Targeting Transcriptional Addictions in Small Cell Lung Cancer with a Covalent CDK7 Inhibitor
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ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Richard Young
PROVIDER: E-GEOD-62614 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:Small cell lung cancer (SCLC) is an aggressive disease with high mortality. The identification of effective pharmacological strategies to target SCLC biology represents an urgent need. Using a high-throughput cellular screen of a diverse chemical library we observe that SCLC is sensitive to transcription-targeting drugs, and in particular to THZ1, a newly identified covalent inhibitor of cyclin-dependent kinase 7 (CDK7). We find that expression of super-enhancer associated transcription factor genes including MYC family proto-oncogenes and neuroendocrine lineage-specific factors are highly vulnerability to THZ1 treatment. We propose that downregulation of these transcription factors contributes, in part, to SCLC sensitivity to transcriptional inhibitors and that THZ1 represents a novel treatment paradigm for targeted SCLC therapy. ChIP-Seq for H3K27ac in small cell lung cancer lines
2014-10-23 | E-GEOD-62412 | biostudies-arrayexpress
Project description:Small cell lung cancer (SCLC) is an aggressive disease with high mortality. The identification of effective pharmacological strategies to target SCLC biology represents an urgent need. Using a high-throughput cellular screen of a diverse chemical library we observe that SCLC is sensitive to transcription-targeting drugs, and in particular to THZ1, a newly identified covalent inhibitor of cyclin-dependent kinase 7 (CDK7). We find that expression of super-enhancer associated transcription factor genes including MYC family proto-oncogenes and neuroendocrine lineage-specific factors are highly vulnerability to THZ1 treatment. We propose that downregulation of these transcription factors contributes, in part, to SCLC sensitivity to transcriptional inhibitors and that THZ1 represents a novel treatment paradigm for targeted SCLC therapy. Microarray expression in small cell lung cancer lines treated with DMSO or THZ1
2014-10-23 | E-GEOD-62609 | biostudies-arrayexpress
Project description:NF-Y, a trimeric transcription factor (TF) composed of two histone-like subunits (NF-YB (NFYB) and NF-YC (NFYC)) and a sequence-specific subunit (NF-YA), binds to the CCAAT motif, a common promoter element. Genome-wide mapping reveals 5,000-15,000 NF-Y binding sites depending on the cell type, with the NF-YA and NF-YB subunits binding asymmetrically with respect to the CCAAT motif. Despite being characterized as a proximal promoter TF, only 25% of NF-Y sites map to promoters. A comparable number of NF-Y sites are located at enhancers, many of which are tissue specific, and nearly half of NF-Y sites are in select subclasses of HERV LTR repeats. Unlike most TFs, NF-Y can access its target DNA motif in inactive (non-modified) or polycomb-repressed chromatin domains. Unexpectedly, NF-Y extensively co-localizes with FOS in all genomic contexts, and at promoters and enhancers this often occurs in the absence of JUN and the AP-1 motif. NF-Y also co-associates with a select cluster of growth-controlling and oncogenic TFs, consistent with the abundance of CCAAT motifs in the promoters of genes overexpressed in cancer. Interestingly, NF-Y and several growth-controlling TFs bind in a stereo-specific manner, suggesting a mechanism for cooperative action at promoters and enhancers. Our results indicate that NF-Y is not merely a commonly-used, proximal promoter TF, but rather performs a more diverse set of biological functions, many of which are likely to involve co-association with FOS. Scrambled control (shSCM) and NF-YA pLKO.1-shRNAs were designed by Sigma-Aldrich. The puromycin resistance cassette was replaced with an EGFP cassette. Viral production and transduction were carried out as previously described (Benatti et al. 2011). HeLaS3 cells were transduced with shSCM or shNF-YA viral supernatants, in triplicate, and cells collected after 48 hr of incubation. Total RNA was prepared by Trizol extraction and Qiagen RNeasy kit purification, converted to biotinylated aRNA and hybridized to U133 Plus 2.0 GeneChip expression arrays using the 3’ IVT Express Kit (Affymetrix, USA) following the manufacturer’s protocol. Arrays were RMA normalized (Irizarry et al. 2003), gene expression levels calculated, differential expression determined using the following R packages from the Bioconductor project: affy (Gautier et al. 2004), limma (Smyth 2004). HelaS3 cells were transduced with shSCM or shNF-YA in triplicate for 48hrs.
