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Unknown,Transcriptomics,Genomics,Proteomics

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Preclinical and clinical studies of estrogen deprivation support the PDGF/Abl pathway as a novel therapeutic target for overcoming endocrine resistance in breast cancer

ABSTRACT: The human oestrogen-positive breast cancer cell line MCF7 was cultured in phenol red-free RPMI medium supplemented with 10% fetal bovine serum, 10mg/ml insulin and 1nM estradiol (E2) and was referred to as wild-type MCF7. The wt-MCF7 cells were passaged weekly and medium was replenished every two to three days. To model acquisition of resistance to long term estrogen deprivation (LTED) on an aromatase inhibitor (AI), wt-MCF7 cells were cultured in phenol red-free RPMI medium supplemented with 10% dextran charcoal-stripped bovine serum (DCC) and 10mg/ml insulin. RNA was extracted from the LTED monolayers using RNeasy columns (Qiagen) according to manufacturerﾒs protocol. RNA amplification, labeling and hybridization on HumanWG-6 v3 Expression BeadChips were performed according to the manufacturer's instructions (http://www.illumina.com) to assess changes in gene expression during adaptation to LTED.

ORGANISM(S): Homo sapiens

SUBMITTER: Zara Ghazoui

PROVIDER: E-MTAB-922 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Quantitative proteomics on wt-MCF7 versus their LTED derivatives using dimethyl labelling

Project description:Wild type (wt) MCF7 cells, modelling breast cancer at primary diagnosis, were cultured in phenol red-free RPMI supplemented with 10% FBS and 1nM estradiol (E2). Long-term oestrogen deprived (LTED) cell lines, which model resistance to endocrine therapy, were cultured in phenol red-free RPMI in the absence of exogenous E2 and supplemented with 10% dextran charcoal-stripped bovine serum (DCC). Samples were harvested at baseline and at the point of resistance (LTED). To reveal differential protein abundances between wt-MCF7 and MCF7 LTED, the peptides were labelled with chemical labelling and underwent fractionation using OFFGEL electrophoresis.

2016-06-01 | PXD004085 | Pride

ER-RIME in wt-MCF7 and MCF7-LTED

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2020-07-13 | PXD004648 | Pride

Comparative shotgun proteomics in wt-SUM44 and SUM44-LTED, using RIME and dimethyl-labelling

Project description:Wild type (wt) SUM44 cells, modelling breast cancer at primary diagnosis, were cultured in phenol red-free RPMI supplemented with 10% FBS and 1nM estradiol (E2). Long-term oestrogen deprived (LTED) cell lines, which model resistance to endocrine therapy, were cultured in phenol red-free RPMI in the absence of exogenous E2 and supplemented with 10% dextran charcoal-stripped bovine serum (DCC). Samples were harvested at baseline and at the point of resistance (LTED). To reveal differential protein abundances between wt-SUM44 and SUM44-LTED, the peptides were labelled with dimethyl labelling and underwent fractionation using OFFGEL electrophoresis. In order to reveal the ER-interactome, RIME (rapid immunoprecipitation mass spectrometry of endogenous proteins) was conducted in wt-SUM44 and SUM44-LTED.

2017-12-11 | PXD004807 | Pride

Elacestrant demonstrates strong anti-estrogenic activity in PDX models of estrogen receptor positive endocrine-resistant and fulvestrant-resistant breast cancer

Project description:Human breast cancer cell lines were cultured in phenol red-free RPMI1640 medium supplemented with 10% fetal bovine serum and 1nM E2. All cell lines were banked in multiple aliquots to reduce the risk of phenotypic drift and identity confirmed using STR. Cells were routinely screened for mycoplasma contamination. Long-term E deprived (LTED) cells modelling resistance to an AI were derived from all parental cell lines as previously described. Two models of MCF7-LTED cells were available: one harbouring a Y537C mutation in ESR1 and the other with wt-ESR1. SUM44-LTED was shown to harbour a natural heterozygous ESR1Y537S mutation. Elacestrant resistant cell lines were generated by growing parental cells (MCF7-LTEDY537C) long-term in the presence of RPMI1640 containing 10% DCC + 0.1nM E2 + 1μM elacestrant. All cell lines were stripped of steroids for 48-72 hours prior to the start of experiments.

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TUMOUR KINOME RE-WIRING GOVERNS RESISTANCE TO PALBOCICLIB IN OESTROGEN RECEPTOR POSITIVE BREAST CANCERS, HIGHLIGHTING NEW THERAPEUTIC MODALITIES

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2020-04-29 | PXD005514 | Pride

Resistance to Palbociclib requires multiple targetable mechanisms highlighting the potential of drug holidays and drug switching to improve therapeutic outcome

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2020-03-25 | PXD005611 | Pride

Coativator Function Defines the Active Estrogen Receptor Alpha Cistrome

Project description:Genome-wide ChIP-on-Chip against RNA Pol II in untreated MCF7 cells (phenol-red free media) and H3R17me2 in untreated and estrogen (45 min) treated cells.

2009-12-09 | GSE19328 | GEO

Drug specific epigenetic reprogramming leads to increased cellular invasion in ERÎ± positive breast cancer via de novo cholesterol biosynthesis.

Project description:Endocrine therapy resistance remains a critical problem in the treatment of estrogen receptor alpha (ERÎ±) breast cancer. Endocrine therapies target ERÎ± via different modes of action. Drug resistance involves drug specific remodeling of the transcriptional and regulatory landscape. Using epigenomics and transcriptomics, we demonstrate that resistance to aromatase inhibitors (AI) induces phenotypical changes through epigenetic activation of cholesterol biosynthesis (CB) and keratin 80. Epigenetic activation is stable and involves both large topological domains and punctuated activation of single enhancers and super-enhancers. Specialized cancer cells expressing high levels of keratin 80 lead invasion through the extracellular matrix. Strikingly, we demonstrate that anti-cholesterol strategies can effectively arrest breast cancer invasion. Our work identifies a robust strategy to target resistant invasive breast cancer. All cell lines were grown in their respective media until they reached 70% confluency. Specifically, MCF7 were grown in DMEM+10% FBS and 1% Pen-Sprep-Glutamine. MCF7T as MCF7 with the addition of 100nM Tamoxifen. MCF7F as MCF7 with the addition of 100nM Fulvestrant. LTED were grown in DMEM without phenol + 10% DCS-FBS and1% Pen-Sprep-Glutamine. LTEDT as LTED with the addition of 100nM Tamoxifen. LTEDF as LTED with the addition of 100nM of Fulvestrant. All chemicals are purchased from Sigma. ChIP-seq was performed using Illumina methodology.

2015-12-31 | E-GEOD-60517 | biostudies-arrayexpress

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2013-10-25 | GSE51448 | GEO

Expression data from androgen treated LNCaP cells

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