Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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ChIP-Seq of ERalpha and DOT1L in MCF7 cell before and after EPZ004777 and ICI 182,780 treatment

ABSTRACT: ChIP-Seq of ERalpha and DOT1L in MCF7 cell before and after EPZ and ICI treatment

INSTRUMENT(S): Illumina HiSeq 2500

ORGANISM(S): Homo sapiens

SUBMITTER: Giorgio Giurato

PROVIDER: E-MTAB-6883 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Descrizione ProvvisoriaPurpose OC is characterized by a low response rate and high frequency of resistance development to currently available treatments, indicating to an urgent need for the identification of novel and more effective therapies. Several studies demonstrated DOT1L therapeutic potential in OC cells. However, DOT1L inhibitors efficacy and safety still require improvement that may be reached by combination targeting of this epigenetic modifier and its protein co-factors such as menin. Methods We applied siRNA-mediated gene knock-down and pharmacological inhibition coupled to cell proliferation assay on a panel of high grade serous OC cell lines in order to evaluate menin depletion and blockade effect on the proiferation of OC cells. We fully characterized processes, transcriptionally regulated by menin in chemotherapy-sensitive and resistant OC cells and tested the effect of simultaneous inhibition of menin and DOT1L on proliferation and transcriptional profile in these cells. Results Menin silencing and pharmacological inhibition exert an antiproliferative effect in OC cells and profound impact on transcriptome of PEO1 and PEO4 cells via down-regulation of cell cycle-related genes, aryl hydrocarbon receptor, MYC and KRAS signaling. We demonstrated assoiation of menin and DOT1L in OC cells and exsistence of a subset of genes concordantly regulated by the two proteins. Importantly, we found that simultaneous administration of DOT1L and menin-targeting drugs exerts an additive effect on the growth of chemotherapy-sensitive and -refractory relapsed OC cells driven by profound transcriptome changes related to both menin and DOT1L activities. Conclusions Described in this study results suggest that menin functionally cooperates with DOT1L in OC cells modulating transcription of genes involved in key cellular functions including cell proliferation. Dual inhibition of these two epigenetic regulators may represent a worth exploring mode to further improve the therapy response and survival of OC patients.

Project description:TGFBR1*6A is a common hypomorphic variant of the type 1 Transforming Growth Factor Beta Receptor (TGFBR1), which has been associated with increased cancer risk in some studies. Although TGFBR1*6A is capable of switching TGF-Î² growth inhibitory signals into growth stimulatory signals when stably transfected into MCF-7 breast cancer cells, TGFBR1*6A biological effects are largely unknown. To broadly explore TGFBR1*6A potential oncogenic properties, we assessed its impact on the migration and invasion of MCF-7 cells. We found that TGFBR1*6A significantly enhances MCF-7 cell migration and invasion in a TGF-Î² signaling independent manner. We set up and performed a gene array using the conditions mimicking the cell migration experiments to determine which genes in the migratory pathway were differentially regulated between the MCF-7*6A cells and the MCF-7*9A (wild type transfected) cells. The gene array identified two downregulated genes in *6A compared to *9A that are involved in cell migration and invasion: ARHGAP5, encoding ARHGAP5, and FN1, encoding fibronectin-1 (FN1). We were subsequently able to use this information in further studies in the lab. Experiment Overall Design: MCF-7 cells were stably transfected with pIRES-TGFBR1-HA-FLAG or pIRES-TGFBR1*6A-HA-FLAG. Clones were picked up and named 6A-SA5 and WT-WB1 referring to the *6A and TGFBR1 clones, respectively. We have found that the *6A mutation causes an increase in migration and invasion of MCF-7 cells which is independent of TGF-beta. We used the gene array to identify genes that were differentially regulated between the two cell lines that could lead to this phenotype. We collected RNA from samples that were serum-deprived overnight prior to being fed with complete media to mimic the conditions in the migration experiment. No exogenous growth factors were added to the media besides the normal 10% heat inactivated FBS. Samples were collected and run in triplicate (referred to in this data set as sample 1, sample 2, and sample 3). The âuntreatedâ refers to the fact that samples were grown in complete media alone with no exogenously added growth factors.

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ChIP-Seq of ERalpha and DOT1L in MCF7 cell before and after EPZ004777 and ICI 182,780 treatment

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