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      GATA2 Inhibition Sensitizes Acute Myeloid Leukemia Cells to Chemotherapy

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          Abstract

          Drug resistance constitutes one of the main obstacles for clinical recovery of acute myeloid leukemia (AML) patients. Therefore, the treatment of AML requires new strategies, such as adding a third drug. To address whether GATA2 could act as a regulator of chemotherapy resistance in human leukemia cells, we observed KG1a cells and clinical patients’ AML cells with a classic drug (Cerubidine) and Gefitinib. After utilizing chemotherapy, the expression of GATA2 and its target genes (EVI, SCL and WT1) in surviving AML cells and KG1a cells were significantly enhanced to double and quadrupled compared to its original level respectively. Furthermore, with continuous chemotherapeutics, AML cells with GATA2 knockdown or treated with GATA2 inhibitor (K1747) almost eliminated with dramatically reduced expression of WT1, SCL, EVI, and significantly increased apoptotic population. Therefore, we propose that reducing GATA2 expression or inhibition of its transcription activity can relieve the drug resistance of acute myeloid leukemia cells and it would be helpful for eliminating the leukemia cells in patients.

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          Most cited references30

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          An update of current treatments for adult acute myeloid leukemia.

          Recent advances in acute myeloid leukemia (AML) biology and its genetic landscape should ultimately lead to more subset-specific AML therapies, ideally tailored to each patient's disease. Although a growing number of distinct AML subsets have been increasingly characterized, patient management has remained disappointingly uniform. If one excludes acute promyelocytic leukemia, current AML management still relies largely on intensive chemotherapy and allogeneic hematopoietic stem cell transplantation (HSCT), at least in younger patients who can tolerate such intensive treatments. Nevertheless, progress has been made, notably in terms of standard drug dose intensification and safer allogeneic HSCT procedures, allowing a larger proportion of patients to achieve durable remission. In addition, improved identification of patients at relatively low risk of relapse should limit their undue exposure to the risks of HSCT in first remission. The role of new effective agents, such as purine analogs or gemtuzumab ozogamicin, is still under investigation, whereas promising new targeted agents are under clinical development. In contrast, minimal advances have been made for patients unable to tolerate intensive treatment, mostly representing older patients. The availability of hypomethylating agents likely represents an encouraging first step for this latter population, and it is hoped will allow for more efficient combinations with novel agents.
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            Optimizing sgRNA structure to improve CRISPR-Cas9 knockout efficiency

            Background Single-guide RNA (sgRNA) is one of the two key components of the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 genome-editing system. The current commonly used sgRNA structure has a shortened duplex compared with the native bacterial CRISPR RNA (crRNA)–transactivating crRNA (tracrRNA) duplex and contains a continuous sequence of thymines, which is the pause signal for RNA polymerase III and thus could potentially reduce transcription efficiency. Results Here, we systematically investigate the effect of these two elements on knockout efficiency and showed that modifying the sgRNA structure by extending the duplex length and mutating the fourth thymine of the continuous sequence of thymines to cytosine or guanine significantly, and sometimes dramatically, improves knockout efficiency in cells. In addition, the optimized sgRNA structure also significantly increases the efficiency of more challenging genome-editing procedures, such as gene deletion, which is important for inducing a loss of function in non-coding genes. Conclusions By a systematic investigation of sgRNA structure we find that extending the duplex by approximately 5 bp combined with mutating the continuous sequence of thymines at position 4 to cytosine or guanine significantly increases gene knockout efficiency in CRISPR-Cas9-based genome editing experiments. Electronic supplementary material The online version of this article (doi:10.1186/s13059-015-0846-3) contains supplementary material, which is available to authorized users.
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              The GATA2 transcriptional network is requisite for RAS oncogene-driven non-small cell lung cancer.

