Determine whether AsiDNA and belinostat alter chromatin accessbility genome-wide in glioblastoma cells. [NicE-Seq]
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ABSTRACT:
PROVIDER: PRJNA1107962 | ENA |
REPOSITORIES: ENA
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Project description:An effective long-term inhibition of multiple DNA repair signals is required to design a novel and effective therapeutic for glioblastoma (GBM), a highly DNA repair ‘addicted’ cancer. AsiDNA, a double-strand DNA break (DSB) mimetic, is an innovative approach that shuts down the entire DNA repair system in cancer cells by sequestering DSB repair factors. We showed that inhibition of class I histone deacetylases (HDACs) dislodges repair factors from sites of DNA damage. We therefore tested whether AsiDNA and pan HDAC inhibitor Belinostat combination can impart a chromatin-based long-term DNA damage and impair DNA repair in GBM cells. We performed mass spectrometry with chromatin isolated from AsiDNA and Belinostat and found that Belinostat reduces histone H1.2 levels. To this end, we assessed whether AsiDNA and Belinostat alter histone H1.2 occupancy genome-wide. In order to understand the long-term effects of AsiDNA on DNA repair, we treated U87 cells with AsiDNA for 6 days followed by 6 days recovery from the drug. We used this protocol to create long-term AsiDNA treated cells (LTAU87) according to the clinical trial protocol. We performed NicE-seq with LTAU87 cells and MTU87 (mock treated U87; control) in the presence or absence of belinostat treatment in order to address whether they alter the global chromatin structure.
2024-05-13 | GSE266621 | GEO
Project description:An effective long-term inhibition of multiple DNA repair signals is required to design a novel and effective therapeutic for glioblastoma (GBM), a highly DNA repair ‘addicted’ cancer. AsiDNA, a double-strand DNA break (DSB) mimetic, is an innovative approach that shuts down the entire DNA repair system in cancer cells by sequestering DSB repair factors. We showed that inhibition of class I histone deacetylases (HDACs) dislodges repair factors from sites of DNA damage. We therefore tested whether AsiDNA and pan HDAC inhibitor Belinostat combination can impart a chromatin-based long-term DNA damage and impair DNA repair in GBM cells. We performed mass spectrometry with chromatin isolated from AsiDNA and Belinostat and found that Belinostat reduces histone H1.2 levels. To this end, we assessed whether AsiDNA and Belinostat alter histone H1.2 occupancy genome-wide. In order to understand the long-term effects of AsiDNA on DNA repair, we treated U87 cells with AsiDNA for 6 days followed by 6 days recovery from the drug. We used this protocol to create long-term AsiDNA treated cells (LTAU87) according to the clinical trial protocol. We performed H1.2 ChIP-Seq with LTAU87 cells and MTU87 (mock treated U87; control) in the presence and absence of belinostat in order to address whether they alter the global chromatin structure.
2024-05-13 | GSE266619 | GEO
Project description:Determine whether AsiDNA and belinostat alter H1.2 occupancy genome-wide. [ChIP-Seq]
| PRJNA1107964 | ENA
Project description:An effective long-term inhibition of multiple DNA repair signals is required to design a novel and effective therapeutic for glioblastoma (GBM), a highly DNA repair ‘addicted’ cancer. AsiDNA, a double-strand DNA break (DSB) mimetic, is an innovative approach that shuts down the entire DNA repair system in cancer cells by sequestering DSB repair factors. We showed that inhibition of class I histone deacetylases (HDACs) dislodges repair factors from sites of DNA damage. We therefore tested whether AsiDNA and pan HDAC inhibitor Belinostat combination can impart a chromatin-based long-term DNA damage and impair DNA repair in GBM cells. In order to understand the long-term effects of AsiDNA on DNA repair, we treated U87 cells with AsiDNA for 6 days followed by 6 days recovery from the drug. We used this protocol to create long-term AsiDNA treated cells (LTAU87) according to the clinical trial protocol. We performed RNA-Seq with LTAU87 cells and MTU87 (mock treated U87; control) in the presence and absence of belinostat.
2024-05-13 | GSE266620 | GEO
Project description:Effect of AsiDNA and belinostat on gene expression in U87 cells. [RNA-Seq]
| PRJNA1107963 | ENA
Project description:Transcriptome analysis revealed a major change in gene expression with a large excess of genes down regulated in 3 independent AsiDNA-treated “evolved” populations as compared to three independent not-treated “naïve” populations The Achilles heel of anticancer treatments is intrinsic or acquired resistance. Among many targeted therapies, the DNA repair inhibitors show limited efficacy due to rapid emergence of resistance. We examined evolution of cancer cells and tumors treated with AsiDNA, a new DNA repair inhibitor targeting all DNA break repair pathways. Effects of AsiDNA or Olaparib were analyzed in various cell lines. Frequency of AsiDNA- and olaparib-resistant clones was measured after 2 weeks of continuous treatment in KBM7 haploid cells. Cell survivals were also measured after one to six cycles of 1-week treatment and 1-week recovery in MDA-MB-231 and NCI-H446. Transcriptomes of cell populations recovering from cyclic treatments or mock treatment were compared. MDAMB- 231 xenografted models were treated with three cycles of AsiDNA to monitor the effects of treatment on tumor growth and transcriptional modifications. No resistant clones were selected after AsiDNA treatment (frequency b 3x10−8) in treatment conditions that generate resistance to olaparib at a frequency of 7.2x10−7 resistant clones per treated cell. Cyclic treatments promote cumulative sensitivity characterized by a higher mortality of cells having undergone previous treatment cycles. This sensitization was stable, and transcriptome analysis revealed a major gene downregulation with a specific overrepresentation of genes coding for targets of DNA-PK. Such changes were also detected in tumor models which showed impaired growth after cycles of AsiDNA treatment.
