells with IC50 in a range of 8.32 to 49.73 μM. The most active derivative 4 d was &gt;2.5 folds more potent than etoposide against Hela, HCT‐116, A549 and drug‐resistant MCF‐7/DOX (Doxorubicin‐resistant MCF‐7) cells and was comparable to etoposide against DU145, SKOV3 and MCF‐7. Tubulin inhibitory results suggested that this kind of bis‐istain derivatives could exert their anti‐cancer activities through tubulin inhibition. The eight bis‐isatin derivatives with high anti‐tumor activity were selected for further screen to investigate their anti‐mycobacterial and anti‐HIV activities, but the majority of them only exhibited weak to moderate activities.

MiR-433 reverses chemoresistance to docetaxel by targeting Notch1 in breast cancer cells

To investigate the role of miR- 433 in chemoresistance to docetaxel in breast cancer cells. A docetaxel-resistant MCF-7 breast cancer cell line (MCF-7/DOX) was established by exposure of parental MCF-7 cells to progressively increased docetaxel concentrations. The functional role of miR-433 was investigated by assessing the changes in viability and apoptosis of the cells transfected with a miR-433 inhibitor or a miR-433 mimics. The downstream targets of miR- 433 were determined by bioinformatics analysis, cell transfection and luciferase reporter assay. Quantitative real-time PCR analysis showed that miR- 433 was down-regulated in MCF-7/DOX cells. Transfection of the cells with the miR-433 inhibitor obviously enhanced chemoresistance to docetaxel and attenuated cell apoptosis in MCF-7 cells; miR-433 overexpression significantly increased the sensitivity to docetaxel and promoted apoptosis in MCF- 7/DOX cells. Luciferase reporter assay showed that the down-regulation miR-433 expression was associated with significantly increased expressions of Notch1 at both mRNA and protein levels in MCF-7 cells. Compared with the control cells, McF-7/DOX cells transfected with miR-433 mimics exhibited significantly decreased mRNA and protein expressions of Notch1. miR-433 may reverse chemoresistance to docetaxel by targeting Notch1 in breast cancer cells.

Abstract 3708: Targeted delivery of doxorubicin to glioblastomas by thermally sensitive polypeptides

Abstract Glioblastoma (GBM) ranks among the most common, aggressive, and least curable cancers due to a strong tendency for intracranial dissemination, high proliferation potential and inherent tumor resistance to radiation or chemotherapy. Current treatments face the critical challenge of adverse treatment effects in normal tissue within the brain itself and also when the agent cannot adequately penetrate the blood brain barrier (BBB) to reach the tumor microenvironment. We have developed a system to selectively deliver chemotherapeutic doxorubicin to GBM tumors. This carrier is based on elastin-like polypeptide (ELP), which is soluble at physiological temperatures but undergoes a phase transition and accumulates at tumor sites with externally applied, mild (40-41°C) hyperthermia. The CPP-ELP-Dox conjugate consists of a cell penetrating peptide (CPP), which facilitates transcytosis through the BBB and cell entry, and the 6-maleimidocaproyl hydrazone derivative of doxorubicin (Dox) at the C-terminus of ELP. The acid-sensitive hydrazone linker ensures release of Dox in the lysosomes/endosomes after cellular uptake of the drug conjugate. We have shown that CPP-ELP-Dox effectively inhibits cell proliferation in three GBM cell lines; GBM6, D54, and U251 with IC50 values of 250 nM, 60nM, and 30nM, respectively. Both the free drug and CPP-ELP-Dox conjugate exhibited similar in vitro cytotoxicity, although their subcellular localization was considerably different. The Dox conjugate was mainly dispersed in the cytoplasm, while free drug was largely dispersed in the cytoplasm but also had partial nuclear accumulation. The accumulation of free Dox in GBM cells was measured by flow cytometry to determine whether the cytotoxic activity of free Dox or CPP- ELP-Dox is related to intracellular Dox levels. Intracellular Dox concentration was increased in the CPP-ELP-Dox cells compared to that in the free Dox cells, which positively correlates with cytotoxic activity. Flow cytometry was also used to quantitatively investigate apoptosis of GBM cells. After incubation for 24 h, the total apoptosis percentage of free Dox was 1.5% in GBM6, 6% in D54, and 7% in U251 cells. However, the total apoptosis rates of CPP-ELP-Dox treated cells were 5%, 12% and 5% in GBM6, D54, and U251 cells respectively, which confirms increased apoptosis compared to free Dox treated cells. Cell cycle analysis of GBM cells treated with free Dox showed enhanced arrest of the cells in G1 phase, while there was increased percentage arrest of cells in G2 phase in cells treated with CPP-ELP-Dox.In summary, our findings demonstrate that CPP-ELP-Dox effectively kills GBM cells. Development of such a drug carrier has the potential to greatly improve current therapeutic approaches for GBM by increasing the specificity and efficacy of treatment and reducing cytotoxicity in normal tissues. Citation Format: Sonja Dragojevic, Rebecca B. Mackey, Felix Kratz, Drazen Raucher. Targeted delivery of doxorubicin to glioblastomas by thermally sensitive polypeptides [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3708.

