Differential gene and microRNA expression between etoposide resistant and etoposide sensitive MCF7 breast cancer cell lines.
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ABSTRACT:
SUBMITTER: Moitra K
PROVIDER: S-EPMC3445463 | biostudies-literature | 2012
REPOSITORIES: biostudies-literature
Publications
Differential gene and microRNA expression between etoposide resistant and etoposide sensitive MCF7 breast cancer cell lines.
Moitra Karobi K Im Kate K Limpert Katy K Borsa Alexander A Sawitzke Julie J Robey Rob R Yuhki Naoya N Savan Ram R Huang Da Wei da W Lempicki Richard A RA Bates Susan S Dean Michael M
PloS one 20120918 9
In order to develop targeted strategies for combating drug resistance it is essential to understand it's basic molecular mechanisms. In an exploratory study we have found several possible indicators of etoposide resistance operating in MCF7VP cells, including up-regulation of ABC transporter genes, modulation of miRNA, and alteration in copy numbers of genes. ...[more]
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Project description:BackgroundMicroRNAs (miRs) regulate gene expression through translation inhibition of target mRNAs. One of the most promising approaches for cancer therapy is through mimicking or antagonizing the action of miRs. In this report, we analyzed the miRnome profile of several human breast cancer cell lines to determine the influence of estrogen receptor (ER) silencing previously shown to result in epithelial to mesenchymal transition (EMT) and enhanced tumor invasion.MethodsMicroRNA extracted from MDA-MB-231 (de novo ER-) and ER-silenced (acquired ER-) pII and IM-26 or ER-expressing (YS1.2) siRNA transfected derivatives of MCF7 cells was deep sequenced on Illumina NextSeq500. Respective miRnomes were compared with edgeR package in R and Venny2.1 and target prediction performed with miRTarBase. Mimics and inhibitors of selected differentially expressed miRs associated with EMT mediators (miR-200c-3p targeting ZEB1, miR-449a targeting δ-catenin and miR-29a-3p) were transfected into pII cells and mRNA targets, as well as E-cadherin and keratin 19 (epithelial and mesenchymal markers respectively) were measured using taqman PCR.ResultsEach cell line expressed about 20% of the total known human miRnome; There was a high degree of similarity between the 3 tested ER-lines. Out of these expressed miRs, 50-60% were significantly differentially expressed between ER- and ER + lines. Transfection of miR-200c-3p mimic into pII cells down regulated ZEB1 and vimentin, and increased E-cadherin and keratin 19 with accompanying morphological changes, and reduced cell motility, reflecting a reversal back into an epithelial phenotype. On the other hand, transfecting pII with miR-449a inhibitor reduced cell invasion but did not induce EMT. Transfecting pII cell line with the mimic or inhibitor of miR-29a-3p showed no change in EMT markers or cell invasion suggesting that the EMT induced by loss of ER function can be reversed by blocking some but not just any random EMT-associated genes.ConclusionsThese data suggest that differences in miR expression can be exploited not only as mediators (using mimics) and targets (using miR antagonists) for general cancer therapies aimed at regulating either individual or multiple mRNAs, but also to re-sensitize endocrine resistant breast cancers by turning them back into a type that will be susceptible to endocrine agents.
