Doxorubicin-resistant Breast Cancer Cells Research Articles

Blockage of AMPK-ULK1 pathway mediated autophagy promotes cell apoptosis to increase doxorubicin sensitivity in breast cancer (BC) cells: an in vitro study

BackgroundActivation of autophagy flux contributed to resistance of breast cancer (BC) cells to current chemotherapeutic drugs, which seriously limited their therapeutic efficacy and facilitated BC recurrence in clinic. However, the detailed mechanisms are still not fully understood. In the present study, we identified that inactivation of AMPK-ULK1 signaling cascade mediated protective autophagy sensitized BC cells to doxorubicin in vitro.MethodsCell counting kit-8 (CCK-8) assay and colony formation assay were performed to evaluate cell proliferation abilities. Trypan blue staining assay was used to examine cell viability, and Annexin V-FITC/PI double staining method was conducted to determine cell apoptosis. The autophagosomes in BC cells were observed and photographed by electronic microscope (EM). Western Blot analysis was employed to examine genes expressions at protein levels.ResultsThe parental doxorubicin-sensitive BC (DS-BC) cells were exposed to increasing concentrations of doxorubicin to establish doxorubicin-resistant BC (DR-BC) cells, and the DR-BC cells were much more resistant to high-dose doxorubicin treatment compared to the DS-BC cells. Interestingly, high-dose doxorubicin specifically increased LC3B-II/I ratio, promoted autophagosomes formation and decreased p62 expression levels to facilitate autophagy in DR-BC cells, instead of DS-BC cells, and the autophagy inhibitor 3-methyladenine (3-MA) enhanced the cytotoxic effects of high-dose doxorubicin on DR-BC cells. In addition, we proved that high-dose doxorubicin triggered protective autophagy in DR-BC cells by activating AMPK-ULK1 pathway. Functionally, high-dose doxorubicin increased the expression levels of phosphorylated AMPK (p-AMPK) and ULK1 (p-ULK1) to activate AMPK-ULK1 pathway in DR-BC cells, and the inhibitors for AMPK (compound C) and ULK1 (SBI-0206965) blocked autophagy to promote cell death and slow down cell growth in DR-BC cells treated with high-dose doxorubicin.ConclusionsCollectively, our in vitro data indicated that blockage of AMPK-ULK1 signaling cascade mediated protective autophagy might be a promising strategy to increase doxorubicin sensitivity for BC treatment.

Ardeemins and citrinin dimer derivatives from Aspergillus terreus harbored in Pinellia ternate

One new alkaloid, 15b-β-hydroxy-16α-hydroxy-5-N-acetylardeemin (1), and one new dimeric derivative of citrinin, penicitrinol Q (5), together with three known alkaloids (2–4) and two citrinin dimers (6–7), were isolated from the solid culture of an endophytic Aspergillus terreus resided in Pinellia ternata. Their structures were established by detailed spectroscopic analysis, and the absolute configurations were determined by single crystal X‑ray diffraction and the comparison of ECD spectra. Penicitrinol Q (5) exhibited inhibitory effects against Staphylococcus aureus and Bacillus subtilis with MIC50 values of 4.3 and 6.2 μg/mL, respectively. When using a concentration of 30 μM, compound 2 showed 2.1-fold enhancement of doxorubicin susceptibility in treating doxorubicin-resistant human breast cancer cell line.

Honokiol antagonizes doxorubicin resistance in human breast cancer via miR-188-5p/FBXW7/c-Myc pathway.