Catechin-conjugated mesoporous hydroxyapatite nanoparticle: A novel nano-antioxidant with enhanced osteogenic property.

Hydroxyapatite is the main component of mineral phase of bone which is widely employed for coating metal implants and scaffold materials in synthetic bone grafts owing to its osteoinductive property. In order to improve the bioactivity of hydroxyapatite, mesoporous hydroxyapatite nanoparticles (MHAP) were synthesized and chemically functionalized with 3-aminopropyltriethoxysilane. The amine-functionalized nanoparticles were conjugated with a natural antioxidant, catechin (Cat), through a stable amide linkage. The true structure of the bioconstruct was confirmed by calculating condensed Fukui indices. The functionalized-hydroxyapatite nanoparticles (Cat@MHAP) showed an outstanding antioxidant activity, having reactivity toward hydroxyl and superoxide radicals larger than that of free catechin. To explore the bone cell responses to this material, multilayer nanoparticle films were prepared by MHAP and Cat@MHAP on a glass substrate. Afterward, the short- and long-term responses of cultured mesenchymal stem cells (MSCs), osteosarcoma cells (Saos-2), and doxorubicin-resistant cells (RSaos-2/Dox) on the surface of the prepared films were investigated. Both the MSCs and bone tumor cells selectively adhered onto Cat@MHAP surface as compared with glass and MHAP at initial culture time. Moreover, it was found that Cat@MHAP decreases the proliferation of Saos-2 and RSaos-2/Dox cells in a time-dependent manner, while it supports the growth of MSCs, indicating the ability of Cat@MHAP to distinguish tumor cells from normal ones. Further, Cat@MHAP promotes the osteogenic differentiation in both the MSCs and tumor cells, accompanied by the attenuation of intracellular ROS. From these results, Cat@MHAP is a novel "nano-antioxidant," which could be considered as a promising biomaterial in treating bone defects, particularly after surgery in osteosarcoma patients.

Trijugin- and mexicanolide-type limonoids from the fruits of Heynea trijuga that reverse multidrug resistance in MCF-7/DOX cells

Eleven previously undescribed limonoids, trichisins A-K, including eight structural analogues A-H of trijugin and three H-J mexicanolide derivatives, together with two known mexicanolide derivatives were isolated from the fruits of Heynea trijuga Roxb. ex Sims. The structure determination was based on extensive physical data analyses (NMR, MS), and their basic skeletons and the absolute configurations of trichisins A, B, E, K and trichiconnarone A were assigned via X-ray crystallographic analysis (Cu Kα radiation). The hemiketal motifs in trijugins A, B, and E-G are rare in limonoids. Bioactivity screenings suggested that the trijugin H and mexicanolide-type trichiconnarones A and B limonoids were effective in reversing resistance in MCF-7/DOX cells at a nontoxic concentration of 50 μM with IC50 values of 12.45, 10.86, and 14.96 μM, respectively.