2012-12-31 | E-GEOD-40215 | biostudies-arrayexpress
Project description:Chromatin-organizing factors, like CTCF and cohesins, have been implicated in the control of complex viral regulatory programs. We investigated the role of CTCF and cohesin in the control of the latent to lytic switch for Kaposi's Sarcoma-Associated Herpesvirus (KSHV). We found that cohesin subunits, but not CTCF, were required for the repression of KSHV immediate early gene transcription. Depletion of cohesin subunits Rad21, SMC1, or SMC3 resulted in lytic cycle gene transcription and viral DNA replication. In contrast, depletion of CTCF failed to induce lytic transcription or DNA replication. ChiP-Seq analysis revealed that cohesins and CTCF bound to several sites within the immediate early control regions for ORF50 and more distal 5' sites that also regulate the divergently transcribed ORF45-46-47 gene cluster. Rad21 depletion led to a robust increase in ORF45 and ORF47 transcripts, with similar kinetics to that observed with chemical induction by sodium butyrate. During latency, the chromatin between the ORF45 and ORF50 transcription start sites was enriched in histone H3K4me3 with elevated H3K9ac at the ORF45 promoter and elevated H3K27me3 at the ORF50 promoter. A paused form of RNA pol II was loosely associated with the ORF45 promoter region during latency, but was converted to an active elongating form upon reactivation induced by Rad21 depletion. Butyrate-induced transcription of ORF45 and ORF47 was resistant to cyclohexamide, suggesting that these genes have immediate early features similar to ORF50. Butyrate-treatment caused the rapid dissociation of cohesins and loss of CTCF binding at the immediate early gene locus, suggesting that cohesins may be a direct target of butyrate-mediated lytic induction. Our findings implicate cohesins as a major repressor of KSHV lytic gene activation, and function coordinately with CTCF to regulate the switch between latent and lytic gene activity. Study of chromatin-organizing factors, like CTCF and cohesins.
2012-06-02 | E-GEOD-38411 | biostudies-arrayexpress
Project description:We identified a novel homozygous 15q13.3 microdeletion in a young boy with a complex neurodevelopmental disorder characterized by severe cerebral visual impairment with additional signs of congenital stationary night blindness (CSNB), congenital hypotonia with areflexia, profound intellectual disability, and refractory epilepsy. The mechanisms by which the genes in the deleted region exert their effect are unclear. In this paper we probed the role of downstream effects of the deletions as a contributing mechanism to the molecular basis of the observed phenotype. We analyzed gene expression of lymphoblastoid cells derived from peripheral blood of the proband and his relatives to ascertain the relative effects of the homozygous and heterozygous deletions. The proband with 15q13.3 homozygous deletion and his parents with 15q13.3 heterozygous deletions described in our previous report 15 were evaluated using Affymetrix aCGH-244K arrays (build hg18). The aCGH data were validated by qPCR using primers specific to CHRNA7 as previously described. 15 The extended family and comparison subjects were also screened using the CHRNA7 primers. Immortalized lymphoblastoid lines were created from the proband, his mother, father, a paternal half sister, a paternal half brother, a paternal grandmother and two paternal great uncles. Three lymphoblastoid lines from age- and sex- matched normally developing comparison subjects with normal chromosomes were obtained from the Coriell cell repository [(AG14724 (8 yo), AG14948 (10 yo), AG14980 (9 yo)] were used for gene expression comparison with the proband. The samples from the father, mother and paternal grandmother with heterozygous 15q13.3 deletions were compared to samples from lymphoblastoid cells from adult male and female subjects respectively (normal copy number for 15q13.3 determined by qPCR).
2013-02-17 | E-GEOD-43583 | biostudies-arrayexpress
Project description:The goal was to determine how IL-12 affects gene expression by murine CTL. Experiment Overall Design: Following RBC lysis, splenocytes from OT-1 TCR transgenic mice (5 million/ml) were cultured in six well-plates (5-6ml/well) in IMDM supplemented with 10% FCS and 55μM 2-ME with 1μM SIINFEKL. After 4 days, the cells were harvested by purification with Ficoll-Paque PLUS and placed in fresh medium containing 0 or 20ng/ml rmIL-12. Twenty-four hours later the cells were harvested and used in experiments. The live cells harvested were >98% CD8+Vα2+ (OT-1).
2009-02-08 | E-GEOD-13173 | biostudies-arrayexpress
Project description:The 3-Deazaneplanocin A (DZNep), one of S-adenosylhomocysteine (AdoHcy) hydrolase inhibitors, has shown antitumor activities in a broad range of solid tumors and acute myeloid leukemia. Here, we examined its effects on multiple myeloma (MM) cells and found that, at 500 nM, it potently inhibited growth and induced apoptosis in 2 of 8 MM cell lines. RNA from un-treated and DZNep treated cells was profiled by Affymetrix HG-U133 Plus 2.0 microarray and genes with a significant change in gene expression were determined by significance analysis of microarray (SAM) testing. ALOX5 was the most down-regulated gene (5.8-fold) in sensitive cells and was expressed at low level in resistant cells. The results were corroborated by quantitative RT-PCR. Western-blot analysis indicated ALOX5 was highly expressed only in sensitive cell line H929 and greatly decreased upon DZNep treatment. Ectopic expression of ALOX5 reduced sensitivity to DZNep in H929 cells. Furthermore, down-regulation of ALOX5 by RNA interference could also induce apoptosis in H929. Gene expression analysis on MM patient dataset indicated ALOX5 expression was significantly higher in MM patients compared to normal plasma cells. We also found that Bcl-2 was overexpressed in DZNep insensitive cells, and cotreatment with DZNep and ABT-737, a Bcl-2 family inhibitor, synergistically inhibited growth and induced apoptosis of DZNep insensitive MM cells. Taken together, this study shows one of mechanisms of the DZNep efficacy on MM correlates with its ability to down-regulate the ALOX5 levels. In addition, DZNep insensitivity might be associated with overexpression of Bcl-2, and the combination of ABT-737 and DZNep could synergistically induced apoptosis. These results suggest that DZNep may be exploited therapeutically for a subset of MM. Cells were treated with 0.5 μmol/L DZNep for 48 h. Total RNA was extracted by using the Qiagen RNeasy Mini kit (Germany). Gene expression was performed using the GeneChip® Human Genome U133A Array (Affymetrix) following the manufacturer’s instructions. Data analysis was performed using GeneSpring software from Agilent Technologies.
