Dataset Information

Genome-Wide CRISPR-Cas9 Screening Identifies NF-?B/E2F6 Responsible for EGFRvIII-Associated Temozolomide Resistance in Glioblastoma.

ABSTRACT: Amplification of epidermal growth factor receptor (EGFR) and active mutant EGFRvIII occurs frequently in glioblastoma (GBM) and contributes to chemo/radio-resistance in various cancers, especially in GBM. Elucidating the underlying molecular mechanism of temozolomide (TMZ) resistance in GBM could benefit cancer patients. A genome-wide screening under a clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 library is conducted to identify the genes that confer resistance to TMZ in EGFRvIII-expressing GBM cells. Deep sgRNA sequencing reveals 191 candidate genes that are responsible for TMZ resistance in EGFRvIII-expressing GBM cells. Notably, E2F6 is proven to drive a TMZ resistance, and E2F6 expression is controlled by the EGFRvIII/AKT/NF-?B pathway. Furthermore, E2F6 is shown as a promising therapeutic target for TMZ resistance in orthotopic GBM cell line xenografts and GBM patient-derived xenografts models. After integrating clinical data with paired primary-recurrent RNA sequencing data from 134 GBM patients who received TMZ treatment after surgery, it has been revealed that the E2F6 expression level is a predictive marker for TMZ response. Therefore, the inhibition of E2F6 is a promising strategy to conquer TMZ resistance in GBM.

SUBMITTER: Huang K

PROVIDER: S-EPMC6724471 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

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Genome-Wide CRISPR-Cas9 Screening Identifies NF-κB/E2F6 Responsible for EGFRvIII-Associated Temozolomide Resistance in Glioblastoma.

Huang Kai K Liu Xing X Li Yansheng Y Wang Qixue Q Zhou Junhu J Wang Yunfei Y Dong Feng F Yang Chao C Sun Zhiyan Z Fang Chuan C Liu Chaoyong C Tan Yanli Y Wu Xudong X Jiang Tao T Kang Chunsheng C

Advanced science (Weinheim, Baden-Wurttemberg, Germany) 20190724 17

Amplification of epidermal growth factor receptor (EGFR) and active mutant EGFRvIII occurs frequently in glioblastoma (GBM) and contributes to chemo/radio-resistance in various cancers, especially in GBM. Elucidating the underlying molecular mechanism of temozolomide (TMZ) resistance in GBM could benefit cancer patients. A genome-wide screening under a clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 library is conducted to identify the genes that confer resistance to TMZ ...[more]

PMID: 31508283

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Project description:BackgroundGlioblastoma (GBM) is an aggressive brain tumor with universal recurrence and poor prognosis. The recurrence is largely driven by chemoradiation resistant cancer stem cells (CSCs). Epidermal growth factor receptor (EGFR) and its mutant EGFRvIII are amplified in ~ 60% and ~ 30% of GBM patients, respectively; however, therapies targeting EGFR have failed to improve disease outcome. EGFRvIII-mediated cross-activation of tyrosine kinase receptor, cMET, regulates GBM CSC maintenance and promote tumor recurrence. Here, we evaluated the efficacy of pan-EGFR inhibitor afatinib and Temozolomide (TMZ) combination on GBM in vitro and in vivo.MethodsWe analyzed the effect of afatinib and temozolomide (TMZ) combination on GBM cells U87MG and U251 engineered to express wild type (WT) EGFR, EGFRvIII or EGFRvIII dead kinase, CSCs isolated from U87 and U87EGFRvIII in vitro. The therapeutic utility of the drug combination was investigated on tumor growth and progression using intracranially injected U87EGFRvIII GBM xenografts.ResultsAfatinib and TMZ combination synergistically inhibited the proliferation, clonogenic survival, motility, invasion and induced senescence of GBM cells compared to monotherapy. Mechanistically, afatinib decreased U87EGFRvIII GBM cell proliferation and motility/invasion by inhibiting EGFRvIII/AKT, EGFRvIII/JAK2/STAT3, and focal adhesion kinase (FAK) signaling pathways respectively. Interestingly, afatinib specifically inhibited EGFRvIII-cMET crosstalk in CSCs, resulting in decreased expression of Nanog and Oct3/4, and in combination with TMZ significantly decreased their self-renewal property in vitro. More interestingly, afatinib and TMZ combination significantly decreased the xenograft growth and progression compared to single drug alone.ConclusionOur study demonstrated significant inhibition of GBM tumorigenicity, CSC maintenance in vitro, and delayed tumor growth and progression in vivo by combination of afatinib and TMZ. Our results warrant evaluation of this drug combination in EGFR and EGFRvIII amplified GBM patients.

Project description:Background Glioblastoma (GBM) is a type of highly malignant brain tumor that is known for its significant intratumoral heterogeneity, meaning that there can be a high degree of variability within the tumor tissue. Despite the identification of several subtypes of GBM in recent years, there remains to explore a classification based on genes related to proliferation and growth. Methods The growth-related genes of GBM were identified by CRISPR-Cas9 and univariate Cox regression analysis. The expression of these genes in the Cancer Genome Atlas cohort (TCGA) was used to construct growth-related genes subtypes (GGSs) via consensus clustering. Validation of this subtyping was performed using the nearest template prediction (NTP) algorithm in two independent Gene Expression Omnibus (GEO) cohorts and the ZZ cohort. Additionally, copy number variations, biological functions, and potential drugs were analyzed for each of the different subtypes separately. Results Our research established multicenter-validated GGSs. GGS1 exhibits the poorest prognosis, with the highest frequency of chr 7 gain & chr 10 loss, and the lowest frequency of chr 19 & 20 co-gain. Additionally, GGS1 displays the highest expression of EGFR. Furthermore, it is significantly enriched in metabolic, stemness, proliferation, and signaling pathways. Besides we showed that Foretinib may be a potential therapeutic agent for GGS1, the worst prognostic subtype, through data screening and in vitro experiments. GGS2 has a moderate prognosis, with a slightly higher proportion of chr 7 gain & chr 10 loss, and the highest proportion of chr 19 & 20 co-gain. The prognosis of GGS3 is the best, with the least chr 7 gain & 10 loss and EGFR expression. Conclusions These results enhance our understanding of the heterogeneity of GBM and offer insights for stratified management and precise treatment of GBM patients. Supplementary Information The online version contains supplementary material available at 10.1186/s12885-023-11131-7.

Dataset Information

Genome-Wide CRISPR-Cas9 Screening Identifies NF-?B/E2F6 Responsible for EGFRvIII-Associated Temozolomide Resistance in Glioblastoma.

Publications

Genome-Wide CRISPR-Cas9 Screening Identifies NF-κB/E2F6 Responsible for EGFRvIII-Associated Temozolomide Resistance in Glioblastoma.

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