| S-EPMC9307586 | biostudies-literature
Project description:The second leading cause of cancer death for women in the U.S. is breast cancer. Moreover, a significant number of patients with breast tumors acquire resistance to drugs during therapy. To develop targeted therapeutic strategies to combat drug resistance it is essential to understand the basic molecular mechanisms through which cancer cells control sensitivity to chemotherapeutics. To identify new candidate genes and facilitate the discovery of novel drug resistance pathways, we have generated a resistance profile or ‘resistome’ of etoposide resistant MCF7 breast cancer cells. Differential expression of over 5000 genes (fold change > 2, P value < 0.05) indicate that several drug resistance mechanisms may be operating in these cells, including up-regulation of ABC transporter genes, down-regulation of the drug target and down-regulation of apoptotic genes. Several transcription factors such as RUNX2, SOX9, ETS1 and SMAD3 were up-regulated in the drug resistant cells. Targeted RUNX2 knockdown in the resistant cells using siRNA increased sensitivity to etoposide and also upregulated expression of pro-apoptotic genes indicating that RUNX2 could be a molecular target against etoposide resistance. Differential miRNA (microRNA) expression was observed among the drug resistant and sensitive cells suggesting that miRNA may also play a role in regulation of drug resistance. Hsa-miR-218, which targets ABCC6, was down-regulated in the drug resistant cell line. Transfection of a miR-218 mimic could down-regulate the expression of the efflux pump ABCC6 by 65% in drug resistant cells suggesting that regulation of miRNA may play an important role in etoposide resistance. RNA from MCF7 and etoposide-resistant MCF7 (MCF7VP) cell lines were hybridized to Affymetrix microarrays. Both samples were run in triplicate
| S-ECPF-GEOD-28413 | biostudies-other
Project description:Chemoresistance is the major cause of cancer recurrence, relapse and eventual death. Doxorubicin resistance is one such challenge in breast cancer. The use of quercetin, an antioxidant, in combination with doxorubicin has been investigated for offering protection to normal cells from the toxic side effects of doxorubicin in addition to modulation of its resistance. The present study aimed to investigate the effects of quercetin in prevention of a doxorubicin-chemoresistant phenotype in both doxorubicin-sensitive and -resistant human MCF-7 breast cancer cell lines. A doxorubicin-resistant MCF-7 cell line was established. The development of resistant cells was closely monitored for changes in morphological features. Sensitivity to doxorubicin and the doxorubicin/quercetin combination was assessed using the tetrazolium assay. To determine the mechanism by which quercetin sensitizes the doxorubicin MCF-7-resistant cell line to doxorubicin, gene expression alterations in breast cancer-related genes were examined using the reverse transcription-quantitative PCR (RT-qPCR) array technology. Resistant MCF cells were successfully developed and the inhibitory concentration (IC50) value of doxorubicin increased from 0.133 to 4 µM (wild-type to resistant). The effects of the quercetin/doxorubicin combination exhibited different effects on wild-type vs. resistant cells. The IC50 of doxorubicin was reduced in wild cells, whereas resistant cells showed an increase in cell viability at lower concentrations and a potentiation of the effects of doxorubicin only at higher concentrations. Annexin V/propidium iodide staining demonstrated that quercetin drives cells into late apoptosis and necrosis, but in resistant cells, necrosis predominates. RT-qPCR results revealed that quercetin led to a reversal in doxorubicin effects via up- and downregulation of important genes such as SNAI2, PLAU and CSF1 genes. Downregulation of cell migration genes, SNAI2 (-31.23-fold) and plasminogen activator, urokinase (PLAU; -30.62-fold), and the apoptotic pathway gene, colony stimulating factor 1 (CSF1; -17.25-fold) were the most important querticin-associated events. Other gene alterations were also observed involving cell cycle arrest and DNA repair pathways. The results of the present study indicated that quercetin could lead to a reversal of doxorubicin resistance in breast cancer cells via downregulation of the expression of important genes, such as SNAI2, PLAU and CSF1. Such findings may represent a potential strategy for reversing breast cancer cell-related chemoresistance.
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Project description:Microarrays identified miRNAs differentially expressed and 4-hydroxytamoxifen (4-OHT) regulated in MCF-7 endocrine-sensitive versus resistant LY2 human breast cancer cells. 97 miRNAs were differentially expressed in MCF-7 versus LY2 cells. Opposite expression of miRs-10a, 21, 22, 29a, 93, 125b, 181, 200a, 200b, 200c, 205, and 222 was confirmed. Bioinformatic analyses to impute the biological significance of these miRNAs identified 36 predicted gene targets from those regulated by 4-OHT in MCF-7 cells. Agreement in the direction of anticipated regulation was detected for 12 putative targets. These miRNAs with opposite expression between the two cell lines may be involved in endocrine resistance.
| S-EPMC3214732 | biostudies-literature
Project description:Azacitidine, a DNA methylation inhibitor, is employed for the treatment of acute myeloid leukemia (AML). However, drug resistance remains a major challenge for effective azacitidine chemotherapy, though several studies have attempted to uncover the mechanisms of azacitidine resistance. With the aim to identify the mechanisms underlying acquired azacitidine resistance in cancer cell lines, we developed a computational strategy that can identify differentially regulated gene networks between drug-sensitive and -resistant cell lines by extending the existing method, differentially coexpressed gene sets (DiffCoEx). The technique specifically focuses on cell line-specific gene network analysis. We applied our method to gene networks specific to azacitidine sensitivity and identified differentially regulated gene networks between azacitidine-sensitive and -resistant cell lines. The molecular interplay between the metallothionein gene family, C19orf33, ELF3, GRB7, IL18, NRN1, and RBM47 were identified as differentially regulated gene network in drug resistant cell lines. The biological mechanisms associated with azacitidine and AML for the markers in the identified networks were verified through the literature. Our results suggest that controlling the identified genes (e.g., the metallothionein gene family) and "cellular response"-related pathways ("cellular response to zinc ion", "cellular response to copper ion", and "cellular response to cadmium ion", where the enriched functional-related genes are MT2A, MT1F, MT1G, and MT1E) may provide crucial clues to address azacitidine resistance in patients with AML. We expect that our strategy will be a useful tool to uncover patient-specific molecular interplay that provides crucial clues for precision medicine in not only gastric cancer but also complex diseases.
