Hacking the Cancer Genome: Profiling Therapeutically Actionable Long Non-coding RNAs Using CRISPR-Cas9 Screening

Esposito, Roberta; Bosch, Núria Vergés; Lanzós, Andrés; Polidori, Taisia; Pulido-Quetglas, Carlos; Johnson, Rory

doi:10.1016/j.ccell.2019.01.019

Cited by 170 publications

(137 citation statements)

References 156 publications

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“…Numerous lncRNAs are revealed to be deregulated in many diseases . Furthermore, complex and various functions of lncRNAs in different conditions have also been discovered …”

Section: Introductionmentioning

confidence: 99%

Long non‐coding RNA LINC00467 drives hepatocellular carcinoma progression via inhibiting NR4A3

Wang

Guo

Nampoukime

et al. 2020

J Cellular Molecular Medi

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Hepatocellular carcinoma (HCC) is a main cause of cancer‐related deaths globally. Long non‐coding RNAs (lncRNAs) play important roles in diverse cancers. Our previous microarray‐based lncRNA profiling showed that LINC00467 was highly expressed in HCC. Here, we further explored the expression, role and functional mechanism of lncRNA LINC00467 in HCC. Our findings revealed that LINC00467 was up‐regulated in HCC tissues and HCC cell lines. Increased expression of LINC00467 was positively associated with tumour size and vascular invasion. In vitro functional experiments revealed that LINC00467 accelerated HCC cell proliferation, cell cycle progression and migration and reduced HCC cell apoptosis. In vivo functional assays revealed that LINC00467 drove HCC xenograft growth and HCC cell proliferation and repressed HCC cell apoptosis in vivo. Moreover, LINC00467 inhibited NR4A3 post‐transcriptionally via interacting with NR4A3 mRNA to form double‐stranded RNA, which was further degraded by Dicer. The expression of NR4A3 was inversely associated with LINC00467 in HCC tissues. Functional rescue assays found that restore of NR4A3 expression blocked the oncogenic roles of LINC00467 in HCC. Taken together, our results demonstrated that lncRNA LINC00467 was a novel highly expressed and oncogenic lncRNA in HCC via inhibiting NR4A3. Targeting LINC00467 or enhancing NR4A3 may be potential therapeutic strategies against HCC.

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“…Numerous lncRNAs are revealed to be deregulated in many diseases . Furthermore, complex and various functions of lncRNAs in different conditions have also been discovered …”

Section: Introductionmentioning

confidence: 99%

Long non‐coding RNA LINC00467 drives hepatocellular carcinoma progression via inhibiting NR4A3

Wang

Guo

Nampoukime

et al. 2020

J Cellular Molecular Medi

View full text Add to dashboard Cite

show abstract

“…On the other hand, a subset prognostic lncRNAs may be functional in cells and act as functional ‘drivers’ that activate oncogenic processes or inhibit tumor suppressive pathways through interactions with DNA, RNA and proteins. However, further experiments are needed to validate the prognostic lncRNAs as drivers of cancer phenotypes, such as large-scale genome editing screens that are increasingly targeting the non-coding genome encoding lncRNAs [60]. Our findings of prognostic lncRNAs are ultimately limited by the transcriptional and clinical information that was available for inference and validation.…”

Section: Discussionmentioning

confidence: 99%

Comprehensive analysis of lncRNAs reveals candidate prognostic biomarkers in multiple cancer types