Quercetin reversed MDR in breast cancer cells through down-regulating P-gp expression and eliminating cancer stem cells mediated by YB-1 nuclear translocation.

Overexpression of P-glycoprotein (P-gp) plays an important role in mediating multidrug resistance (MDR), resulting in chemotherapy failure of tumor patients and enhancement of cancer stem cell characteristics. By preparing doxorubicin (Dox) resistant human breast cancer MCF-7 cells, here, we wanted to evaluate the effects of quercetin (Que) on MDR reversal activity and investigate its possible mechanism. MCF-7 and MCF-7/dox cells were respectively treated by Dox, paclitaxel (Pac), or vincristine (Vcr) with or without Que intervention for 24hr. Cell viability, cell apoptosis, cell cycle, intracellular drug accumulation, the expression of P-gp and Y-box binding protein 1 (YB-1), and breast cancer stem cells (BCSCs) were then assessed. The results showed that Que significantly enhanced the antitumor activities of Dox, Pac, and Vcr in breast cancer cells. In addition, combined treatment of Dox, Pac, or Vcr with Que significantly downregulated P-gp expression and eliminated BCSCs. Furthermore, combined treatment of Dox, Pac, or Vcr with Que significantly inhibited nuclear translocation of YB-1. Thus, we speculated that Que reversed MDR in breast cancer cells through downregulating P-gp expression and eliminating cancer stem cells mediated by YB-1 nuclear translocation.

Down-regulation of miR-210-3p encourages chemotherapy resistance of renal cell carcinoma via modulating ABCC1

BackgroundATP-binding cassette transporter super-family including ABCC1 and MDR-1 were involved in multi-drug resistance (MDR) of renal cell carcinoma (RCC) patients. Several miRNAs were confirmed to promote the MDR and the survival of tumor cells.MethodsThe RCC cell lines Caki-2 with vinblastine-resistant (Caki-2/VBL) or doxorubicin-resistant (Caki-2/DOX) were constructed, respectively. The expressions of miR-210-3p, ABCC1 and MDR-1 protein were determined by qRT-PCR and Western blot assays. The viability of RCC cells was assessed by MTT assay. The regulatory relationship between miR-210-3p and ABCC1 was analyzed by Dual Luciferase assay. The effect of miR-210-3p in vivo was investigated with a tumor xenograft model in mice.ResultsMiR-210-3p expression was observed to significantly decrease in Caki-2/VBL and Caki-2/DOX cells. Meanwhile, ABCC1 and MDR-1 were significantly increased in Caki-2/VBL and Caki-2/DOX cells. ABCC1 was a novel target of miR-210-3p and negatively regulated by miR-210-3p. And miR-210-3p improved drug-sensitivity of RCC cells. Down-regulation of ABCC1 could reverse the effect of miR-210-3p knockdown on the drug-resistance and the level of MDR-1 in drug-sensitive RCC cells.ConclusionWe confirmed that down-regulation of miR-210-3p increased ABCC1 expression, thereby enhancing the MRP-1-mediated multidrug resistance of RCC cells.

Reversal of multidrug resistance by icaritin in doxorubicin-resistant human osteosarcoma cells