| S-EPMC10970437 | biostudies-literature
Project description:Introduction: Although changes in microRNA (miRNA) expression correlate with breast cancer diagnostic markers, comparatively little is known about miRNA regulation by selective estrogen receptor modulators (SERMs), e.g., tamoxifen (TAM), or their role in endocrine-resistance. Methods: A microarray approach was used to identify miRNAs differentially expressed and regulated by 4-hydroxyTAM (4-OHT) in MCF-7/TAM-sensitive versus LY2 TAM/endocrine-resistant human breast cancer cells after 6 h treatment. The expression of specific miRNAs was validated by quantitative, real-time PCR. Control miRNAs and time-course studies showed important gene-, time-, and cell- specific differences in miRNA expression. Results: 97 miRNAs were differentially expressed in MCF-7 versus LY2 cells. Bioinformatic analyses to impute the biological significance of these miRNAs by identifying 36 predicted gene targets of these miRNAs from amongst those reported to be regulated by MCF-7 cells was performed and agreement was found in direction of anticipated regulation for 12 of those putative targets. Among the miRNAs differentially expressed in MCF-7 versus LY2 cells, and that putatively target mRNAs regulated by 4-OHT in MCF-7 cells, are: miRs- 10a, 21, 22, 29a, 93, 125b, 181, 200a, 200b, 200c; 205, 221, and 222. Q-PCR assessment showed agreement with microarray data: miR- 10a, 22, 125b, 181, and 222 are higher in LY2 whereas miR-21 and miR-200a are lower in LY2 than MCF-7. 4-OHT increased miR-10a, miR-21, miR-22, miR-125b, miR-181a, miR-200a, and miR-222 in MCF-7 cells and reduced miR-10a in LY2 cells. E2 reduced the miR-21expression in MCF-7 cells. Treatment with ICI 182,780 revealed that ER suppresses miR-10a, miR-21, miR-22, miR-200a, miR-221, and miR-222 expression in MCF-7 cells. Time-dependent changes in select miRNA expression were observed in control, E2, and 4-OHT-treated MCF-7 cells. ESR1/ERα was significantly lower in LY2 than MCF-7, commensurate with higher let-7 family member, miR-221, and miR-222 expression in LY2. Reflecting inverse association with miR-200 family expression, the expression of ZEB1 was higher in LY2 than MCF-7 cells and concomitantly, E-cadherin expression was absent in LY2 cells indicating that LY2 have undergone epithelial to mesenchymal transition. Conclusions: Our studies identifying miRNAs with opposite expression between the two cell lines, indicate the involvement of these miRNAs in endocrine resistance. The purpose of this experiment is: 1. To identify miRNAs that are regulated (up or down) by 4-OHT in MCF-7 and LY2 cell lines (comparison of values from EtOH versus 4-OHT treated cells). 2. To identify miRNAs that are differentially regulated under basal (EtOH) conditions between MCF-7 and LY2 cell lines. 3. To identify miRNAs that are differentially regulated by 4-OHT in MCF-7 versus LY2 cells.
| S-ECPF-GEOD-28267 | biostudies-other
Project description:The second leading cause of cancer death for women in the U.S. is breast cancer, moreover, a significant number of patients with breast tumors acquire resistance to drugs during therapy. To develop targeted therapeutic strategies to combat drug resistance it is essential to understand the basic molecular mechanisms through which cancer cells control sensitivity to chemotherapeutics. To identify new candidate genes and facilitate the discovery of novel drug resistance pathways, we have generated a resistance profile or ‘resistome’ of etoposide resitant MCF7 breast cancer cells. Differential expression of over 5000 genes (fold change > 2, P value < 0.05) indicate that several drug resistance mechanisms may be operating in these cells, including up-regulation of ABC transporter genes, down-regulation of the drug target and down-regulation of apoptotic genes. Several transcription factors such as RUNX2, SOX9, ETS1 and SMAD3 were up-regulated in the drug resistant cells. Targeted RUNX2 knockdown in the resistant cells using siRNA increased sensitivity to etoposide and also upregulated expression of pro-apoptotic genes indicating that RUNX2 could be a molecular target against etoposide resistance. Differential miRNA (microRNA) expression was observed among the drug resistant and sensitive cells suggesting that miRNA may also play a role in regulation of drug resistance. Hsa-miR-218, which targets ABCC6, was down-regulated in the drug resistant cell line. Transfection of a miR-218 mimic could down-regulate the expression of the efflux pump ABCC6 by 65% in drug resistant cells suggesting that regulation of miRNA may play an important role in etoposide resistance. RNA from etoposide-resistant MCF7 (MCF7VP) cell lines were hybridized to Affymetrix microarrays.