2020-08-20 | GSE144023 | GEO
Project description:AsiDNA, a cholesterol-coupled oligonucleotide mimicking double-stranded DNA breaks, was developed to sensitize tumour cells to radio- and chemotherapy. This drug acts as a decoy hijacking the DNA damage response. Previous studies have demonstrated that standalone AsiDNA administration is well tolerated with no additional adverse effects when combined with chemo- and/or radiotherapy. This lack of normal tissue complication encouraged further examination into the role of AsiDNA in normal cells. The delayed development of pulmonary fibrosis after FLASH-RT compared to CONV-RT might be a known phenomenon but the complexity behind this specific tissue response is completely unidentified. Radiation has been shown to impact various cell types presented in the lung, resulting in changes in cellular activity and numerous pathway activations. Current unpublished data suggest a clear variety between the CONV and FLASH-RT gene signatures examined using single cell RNA sequencing. As the combined treatment of AsiDNA with CONV-RT resulted in a delay in radiation induced lung fibrosis compared to CONV-RT standalone, it is to be questioned if the signature of AsiDNA CONV-RT is similar to FLASH-RT or CONV-RT. To characterize the similarity in cellular changes and expression signatures in lungs 5 months post CONV-RT, FLASH-RT, AsiDNA CONV-RT or non-irradiated (Control) treatment. Single cell RNA sequencing of irradiated lungs revealed a closer resemblance between CONV AsiDNA and FLASH-RT in profibrotic gene signatures within the fibroblast and macrophages population, in comparison to CONV-RT standalone. Collectively, this information indicates that the activity of AsiDNA and FLASH-RT could draw upon identical mechanism interference to result in the protective capacities within the normal tissue.
2024-02-27 | GSE240510 | GEO
Project description:Enhancer-promoter communication is known to regulate spatiotemporal gene expression. Here, we present NicE-C, a method that leverages nicking enzyme mediated open chromatin profiling and chromosome conformation capture to enable robust and cost-effective detection of chromatin interactions among open chromatin regions (e.g., between enhancers and promoters). Demonstrating its application utility, NicE-C revealed dynamic enhancer-promoter interactions associated with gene expression changes after TNFɑ stimulation in HeLa cells and during aging process in mice kidney.
2022-01-18 | GSE176066 | GEO
Project description:Mammalian chromatin is dynamic and contains structural information that leads to transcriptional regulation of genes by recruiting transcription factors and altering nucleosome positioning. Here, we describe open chromatin mapping using Nicking Enzyme assisted sequencing (NicE-seq) for integrative epigenomic analysis. NicE-seq captures open chromatin sites in both fixed and native cells and reveals the genomic location of open chromatin sites (OCS) and transcription factor occupancy at single nucleotide resolution, with as few as 250 HCT116 cells. In HCT116 cells a large percentage of the OCS were coincident with DHS (DNaseI hypersensitive site) and ATAC-seq sites. OCSs correlated well with RNA pol II occupancy and transcriptionally active chromatin marks, while displaying a pattern contrasting to CpG methylation. ChIP CTCF peaks common to OCS and DHS displayed a strong correlation with H3K4me3 and H3K27ac marks. Further peaks unique to OCS and ChIP CTCF peaks also correlated strongly with H3K4me3, H3K27ac and higher transcription. Similar results were also obtained for Max and Sp1 transcription factors. Using NicE-seq we have demonstrated that HCT116 cells, treated with anti cancer drug decitabine, displayed time dependent accumulation of OCS coinciding with hypomethylation of the genome, particularly in SINE and satellite repetitive DNA. Differentially methylated regions (DMRs) were more pronounced in promoters, 5’UTR, CpG islands and exons. In summary, sequence specificity offered by a nicking enzyme allows a means to study open chromatin structure and hypomethylation of genomes, revealing the open chromatin landscape in cellular context.
2017-05-24 | GSE97889 | GEO
Project description:Glucocorticoid resistance (GCR), i.e. unresponsiveness to the beneficial anti-inflammatory activities of the glucocorticoid receptor (GR), poses a serious problem in the treatment of inflammatory diseases. One possible solution to try and overcome GCR, is to identify molecules that prevent or revert GCR by hyper-stimulating the biological activity of the GR. To this purpose, we screened for compounds that potentiate the dexamethasone (Dex)-induced transcriptional activity of GR. To monitor GR transcriptional activity, the screen was performed using the lung epithelial cell line A549 in which a glucocorticoid responsive element (GRE) coupled to a luciferase reporter gene construct was stably integrated. Histone deacetylase inhibitors (HDACi) such as Vorinostat and Belinostat are two broad-spectrum HDACi that strongly increased the Dex-induced luciferase expression in our screening system. In sharp contrast herewith, results from a genome-wide transcriptome analysis of Dex-induced transcripts using RNAseq, revealed that Belinostat impairs the ability of GR to transactivate target genes. The stimulatory effect of Belinostat in the luciferase screen further depends on the nature of the reporter construct. In conclusion, a profound discrepancy was observed between HDACi effects on two different synthetic promoter-luciferase reporter systems. The favorable effect of HDACi on gene expression should be evaluated with care, when considering them as potential therapeutic agents.
2017-03-16 | GSE96649 | GEO