              Non-small cell lung cancer (NSCLC) is the most frequent cause of cancer deaths worldwide; nearly half contain mutations in the receptor tyrosine kinase/RAS pathway. Here we show that RAS-pathway mutant NSCLC cells depend on the transcription factor GATA2. Loss of GATA2 reduced the viability of NSCLC cells with RAS-pathway mutations, whereas wild-type cells were unaffected. Integrated gene expression and genome occupancy analyses revealed GATA2 regulation of the proteasome, and IL-1-signaling, and Rho-signaling pathways. These pathways were functionally significant, as reactivation rescued viability after GATA2 depletion. In a Kras-driven NSCLC mouse model, Gata2 loss dramatically reduced tumor development. Furthermore, Gata2 deletion in established Kras mutant tumors induced striking regression. Although GATA2 itself is likely undruggable, combined suppression of GATA2-regulated pathways with clinically approved inhibitors caused marked tumor clearance. Discovery of the nononcogene addiction of KRAS mutant lung cancers to GATA2 presents a network of druggable pathways for therapeutic exploitation. Copyright © 2012 Elsevier Inc. All rights reserved.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                23 January 2017
                2017
                : 12
                : 1
                : e0170630
                Affiliations
                [1 ]Department of Hematology, Tongren Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
                [2 ]Department of Laboratory Medicine, Tongren Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China
                University of Navarra, SPAIN
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                • Conceptualization: WZ HS.

                • Data curation: LY HS.

                • Formal analysis: LY HS.

                • Funding acquisition: LY HS.

                • Investigation: LY HS.

                • Methodology: LY HS.

                • Project administration: LY HS.

                • Resources: LY HS.

                • Software: LY HS.

                • Supervision: YC BX YF.

                • Validation: LY HS.

                • Visualization: LY HS.

                • Writing – original draft: LY HS.

                • Writing – review & editing: LY HS.

                Article
                PONE-D-16-40980
                10.1371/journal.pone.0170630
                5256934
                28114350
                e85e5841-f5b0-4179-b1e6-ef7e04a69857
                © 2017 Yang et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 14 October 2016
                : 6 January 2017
                Page count
                Figures: 5, Tables: 1, Pages: 12
                Funding
                Funded by: Tongren Hospital
                Award ID: TR201407
                Award Recipient :
                This work was supported by the Shanghai Municipal Health Bureau Project (15ZR1437900), Natural Science Research Project of Changning District (CNKW2014Z01), Leading Disciplines of Medical Laboratory in Changning District (2012QZK01) and Tongren Hospital grant (grant TR201407, TRYJ201506). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Medicine and Health Sciences
                Oncology
                Cancers and Neoplasms
                Hematologic Cancers and Related Disorders
                Leukemias
                Myeloid Leukemia
                Acute Myeloid Leukemia
                Medicine and Health Sciences
                Hematology
                Hematologic Cancers and Related Disorders
                Leukemias
                Myeloid Leukemia
                Acute Myeloid Leukemia
                Medicine and Health Sciences
                Pharmaceutics
                Drug Therapy
                Chemotherapy
                Medicine and Health Sciences
                Pharmaceutics
                Drug Therapy
                Biology and Life Sciences
                Cell Biology
                Cell Processes
                Cell Death
                Apoptosis
                Medicine and Health Sciences
                Oncology
                Cancer Treatment
                Cancer Chemotherapy
                Medicine and Health Sciences
                Pharmaceutics
                Drug Therapy
                Chemotherapy
                Cancer Chemotherapy
                Medicine and Health Sciences
                Clinical Medicine
                Clinical Oncology
                Cancer Chemotherapy
                Medicine and Health Sciences
                Oncology
                Clinical Oncology
                Cancer Chemotherapy
                Medicine and Health Sciences
                Oncology
                Cancer Treatment
                Biology and Life Sciences
                Molecular Biology
                Molecular Biology Techniques
                Artificial Gene Amplification and Extension
                Polymerase Chain Reaction
                Research and Analysis Methods
                Molecular Biology Techniques
                Artificial Gene Amplification and Extension
                Polymerase Chain Reaction
                Research and Analysis Methods
                Biological Cultures
                Cell Cultures
                Cultured Tumor Cells
                Leukemia Cells
                Custom metadata
                All relevant data are within the paper and its Supporting Information files.

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                Uncategorized

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