| S-ECPF-GEOD-28414 | biostudies-other
Project description:Testicular germ cell tumors (TGCTs) represent a well curable malignity due to their exceptional response to cisplatin (CDDP). Despite remarkable treatment results, approximately 5% of TGCT patients develop CDDP resistance and die. Exceptional curability makes TGCTs a highly valuable model system for studying the molecular mechanisms of CDDP sensitivity. Our study was aimed at revealing difference in gene expression between the CDDP-resistant and -sensitive TGCT cell lines, and hence at identifying candidate genes that could serve as potential biomarkers of CDDP response. Using gene expression array, we identified 281 genes that are differentially expressed in CDDP-resistant compared to -sensitive TGCT cell lines. The expression of 25 genes with the highest fold change was validated by RT-qPCR. Of them, DNMT3L, GAL, IGFBP2, IGFBP7, L1TD1, NANOG, NTF3, POU5F1, SOX2, WNT6, ZFP42, ID2, PCP4, SLC40A1 and TRIB3, displayed comparable expression change in gene expression array and RT-qPCR, when all CDDP-resistant TGCT cell lines were pairwise combined with all -sensitive ones. Products of the identified genes are pluripotency factors, or are involved in processes, such as cell metabolism, proliferation or migration. We propose that, after clinical validation, these genes could serve as prognostic biomarkers for early detection of CDDP response in TGCT patients.
| S-EPMC7771712 | biostudies-literature
Project description:The second leading cause of cancer death for women in the U.S. is breast cancer. Moreover, a significant number of patients with breast tumors acquire resistance to drugs during therapy. To develop targeted therapeutic strategies to combat drug resistance it is essential to understand the basic molecular mechanisms through which cancer cells control sensitivity to chemotherapeutics. To identify new candidate genes and facilitate the discovery of novel drug resistance pathways, we have generated a resistance profile or ‘resistome’ of etoposide resistant MCF7 breast cancer cells. Differential expression of over 5000 genes (fold change > 2, P value < 0.05) indicate that several drug resistance mechanisms may be operating in these cells, including up-regulation of ABC transporter genes, down-regulation of the drug target and down-regulation of apoptotic genes. Several transcription factors such as RUNX2, SOX9, ETS1 and SMAD3 were up-regulated in the drug resistant cells. Targeted RUNX2 knockdown in the resistant cells using siRNA increased sensitivity to etoposide and also upregulated expression of pro-apoptotic genes indicating that RUNX2 could be a molecular target against etoposide resistance. Differential miRNA (microRNA) expression was observed among the drug resistant and sensitive cells suggesting that miRNA may also play a role in regulation of drug resistance. Hsa-miR-218, which targets ABCC6, was down-regulated in the drug resistant cell line. Transfection of a miR-218 mimic could down-regulate the expression of the efflux pump ABCC6 by 65% in drug resistant cells suggesting that regulation of miRNA may play an important role in etoposide resistance.
2011-04-08 | GSE28413 | GEO
Project description:Cancer is considered one of the most burdensome diseases affecting lives and, hence, the economy. Breast cancer is one of the most common types of cancer. Patients with breast cancer are divided into two groups: one group responds to the chemotherapy, and the other group resists the chemotherapy. Unfortunately, the group which resists the chemotherapy is still suffering the pain associated with the severe side effects of the chemotherapy. Therefore, there is a critical need for a method to differentiate between both groups before the administration of the chemotherapy. Exosomes, the recently discovered nano-vesicles, are often used as cancer diagnostic biomarkers as their unique composition allows them to represent their parental cells, which makes them promising indicators for tumor prognosis. Exosomes contain proteins, lipids, and RNA that exist in most body fluids and are expelled by multiple cell types, including cancer cells. Furthermore, exosomal RNA has been significantly used as a promising biomarker for tumor prognosis. Herein, we have developed an electrochemical system that could successfully differentiate between MCF7 and MCF7/ADR depending on the exosomal RNA. The high sensitivity of the proposed electrochemical assay opens the door for further investigation that will address the other type of cancer cells.
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