Honokiol, a natural phenolic compound derived from Magnolia plants, is a promising anti-tumor compound that exerts a wide range of anti-cancer effects. Herein, we investigated the effect of honokiol on doxorubicin resistance in breast cancer. Doxorubicin-sensitive (MCF-7 and MDA-MB-231) and doxorubicin-resistant (MCF-7/ADR and MDA-MB-231/ADR) breast cancer cell lines were treated with doxorubicin in the absence or presence of honokiol; then, the following tests were performed: flow cytometry for cell apoptosis, WST-1 assay for cell viability, qPCR and western blot for the expression of miR-188-5p, FBXW7, and c-Myc. MiR-188-5p mimic, miR-188-5p inhibitor, siFBXW7, and c-Myc plasmids were transfected into cancer cells to evaluate whether miR-188-5p and FBXW7/c-Myc signaling are involved in the effect of honokiol on doxorubicin resistance in breast cancer. A dual luciferase reporter system was used to study the direct interaction between miR-188-5p and FBXW7. Honokiol sensitized doxorubicin-resistant breast cancer cells to doxorubicin-induced apoptosis. Mechanically, upregulation of miR-188-5p was associated with doxorubicin resistance, and honokiol enhanced doxorubicin sensitivity by downregulating miR-188-5p. FBXW7 was confirmed to be a direct target gene of miR-188-5p. FBXW7/c-Myc signaling was involved in the chemosensitization effect of honokiol. Honokiol induced apoptosis in MCF-7/ADR and MDA-MB-231/ADR cells. However, FBXW7 silencing or c-Myc transfection resulted in resistance to the honokiol-induced apoptotic effect. These findings suggest that downregulation of miR-188-5p by honokiol enhances doxorubicin sensitivity through FBXW7/c-Myc signaling in human breast cancer. Our study finds an important role of miR-188-5p in the development of doxorubicin resistance in breast cancer, and enriches our understanding of the mechanism of action of honokiol in cancer therapy.

MiR-34a Reverses Doxorubicin Resistance in Breast Cancer

Chemotherapy is effectively used for treating breast cancer, but the problem of tumor resistance to chemotherapy drugs has been plaguing scientists. Our study investigated miR-34a?s effect on the sensitivity of drug-resistant strains to chemotherapeutic drugs using doxorubicin-resistant strains of breast cancer cells. Cell survival rate was detected by MTT assay. The doxorubicin-resistant strain rMCF-7 was obtained. The cell scratching method and CCK-8 method were used to detect cell migration and proliferation.Western Blot was performed for measuring SIRT1, p-AKT, AKT, p-mTOR and mTOR level. miR-34a significantly reduced the survival rate o doxorubicin-resistant breast cancer cell line rMCF-7 and significantly enhanced doxorubicin?s effect on inhibiting cell proliferation and cell migration. Compared with the doxorubicin group alone, miR-34a and doxorubicin combination group significantly downregulated SIRT1, p-AKT/AKT and p-mTOR/mTOR related proteins in rMCF-7 cells. miR-34a can reverse the resistance of doxorubicin in breast cancer in vitro and the mechanism may be through inhibition of SIRT and AKT signaling.

Overcoming anticancer resistance by photodynamic therapy-related efflux pump deactivation and ultrasound-mediated improved drug delivery efficiency

One of the major obstacles to successful chemotherapy is multi-drug resistance (MDR). A multi-drug resistant cancerous cell abnormally overexpresses membrane transporters that pump anticancer drugs out of the cell, resulting in low anticancer drug delivery efficiency. To overcome the limitation, many attempts have been performed to inhibit the abilities of efflux receptors chemically or genetically or to increase the delivery efficiency of anticancer drugs. However, the results have not yet been satisfactory. In this study, we developed nanoparticle-microbubble complexes (DOX-NPs/Ce6-MBs) by conjugating doxorubicin loaded human serum albumin nanoparticles (DOX-NPs) onto the surface of Chlorin e6 encapsulated microbubbles (Ce6-MBs) in order to maximize anticancer efficiency by overcoming MDR. Under the ultrasound irradiation, DOX-NPs and Ce6 encapsulating self-assembled liposomes or micelles were effectively delivered into the cells due to the sonoporation effect caused by the microbubble cavitation. At the same time, reactive oxygen (ROS) generated from intracellularly delivered Ce6 by laser irradiation arrested the activity of ABCG2 efflux receptor overexpressed in doxorubicin-resistant breast cancer cells (MCF-7/ADR), resulting in increased the chemotherapy efficacy. In addition, the total number of side population cells that exhibit the properties of cancer stem-like cells were also reduced by the combination of photodynamic therapy and chemotherapy. In conclusion, DOX-NPs/Ce6-MBs will provide a platform for simultaneously overcoming MDR and increasing drug delivery and therefore, treatment efficiency, under ultrasound irradiation.