Isaev

Jiang

Lee

et al. 2019

Preprint

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12Long non-coding RNAs (lncRNAs) are increasingly recognized as functional units in can-13 cer pathways and powerful molecular biomarkers, however most lncRNAs remain un-14 characterized. Here we performed a systematic discovery of prognostic lncRNAs in 9,326 15 patient tumors of 29 types using a proportional-hazards elastic net machine-learning 16 framework. lncRNAs showed highly tissue-specific transcript abundance patterns. We 17 identified 179 prognostic lncRNAs whose abundance correlated with patient risk and im-18 proved the performance of common clinical variables and molecular tumor subtypes. 19 Pathway analysis revealed a large diversity of the high-risk tumors stratified by lncRNAs 20and suggested their functional associations. In lower-grade gliomas, discrete activation 21 of HOXA10-AS indicated poor patient prognosis, neurodevelopmental pathway activation 22 and a transcriptomic similarity to glioblastomas. HOXA10-AS knockdown in patient-de-23 rived glioblastoma cells caused decreased cell proliferation and deregulation of glioma 24 driver genes and proliferation pathways. Our study underlines the pan-cancer potential 25 of the non-coding transcriptome for developing molecular biomarkers and innovative 26 therapeutic strategies. 27 28 cancer patient prognosis with transcript abundance of protein-coding genes and their genetic 61 and epigenetic alterations [22][23][24][25]. However, associations of lncRNAs with cancer patient sur-62 vival and biological function remain largely unexplored. A recent study characterized recurrent 63 hypomethylation patterns affecting a thousand lncRNAs in the TCGA PanCanAtlas cohort and 64 identified the EPIC1 lncRNA as a marker of poor prognosis in a subset of breast cancers [26]. 65Another TCGA study associated mutations and transcript abundance profiles of lncRNAs with 66 regulatory networks and molecular pathways and nominated candidate oncogenic and tumor 67 suppressive lncRNAs, some of which were functionally validated in cancer cell lines [27]. Analy-68 sis of cell-cycle correlated lncRNAs revealed a subset of S-phase enriched lncRNAs whose 69 transcript abundance profiles correlated with patient survival in multiple TCGA cohorts [28]. 70However, those studies did not analyze robust prognostic performance of lncRNAs using ma-71 chine-learning and cross-validation approaches, indicating further potential to systematically dis-72 cover lncRNAs as candidate prognostic biomarkers of multiple cancer types. 73Here we evaluated the transcript abundance profiles of nearly 6,000 lncRNAs as prognostic bi-74 omarkers in human cancers. Using a comprehensive machine-learning analysis, we compiled a 75 robust catalogue of prognostic lncRNAs across nearly 10,000 tumors of 29 types from the 76 TCGA PanCanAtlas project [22, 29]. The majority of our candidate lncRNAs showed improved 77 prognostic potential compared to standard clinical features and molecular tumor subtypes. We 78 associated prognostic lncRNAs with large-scale deregulation of hallmark cancer pathways, re-79 vealing e...

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“…Pooled CRISPR screens employ phenotypic readouts, often in the form of cell proliferation 3,27 . To test whether improved CRISPR-del translates into stronger phenotypes, we developed a reporter assay capable of quantifying the phenotypic effect of CRISPR-del in terms of cell death.…”

Section: Dna-pk Inhibition In the Context Of High-throughput Pooled Smentioning

confidence: 99%

“…Pooled screens require that both sgRNAs are encoded in a single vector to ensure their simultaneous delivery, and are typically performed under conditions of low multiplicity-of-infection (MOI), where each cell carries a single lentiviral insertion 18,20,25,27,28 .…”

Section: Introductionmentioning

confidence: 99%

Enhancing CRISPR deletion via pharmacological delay of DNA-PK

Bosch

Medová

Esposito

et al. 2020

Preprint

Self Cite

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CRISPR-Cas9 deletion (CRISPR-del) is the leading approach for eliminating DNA from mammalian cells and underpins a variety of genome-editing applications. Target DNA, defined by a pair of double strand breaks (DSBs), is removed during non-homologous end-joining (NHEJ). However, the low efficiency of CRISPR-del results in laborious experiments and false negative results. Using an endogenous reporter system, we demonstrate that temporary inhibition of DNA-dependent protein kinase (DNA-PK) -an early step in NHEJ -yields up to 17-fold increase in DNA deletion. This is observed across diverse cell lines, gene delivery methods, commercial inhibitors and guide RNAs, including those that otherwise display negligible activity. Importantly, the method is compatible with pooled functional screens employing lentivirally-delivered guide RNAs. Thus, delaying the kinetics of NHEJ relative to DSB formation is a simple and effective means of enhancing CRISPR-deletion.

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Hacking the Cancer Genome: Profiling Therapeutically Actionable Long Non-coding RNAs Using CRISPR-Cas9 Screening

Cited by 170 publications

References 156 publications

Long non‐coding RNA LINC00467 drives hepatocellular carcinoma progression via inhibiting NR4A3

Long non‐coding RNA LINC00467 drives hepatocellular carcinoma progression via inhibiting NR4A3

Comprehensive analysis of lncRNAs reveals candidate prognostic biomarkers in multiple cancer types

Enhancing CRISPR deletion via pharmacological delay of DNA-PK

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