Multidrug resistance (MDR) is one of the major obstacles in cancer chemotherapy. Our previous study has shown that icariin could reverse MDR in MG-63 doxorubicin-resistant (MG-63/DOX) cells. It is reported that icariin is usually metabolized to icariside II and icaritin. Herein, we investigated the effects of icariin, icariside II, and icaritin (ICT) on reversing MDR in MG-63/DOX cells. Among these compounds, ICT exhibited strongest effect and showed no obvious cytotoxicity effect on both MG-63 and MG-63/DOX cells ranging from 1 to 10 μmol·L−1. Furthermore, ICT increased accumulation of rhodamine 123 and 6-carboxyfluorescein diacetate and enhanced DOX-induced apoptosis in MG-63/DOX cells in a dose-dependent manner. Further studies demonstrated that ICT decreased the mRNA and protein levels of multidrug resistance protein 1 (MDR1) and multidrug resistance-associated protein 1 (MRP1). We also verified that blockade of STAT3 phosphorylation was involved in the reversal effect of multidrug resistance in MG-63/DOX cells. Taken together, these results indicated that ICT may be a potential candidate in chemotherapy for osteosarcoma.

CYTOTOXIC ACTIVITY OF BRAZILEIN ISOLATED FROM SECANG (CAESALPINIA SAPPAN L.) AGAINST MCF7/DOX CELLS BY INHIBITION OF P-GLYCOPROTEIN

Objective: This study was focused on isolation of brazilein from the dried heartwood of Secang (Caesalpinia sappan L.) followed by its characterization using Infrared (IR) spectroscopy, liquid chromatography-mass spectrometry (LC-MS) with electrospray ionization (ESI), proton (1H), carbon-13 (13C) nuclear magnetic resonance (NMR) and two dimensional (2D)-NMR, evaluation the cytotoxic activity of brazilein in MCF-7 resistant doxorubicin (MCF-7/DOX) cells and evaluate the interaction between brazilein and ATP with P-glycoprotein (Pgp) in silico using molecular docking.Methods: Brazilein was isolated and purified from ethyl acetate fraction by flash silica gel column chromatography, eluting with chloroform, ethyl acetate and methanol in gradient concentration. In the cytotoxicity assay, MCF-7/DOX cells were cultured in the presence of brazilein for 24 hour (h) and cell viability was evaluated by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Interactions between brazilein and the target proteins were evaluated and calculated in silico by molecular docking using PLANTS.Results: The infrared, mass spectra with a molecular weight of 284 and NMR signal confirmed that was brazilein. MTT assay showed a dose-dependent inhibition of MCF-7/DOX cell proliferation with brazilein IC50 value of 43 µM. The docking score of brazilein was-71,45 kcal/mol and ATP value-96,23 kcal/mol.Conclusion: Brazilein has a potent cytotoxic value on MCF-7/DOX and high affinity in Pgp protein target. Brazilein can be developed as anticancer especially in cancer resistance incidence. Further study must be established to evaluate the molecular mechanism of brazilein inhibit MCF-7/DOX cell proliferation in vitro.

TanshinoneIIA enhances the chemosensitivity of breast cancer cells to doxorubicin through down-regulating the expression of MDR-related ABC transporters

As the first-line drug for breast cancer chemotherapy, doxorubicin (Dox) has strong cardiotoxicity. Meanwhile, prolonged Dox treatment of patients with breast cancer may result in resistance of breast cancer cells to Dox and an increased number of Dox-resistant breast cancer stem cells (BCSCs), thereby easily leading to breast cancer relapse. TanshinoneIIA (Tan IIA) has anti-tumor activity in addition to its cardiovascular protective effect. By preparing Dox resistant human breast cancer MCF-7 cells, here, we wanted to assess a new use of Tan IIA in enhancing the chemosensitivity of breast cancer cells to Dox and investigated its possible mechanisms. The results showed that Tan IIA could enhance the anti-tumor effect of Dox on MCF-7 and MCF-7/dox cells in a dose-dependent manner, especially that of on MCF-7/dox cells. Even nontoxic dose of Tan IIA could also promote intracellular Dox accumulation of MCF-7 and MCF-7/dox cells through down-regulating the expression of efflux ABC transporters including P-gp, BCRP and MRP1, which can effectively eliminated cancerous cells including BCSCs, thereby enhancing the chemosensitivity of breast cancer. Therefore, Tan IIA can be used as a new potential chemotherapeutic sensitizer for the combination treatment of breast cancer.