Long non-coding RNA H19 regulates proliferation and doxorubicin resistance in MCF-7 cells by targeting PARP1

ABSTRACT Chemoresistance is a major obstacle to effective breast cancer chemotherapy. However, the underlying molecular mechanisms remain unclear. The long noncoding RNA H19 (H19) is involved in various stages of tumorigenesis, however, its role in doxorubicin resistance remains unknown. The goal of this study was to evaluate the role of H19 in the development of doxorubicin-resistant breast cancer. Quantitative real-time PCR (qRT-PCR) analyzed H19 expression in chemotherapy-resistant and sensitive breast cancer tissues. Both knockdown and overexpression of H19 were used to assess the sensitivity to doxorubicin in breast cancer cells in vitro and in vivo. qRT-PCR and Western blot were used to explore the doxorubicin resistance mechanism of H19. We observed that the H19 expression was significantly upregulated in chemotherapy-resistant breast cancer tissues and doxorubicin-resistant breast cancer cell lines. Knockdown of H19 enhanced the sensitivity to doxorubicin both in vitro and in vivo. While H19 overexpression developed doxorubicin-resistant in breast cancer cells both in vitro and in vivo. Furthermore, it was revealed that H19 negatively regulated PARP1 expression in breast cancer cells following doxorubicin treatment. Knockdown of H19 sensitized breast cancer cells to doxorubicin by promoting PARP1 upregulation. H19 overexpression could recapitulate doxorubicin resistance by PARP1 downregulation. Our findings revealed that H19 plays a leading role in breast cancer chemoresistance development, mediated mainly through a H19-PARP1 pathway.

The LQB-223 Compound Modulates Antiapoptotic Proteins and Impairs Breast Cancer Cell Growth and Migration.

Drug resistance represents a major issue in treating breast cancer, despite the identification of novel therapeutic strategies, biomarkers, and subgroups. We have previously identified the LQB-223, 11a-N-Tosyl-5-deoxi-pterocarpan, as a promising compound in sensitizing doxorubicin-resistant breast cancer cells, with little toxicity to non-neoplastic cells. Here, we investigated the mechanisms underlying LQB-223 antitumor effects in 2D and 3D models of breast cancer. MCF-7 and MDA-MB-231 cells had migration and motility profile assessed by wound-healing and phagokinetic track motility assays, respectively. Cytotoxicity in 3D conformation was evaluated by measuring spheroid size and performing acid phosphatase and gelatin migration assays. Protein expression was analyzed by immunoblotting. Our results show that LQB-223, but not doxorubicin treatment, suppressed the migratory and motility capacity of breast cancer cells. In 3D conformation, LQB-223 remarkably decreased cell viability, as well as reduced 3D culture size and migration. Mechanistically, LQB-223-mediated anticancer effects involved decreased proteins levels of XIAP, c-IAP1, and Mcl-1 chemoresistance-related proteins, but not survivin. Survivin knockdown partially potentiated LQB-223-induced cytotoxicity. Additionally, cell treatment with LQB-223 resulted in changes in the mRNA levels of epithelial-mesenchymal transition markers, suggesting that it might modulate cell plasticity. Our data demonstrate that LQB-223 impairs 3D culture growth and migration in 2D and 3D models of breast cancer exhibiting different phenotypes.

Downregulation of STAT3, β-Catenin, and Notch-1 by Single and Combinations of siRNA Treatment Enhance Chemosensitivity of Wild Type and Doxorubicin Resistant MCF7 Breast Cancer Cells to Doxorubicin.

Combinatorial therapeutic strategies using siRNA and small molecules to eradicate tumors are emerging. Targeting multiple signaling pathways decreases the chances of cancer cells switching and adapting new signaling processes that may occur when using a single therapeutic modality. Aberrant functioning of Notch-1, Wnt/β-catenin, and STAT3 proteins and their crosstalk signaling pathways have been found to be involved in tumor survival, drug resistance, and relapse. In the current study, we describe a therapeutic potential of single and combinations of siRNA designed for silencing Notch-1, Wnt/β-catenin, and STAT3 in MCF7_DoxS (wild type) and MCF7_DoxR (doxorubicin resistant) breast cancer cells. The MCF7_DoxR cells were developed through treatment with a gradual increase in doxorubicin concentration, the expression of targeted genes was investigated, and the expression profiling of CD44/CD24 of the MCF7_DoxS and MCF7_DoxR cells were detected by flow cytometry. Both MCF7_DoxS and MCF7_DoxR breast cancer cells were treated with single and combinations of siRNA to investigate synergism and were analyzed for their effect on cell proliferation with and without doxorubicin treatment. The finding of this study showed the overexpression of targeted genes and the enrichment of the CD44−/CD24+ phenotype in MCF7_DoxR cells when compared to MCF7_DoxS cells. In both cell lines, the gene silencing efficacy showed a synergistic effect when combining STAT3/Notch-1 and STAT3/Notch-1/β-catenin siRNA. Interestingly, the chemosensitivity of MCF7_DoxS and MCF7_DoxR cells to doxorubicin was increased when combined with siRNA treatment. Our study shows the possibility of using single and combinations of siRNA to enhance the chemosensitivity of cancer cells to conventional antitumor chemotherapy.