2011-07-04 | E-GEOD-26921 | biostudies-arrayexpress
Project description:Purpose: The phosphoinositide 3-kinase (PI3K) pathway is fundamental for cell proliferation and survival and is frequently altered and activated in neoplasia, including carcinomas of the lung. In this study we investigated the potential of targeting the catalytic class IA PI3K isoforms in small cell lung cancer (SCLC), which is the most aggressive of all lung cancer types. Experimental Design: The expression of PI3K isoforms in patient specimens was analyzed. The effects on SCLC cell survival and downstream signaling were determined following PI3K isoform inhibition by selective inhibitors or down-regulation by small interfering RNA. Results: Over-expression of the PI3K isoforms p110 -alpha and p110-alpha was shown by immunohistochemistry in primary SCLC tissue samples. Targeting the PI3K p110 -alpha with RNA interference (RNAi) or selective pharmacological inhibitors resulted in strongly affected cell proliferation of SCLC cells in vitro and in vivo, while targeting p110-alph was less effective. Inhibition of p110 -alpha also resulted in increased apoptosis and autophagy, which was accompanied by decreased phosphorylation of Akt and components of the mammalian target of rapamycin (mTOR) pathway, such as the ribosomal S6 protein, and the eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1). A DNA microarray analysis revealed that p110-alpha inhibition profoundly affected the balance of pro- and anti-apoptotic Bcl-2 family proteins. Finally, p110 -alpha inhibition led to impaired SCLC tumor formation and vascularization in vivo. Conclusion: Together our data demonstrate the key involvement of the PI3K isoform p110 -alpha in multiple tumor-promoting processes in SCLC. 3 control samples, 3 samples for two treaments
2012-12-01 | E-GEOD-40564 | biostudies-arrayexpress
Project description:The trimeric transcription factor -TF- NF-Y binds to the CCAAT box, an element enriched in promoters of genes overexpressed in tumors. The NF-YC subunit was recently identified as an oncogene in choroid plexus carcinomas. Previous studies on the NF-Y regulome identified the general term metabolism as significantly enriched. We dissect here in detail the targeting of metabolic genes by integrating analysis of NF-Y genomic binding and profilings after inactivation of NF-Y subunits in different cell types. NF-Y comprehensively controls de novo biosynthetic pathways of lipids, teaming up with the master SREBPs regulators. It activates glycolytic genes, but, surprisingly, is neutral, or represses mitochondrial respiratory genes. NF-Y targets the SOCG (Serine, One Carbon, Glycine) and Glutamine pathways, as well as genes involved in the biosynthesis of polyamines and purines. Within the different pathways, cancer-driving nodes are under NF-Y positive control. Altogether, these data delineate a coherent strategy to promote expression of metabolic genes fuelling anaerobic energy production, and other anabolic pathways commonly altered in cancer cells. Scrambled control (shSCM) and NF-YA pLKO.1-shRNAs were designed by Sigma-Aldrich. The puromycin resistance cassette was replaced with an EGFP cassette. Viral production and transduction were carried out as previously described (Benatti et al. 2011). H322 and HCT116 were transduced with shSC or shNF-YA viral supernatants, in triplicates and cells collected after 72 hours. RNA was prepared according to Affymetrix standard protocol and hybridized to Hu-Gene 2.0 expression arrays. H322 and HCT116 were transduced with shSC and shNF-YA and harvest after 72 hours
2016-07-03 | E-GEOD-70543 | biostudies-arrayexpress
Project description:Mouse BAL alveolar macrophages from pregnant E14 and control dams are compared to RAW cells cultured with estradiol to determine shared gene expression change patterns induced by pregnancy and estrogen BAL cells from intact pregnant and control mice were directly processed for RNA extraction. RAW cells were cultured overnight with estradiol or vehicle, then RNA was exctracted.
2014-01-01 | E-GEOD-52649 | biostudies-arrayexpress