Silybin counteracts doxorubicin resistance by inhibiting GLUT1 expression

Despite significant advances in the diagnosis and treatment of cancer, the development of drug resistance still remains one of the principal causes that hampers the effectiveness of the therapy. Emerging evidences support the idea that the dysregulated metabolism could be related to drug resistance. The major goal of this study was to target cancer metabolic pathways using new pharmacological approaches coming from natural sources in order to possibly prevent or overcome this phenomenon. Firstly, the metabolic profile of human colorectal adenocarcinoma cells sensitive (LoVo WT) and resistant to doxorubicin (LoVo DOX) was delineated demonstrating that resistant cells remodel their metabolism toward a glycolytic phenotype. In particular it was observed that doxorubicin-resistant cancer cells exhibit an increased dependency from glucose for their survival, associated with overexpression of the glycolytic pathway. Moreover, both GLUT1 mRNA and protein expression significantly increased in LoVo DOX cells. Given the results about the metabolic profile, silybin, modulator of GLUTs, was selected as potential candidate to overcome doxorubicin resistance and, intriguingly, data revealed not only that silybin is more active in resistant cells than in wild type cells, but also that the combined treatment with doxorubicin and silybin presents a synergistic effect in LoVo DOX cells. Although many unanswered questions still remain about the molecular mechanism of silybin, these data suggest that targeting GLUTs may be a good strategy to restore doxorubicin sensitivity and elude drug resistance.

Discovery of monocarbonyl curcumin-BTP hybrids as STAT3 inhibitors for drug-sensitive and drug-resistant breast cancer therapy

Signal transducer and activator of transcription 3 (STAT3) is a well-known antitumor target. Exogenous ROS insult can lead to selective cytotoxicity against cancer cells. A combination of STAT3 inhibition and “oxidation therapy” may be a new strategy to address the multidrug-resistance issue due to their important roles in the survival and drug resistance of cancer cells. Here, a series of novel curcumin-BTP hybrids were designed and evaluated as STAT3 inhibitors with ROS production activity. Compound 6b exerted the best antitumor activity and selectivity for MCF-7 and MCF-7/DOX cells (IC50 = 0.52 μM and 0.40 μM, respectively), while its IC50 value for MCF-10A breast epithelial cells was 7.72 μM. Furthermore, compound 6b suppressed STAT3 phosphorylation, nuclear translocation and DNA-binding activity and the expression of STAT3 specific oncogenes. Increases in the level of IL-6-induced p-STAT3 were also inhibited by 6b without influencing IFN-γ-induced p-STAT1 expression. Additionally, 6b effectively promoted intracellular ROS accumulation, induced cancer cell apoptosis and cell cycle arrest, abolished the colony formation ability of breast cancer cells, and inhibited P-gp expression in MCF-7/DOX cells. Finally, 6b suppressed the growth of implanted human breast cancer in vivo. Our findings highlight that 6b may be a promising therapeutic agent for drug-sensitive and drug-resistant breast cancers.