Effectiveness of Small Interfering RNA Delivery via Arginine-Rich Polyethylenimine-Based Polyplex in Metastatic and Doxorubicin-Resistant Breast Cancer Cells.

Poor cellular uptake, rapid degradation in the presence of serum, and inefficient transfection are some of the major barriers in achieving therapeutic efficacy of naked small interfering RNAs (siRNAs). We investigated the efficacy of the polyplex formulated using our synthesized polymer, polyethylene glycol (PEG)-modified l-arginine oligo(-alkylaminosiloxane) that is grafted with poly(ethyleneimine) (PEI) for siRNA delivery. We hypothesized that the polyplex formulated using the polymer with a balanced composition of PEI for siRNA condensation and its protection, PEG for polyplex stability and to minimize the PEI-associated toxicity, and with arginine facilitating cellular uptake would overcome the aforementioned issues with siRNA delivery. We tested our hypothesis using antiluciferase siRNA in luciferase-expressing metastatic breast cancer cells (MDA-MB-231-Luc-D3H2LN) and anti-ABCB1 siRNA against an efflux membrane protein, ABCB1, in doxorubicin (DOX)-resistant breast cancer cells (MCF-7/Adr). The results demonstrated that the polyplex at an optimal nucleotide/polymer ratio is stable in the presence of excess polyanions, has no cellular toxicity, and protects siRNA from RNase degradation. Transfection of MDA-MB-231-Luc-D3H2LN cells with antiluciferase siRNA polyplex showed almost complete knockdown of luciferase expression. In MCF-7/Adr cells, transfection with anti-ABCB1 siRNA effectively downregulated its target efflux protein, ABCB1; increased cellular uptake of DOX; and enhanced its cytotoxic effect. However, the cotreatment did not completely overcome drug resistance, suggesting that further optimization is needed and/or a mechanism(s) other than the efflux protein ABCB1 may be involved in drug resistance. In conclusion, our polyplex is effective for siRNA delivery and can be explored for different therapeutic applications.

Transferrin-Conjugated Polymeric Nanoparticle for Receptor-Mediated Delivery of Doxorubicin in Doxorubicin-Resistant Breast Cancer Cells.

In this study, a transferrin (Tf)-conjugated polymeric nanoparticle was developed for the targeted delivery of the chemotherapeutic agent doxorubicin (Dox) in order to overcome multi-drug resistance in cancer treatment. Our objective was to improve Dox delivery for producing significant antitumor efficacy in Dox-resistant (R) breast cancer cell lines with minimum toxicity to healthy cells. The results of our experiments revealed that Dox was successfully loaded inside a transferrin (Tf)-conjugated polymeric nanoparticle composed of poloxamer 407 (F127) and 123 (P123) (Dox/F127&P123-Tf), which produced nanosized particles (~90 nm) with a low polydispersity index (~0.23). The accelerated and controlled release profiles of Dox from the nanoparticles were characterized in acidic and physiological pH and Dox/F127&P123-Tf enhanced Dox cytotoxicity in OVCAR-3, MDA-MB-231, and MDA-MB-231(R) cell lines through induction of cellular apoptosis. Moreover, Dox/F127&P123-Tf inhibited cell migration and altered the cell cycle patterns of different cancer cells. In vivo study in MDA-MB-231(R) tumor-bearing mice demonstrated enhanced delivery of nanoparticles to the tumor site when coated in a targeting moiety. Therefore, Dox/F127&P123-Tf has been tailored, using the principles of nanotherapeutics, to overcome drug-resistant chemotherapy.

MiR-135b-5p enhances doxorubicin-sensitivity of breast cancer cells through targeting anterior gradient 2

BackgroundThe pro-oncogenic anterior gradient 2 (AGR2) is involved in tumor growth and drug resistance of breast cancer. Mechanisms that regulate expression of AGR2 still need to be elucidated.MethodsIn this study, expression levels of AGR2 and miR-135b-5p were analyzed in different breast cancer cell lines as well as in clinical breast cancer tissues. The in vitro and in vivo functional effect of AGR2 and miR-135b-5p were also investigated. A luciferase reporter assay was applied to confirm the interaction between miR-135b-5p and AGR2 mRNA.ResultsWe identified AGR2 as a target of miR-135b-5p. Expression of AGR2 was up-regulated in doxorubicin-resistant breast cancer cells. AGR2 mediated doxorubicin-sensitivity of breast cancer cells both in vitro and in vivo. miR-135b-5p negatively regulated AGR2-expression of breast cancer cells increasing doxorubicin-sensitivity. However, miR-135b-5p was down-regulated in doxorubicin-resistant breast cancer cells as well as during treatment with doxorubicin, which might be a probable reason for over-expression of AGR2. Up-regulation of miR-135b-5p increased doxorubicin-sensitivity of breast cancer cells in vivo. In addition, levels of AGR2 negatively correlated with levels of miR-135b-5p in clinical breast cancer tissue samples.ConclusionOur results highlight the potential of miR-135b-5p as a target for treating AGR2-expressing breast cancer with doxorubicin-resistance.

Seven new guanacastane-type diterpenoids from the fungus Verticillium dahliae

Seven new guanacastane-type diterpenoids, namely dahlianes E-K (1–7), and three known ones (8–10) were isolated from the cultures of the fungus Verticillium dahliae. Their structures were established by extensive spectroscopic data analysis along with Rh2(OCOCF3)4- and Mo2(OAc)4-induced electronic circular dichroism (ECD) experiment. Dahliane G showed an 80-fold potentiation effect on the sensitization of doxorubicin at the concentration of 15 μM when screening the reversal activity on doxorubicin-resistant human breast cancer cells (MCF-7/DOX).

Hypoxia-induced gene expression pattern in doxorubicin resistant MCF7 cells

Purpose: To investigate hypoxia-induced gene expression pattern in doxorubicin-resistant human breast cancer cells (MCF7). Methods: Human breast cancer cells (MCF7) were exposed to 60 episodes of 8 h hypoxia thrice a week for three months. Chemo-resistance to doxorubicin was assessed using 3-(4,5-dimethylthiazol-2- yl)-2, 5-diphenyl tetrazolium bromide (MTT) cell proliferation assay. Real-time quantitative polymerase chain reaction (qRT-PCR) assay was performed to assess gene expression pattern in doxorubicinresistant cells on exposure to hypoxia. Results: Hypoxia significantly increased the resistance of MCF7 cells to doxorubicin, with a maximum of 16.42-fold enhancement after 25 episodes of 8-h hypoxia, while the resistance thereafter significantly decreased with prolonged episodes of hypoxia (p < 0.05). Gene expression analysis revealed significant changes in 42 genes. The expressions of 10 of these genes were significantly upregulated, while those of 32 genes were significantly down-regulated (p < 0.05). Cytochrome P450 family 1, subfamily A, member1 (CYP1A1) was the most conspicuous upregulated gene (13.32-fold), while breast cancer gene 1 (BRCA1) was the most down-regulated (8.23-fold). Gene expression analysis after 60 episodes of 8-h hypoxia revealed the upregulation of CYP1A1 (5.77-fold). Similarly, 27 genes were significantly down-regulated, with BRCA2 as the most down-regulated gene (8.11-fold). Topoisomerase (DNA) II alpha (TOP2A) was the most down-regulated among genes involved in drug metabolism and resistance (6.37-fold), while cyclin-dependent kinase 2 (CDK2) was the most profoundly downregulated among genes involved in cell cycle regulation (3.56-fold). Conclusion: These results indicate that development of resistance to doxorubicin by MCF7 cells after short-term hypoxia results from the upregulation of genes responsible for the metabolism of doxorubicin and for shifting the cells to alternative pathway driven principally by EGF and ESR2. The observed down-regulation is an adaptation of the MCF7 cells to survive under long-term hypoxia.

Regulation of ABCB1 activity by microRNA-200c and microRNA-203a in breast cancer cells: the quest for microRNAs' involvement in cancer drug resistance.