ARTEMISININ MODULATING EFFECT ON HUMAN BREAST CANCER CELL LINES WITH DIFFERENT SENSITIVITY TO CYTOSTATICS

To explore effects of Artemisinin on a series of breast cancer cells with different sensitivity to typical cytotoxic drugs (doxorubicin - Dox; cisplatin - DDP) and to investigate possible artemisinin-induced modification of the mechanisms of drug resistance. The study was performed on wild-type breast cancer MCF-7 cell line (MCF-7/S) and its two sublines MCF-7/Dox and MCF-7/DDP resistant to Dox and DDP, respectively. The cells were treated with artemisinin and iron-containing magnetic fluid. The latter was added to modulate iron levels in the cells and explore its role in artemisinin-induced effects. The MTT assay was used to monitor cell viability, whereas changes of expression of selected proteins participating in regulation of cellular iron homeostasis were estimated using immunocytochemical methods. Finally, relative expression levels of miRNA-200b, -320a, and -34a were examined by using qRT-PCR. Artemisinin affects mechanisms of the resistance of breast cancer cells towards both Dox and DDP at sub-toxic doses. The former drug induces changes of expression of iron-regulating proteins via different mechanisms, including epigenetic regulation. Particularly, the disturbances in ferritin heavy chain 1, lactoferrin, hepcidin (decrease) and ferroportin (increase) expression (р ≤ 0.05) were established. The most enhanced increase of miRNA expression under artemisinin influence were found for miRNA-200b in MCF-7/DDP cells (7.1 ± 0.98 fold change), miRNA-320a in MCF-7/Dox cells (2.9 ± 0.45 fold change) and miRNA-34a (1.7 ± 0.15 fold change) in MCF-7/S cells. It was observed that the sensitivity to artemisinin can be influenced by changing iron levels in cells. Artemisinin can modify iron metabolism of breast cancer cells by its cytotoxic effect, but also by inducing changes in expression of iron-regulating proteins and microRNAs (miRNAs), involved in their regulation. This modification affects the mechanisms that are implicated in drug-resistance, that makes artemisinin a perspective modulator of cell sensitivity towards chemotherapeutic agents in cancer treatment.

Brazilein Increased Cytotoxic Activity of Doxorubicin on MCF-7/DOX Cells

Brazilein is a compound obtained in a large amount from the dried heartwood of Secang (Caesalpinia sappan L.). Brazilein has strong cytotoxic effect in several cancer cell lines. This research was designed to evaluate the cytotoxic effect of brazilein and its combination with a chemotherapy agent, doxorubicin on MCF-7/DOX breast cancer cells. In the cytotoxicity assay, MCF-7/DOX cells were cultured in the presence of brazilein solely and in combination with doxorubicin for 24 hours and cell viability was evaluated by using MTT assay. MTT assay showed a dose-dependent inhibition of cell proliferation with IC50 value of 37 µM. Brazilein increased doxorubicin’s cytotoxic activity on MCF-7/DOX cells. Both of single treatment with different concentration of brazilein 12.5 and 25 mM or doxorubicin 0.8 and 1 mM gave cell viability percentage above 80%, but combination of them led to decrease the cell viability percentage significantly. Based on this research, it can be concluded that brazilein is potential to be developed as a co-chemotherapy agent on breast cancer cell that have been resistant to doxorubicin. Futher study must be held to evaluate its molecular mechanism.Keywords : brazilein, doxorubicin, MCF-7/DOX, cytotoxic.

A redox-responsive strategy using mesoporous silica nanoparticles for co-delivery of siRNA and doxorubicin.

Co-delivery of gene and drug therapies for cancer treatment remains a major goal of nanocarrier research. In this study, mesoporous silica nanoparticles (MSNs) were used to co-deliver siRNA and doxorubicin (Dox) for redox-controlled release. The present nanocarrier (MSNs-SS-siRNA@Dox) has mesoporous silica cores that can be loaded with Dox, while siRNA connects to the core surface by disulfide linkage and plays a gatekeeper role. Disulfide linkages were also utilized to target intracellular GSH, and their cleavage led to the release of Dox and siRNA. Release of siRNA and Dox was correlated with GSH concentrations, and rapid release at 10 mM GSH reflected reductive cleavage of intermediate disulfide linkages. Subsequent experiments using an in vitro Dox delivery and release assay indicated that MSNs-SS-siRNA@Dox significantly enhanced the accumulation of Dox in cells compared with that after treatment with free Dox. Moreover, MSNs-SS-siRNA@Dox has sufficient efficiency to knock down target protein expression. More importantly, MSNs-SS-siRNA@Dox displayed great potential for tumor targeting and achieved satisfactory therapeutic effects on tumor growth inhibition in vivo. In summary, the present nanoparticles may provide an effective strategy for the design and development of controlled release and co-delivery of siRNA and drugs for cancer therapy.