Aim: ABCB1 is a major player in cancer drug resistance. The purpose of this study was to functionally assess the regulation of ABCB1 activity in a doxorubicin-resistant breast cancer cell line by miR-200c and miR-203.Methods: Human breast carcinoma cell lines MCF-7 (Doxorubicin-sensitive and not expressing ABCB1) and KCR (Doxorubicin-resistant and expressing ABCB1) were used to evaluate the expression levels of miR-200c and miR-203 by Real-time quantitative PCR (RT-qPCR). The effects of transient ectopic expression of miRNA-200c and miR-203 on the expression of ABCB1 in KCR and MCF-7 cells was verified by RT-qPCR and Western Blot. The extrusion activity of the ABCB1 pump was analyzed by fluorescence microscopy and flow cytometry through fluorescence substrate retention assays (DiOC2) in the presence and absence of the ABCB1 inhibitor verapamil.Results: RT-qPCR results indicated a 100,000-fold increase in ABCB1 mRNA expression levels in KCR cells compared to MCF-7 cells, and is inversely correlated with the expression of miR-203 and miR-200c. The insertion of miR-200c and miR-203 led to a higher retention of DiOC2 within KCR cells, and slightly reduced the protein levels of ABCB1 in KCR cells, although the high initial expression of ABCB1 masked the reduction in protein levels. The increased intracellular accumulation of the fluorescent due DiOC2 in the presence of the ABCB1 inhibitor verapamil correlated with the inhibition caused by miR-203 and miR-200c in transfected cells.Conclusion: The present study demonstrates that miR-200c and miR-203 exert a negative modulating effect on the activity of ABCB1 associated with doxorubicin resistance.

Cytotoxic seco-cytochalasins from an endophytic Aspergillus sp. harbored in Pinellia ternata tubers

Six new seco-cytochalasins A-F (1–6), two new asperlactones G-H (7–8) along with three known cytochalasins (9–11) were isolated from the solid cultures of an endophytic fungus Aspergillus sp. Their structures were elucidated by comprehensive spectral analysis, and their absolute configurations were determined through Mo2(OCOCH3)4-induced electronic circular dichroism (ECD) spectra and Rh2(OCOCF3)4-induced ECD experiment. Compounds 5 and 6 were rare seco-cytochalasins possessing an α, β-unsaturated furanone structure in their side-chains. These isolates exhibited cytotoxicity against human lung cancer A-549 cell line with IC50 values ranging from 7.8 to 70.2 μM. At the concentration of 16 μM, compound 4 also exerted a 3-fold enhancement of doxorubicin susceptibility on doxorubicin-resistant human breast cancer (MCF-7/DOX) cell line.

Rare dimeric guaianes from Xylopia vielana and their multidrug resistance reversal activity

Thirteen undescribed dimeric guaianes were isolated from the leaves of Xylopia vielana Pierre. Their structures were elucidated by NMR spectroscopy and mass spectrometry, and the absolute configurations of vielanins G-Q were determined by a combination of the circular dichroism (CD) exciton chirality method, chemical conversion, and electronic CD (ECD) spectroscopy analysis. Vielaninors A and B are the first examples of trinor-guaiane-dimers. Multidrug resistance reversal activity assay of the isolates was evaluated in doxorubicin-resistant human breast cancer cells. Vielanins H, K-M, P, and Q were noncytotoxic and enhanced the cytotoxicity of doxorubicin by 2.1–41.6-fold at 10 μM.

A Microbore Tubing Based Spiral for Multistep Cell Fractionation.

Cells were separated with the aid of a multistep spiral fractionation device, utilizing hydrodynamic forces in a spiral tubing. The spiral was fabricated using "off-the-shelf" microbore tubing, allowing for cheap and fast prototyping to achieve optimal cell separation. As a first step, a model system with 20 and 40 μm beads was used to demonstrate the effectiveness of the multistep separation device. With an initial purity of 5%, a separation purity of 83% was achieved after a two-step separation with the addition of 0.1% polyethylene glycol (PEG)-8000. Next, doxorubicin-resistant polyploid giant breast cancer cells (MDA-MB-231) were separated from doxorubicin-sensitive monoploid small breast cancer cells in the same fashion as the beads, resulting in a purity of around 40%, while maintaining a cell viability of more than 90%. Combined with basic cell analytical methods, the hydrodynamic separation principle of the device could be envisaged to be useful for a variety of cell fractionation needs in cell biology and in clinical applications.

Anti-cancer effects of Rhizoma Curcumae against doxorubicin-resistant breast cancer cells

BackgroundChemotherapy is a primary approach in cancer treatment after routine surgery. However, chemo-resistance tends to occur with chemotherapy in clinic, resulting in poor prognosis and recurrence. Nowadays, Chinese medicine may shed light on design of new therapeutic modes to overcome chemo-resistance. Although Rhizoma Curcumae possesses anti-cancer activities in various types of cancers, the effects and underlying mechanisms of its bioactive components against chemo-resistance are not clear. Therefore, the present study aims to explore the potential effects of Rhizoma Curcumae on doxorubicin-resistant breast cancer cells.MethodsThe expression and function of ABC transporters in doxorubicin-resistant MCF-7 breast cancer cells were measured by western blotting and flow cytometry. Cell viability was detected using MTT assay. The combination index was analyzed using the CalcuSyn program (Biosoft, Ferguson, MO), based on the Chou–Talalay method.ResultsIn our present study, P-gp was overexpressed at protein level in doxorubicin-resistant MCF-7 cell line, but short of MRP1 and BCRP1. Essential oil of Rhizoma Curcumae and the main bioactive components were assessed on doxorubicin-resistant MCF-7 cell line. We found that the essential oil and furanodiene both display powerful inhibitory effects on cell viability, but neither of these is the specific inhibitor of ABC transporters. Moreover, furanodiene fails to enhance the efficacy of doxorubicin to improve multidrug resistance.ConclusionOverall, our findings fill the gaps of the researches on chemo-resistance improvement of Rhizoma Curcumae and are also beneficial for Rhizoma Curcumae being developed as a promising natural product for cancer adjuvant therapy in the future.

Silymarin increases the sensitivity of breast cancer cells to doxorubicin in doxorubicin-induced MCF-7 cells by inhibiting breast cancer resistance protein expression

The efflux transporter breast cancer resistance protein (BCRP) is typically overexpressed in cancer cells with reduced doxorubicin sensitivity. Silymarin is known to exert inhibitory effects against BCRP; therefore, the exposure of doxorubicin-resistant breast cancer cells to silymarin is expected to increase their sensitivity to doxorubicin. Here, we examined the effect of silymarin addition on the sensitivity of breast cancer cells to doxorubicin. MCF-7 breast cancer cells were exposed to doxorubicin for 14 days. Subsequently, the cells were treated with different concentrations of silymarin (10, 25, 50, or 100 μM) with or without doxorubicin. On days 3 and 7 following the treatment, cells were analyzed for viability and BCRP mRNA expression. MCF-7 cells exposed to doxorubicin for 14 days showed reduced sensitivity to doxorubicin, as indicated by the 9.5-fold shift in cytotoxicity concentration and the 9.7-fold increase in BCRP mRNA expression. Treatment with the combination of different concentrations of silymarin and doxorubicin significantly decreased the percent cell viability and reduced BCRP mRNA expression on days 3 and 7. The combination of doxorubicin and silymarin increased the sensitivity of MCF-7 cells to doxorubicin through the inhibition of BCRP mRNA expressions by silymarin.

Therapeutic Potential of DNAzyme Loaded on Chitosan/Cyclodextrin Nanoparticle to Recovery of Chemosensitivity in the MCF-7 Cell Line.

Commonly, acquired resistances to anticancer drug are mediated by overexpression of a membrane-associated protein that encode via multi-drug resistance gene-1 (MDR1). Herein, the mRNA-cleaving DNAzyme that targets the mRNA of MDR1 gene in doxorubicin-resistant breast cancer cell line (MCF-7/DR) loaded on the chitosan β-cyclodextrin complexes was used as a tropical agent. Chitosan/β-cyclodextrin complexes were used to deliver DNAzymes into cancer cells. Determination of the physicochemical characteristics of the particles was done by photon correlation spectroscopy and scanning electron microscopy. The encapsulation efficiency of the complexes was tested by using gel retardation assay. Positively charged nanoparticles interacted with DNAzyme that could perform as an efficient DNAzyme transfection system. The rationale usage of this platform is to sensitize MCF-7/DR to doxorubicin by downregulating the drug-resistance gene MDR1. Results demonstrated a downregulation of MDR1 mRNAs in MCF-7/DR/DNZ by real-time PCR, compared to the MCF-7/DR as control. WST1 assay showed the 22-fold decrease in drug resistance on treated cells 24 h after transfection. Results showed the intracellular accumulation of Rh123 increased in the treated cells with DNAzyme. Results suggested a potential platform in association with chemotherapy drug for cancer therapy and indicated extremely efficient at delivery of DNAzyme in restoring chemosensitivity.