Khám phá sự phụ thuộc vào môi trường vi mô trong ung thư bạch cầu cấp tính ở chuột thông qua sàng lọc CRISPR toàn bộ bộ gen tại chỗ

Blood Advances - Tập 6 - Trang 5072-5084 - 2022
Francois E. Mercier1,2,3, Jiantao Shi4, David B. Sykes1,2,3, Toshihiko Oki1,2,3, Maja Jankovic5, Cheuk Him Man6, Youmna S. Kfoury1,2,3, Elizabeth Miller1,2,3, Shutao He7, Alexander Zhu1,2,3, Radovan Vasic1,2,3, John Doench8, Alexandre Orthwein9, Franziska Michor2,4,8,10, David T. Scadden1,2,3,10
1Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA
2Department of Stem Cell and Regenerative Biology, Harvard University, Boston, MA
3Harvard Stem Cell Institute, Harvard University, Cambridge, MA
4Center for Cancer Evolution and Department of Data Science, Dana-Farber Cancer Institute, Department of Biostatistics, Harvard T.H. Chan School of Public Health, Boston, MA
5Division of Experimental Medicine, Department of Medicine, McGill University, Montreal, QC, Canada
6Li Ka Shing Faculty of Medicine, University of Hong Kong, Hong Kong
7State Key Laboratory of Molecular Biology, Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, Beijing, China
8Broad Institute, Cambridge, MA
9Gerald Bronfman Department of Oncology, McGill University, Montreal, QC, Canada
10Ludwig Center at Harvard, Boston, MA

Tóm tắt

Tóm tắt Các nghiên cứu CRISPR trên toàn bộ bộ gen đã rất hữu ích trong việc xác định các mục tiêu điều trị trong nhiều loại ung thư bằng cách định nghĩa các gen thiết yếu cho sự phát triển ác tính. Tuy nhiên, hầu hết các nghiên cứu CRISPR được thực hiện trong vitro và do đó không thể xác định các gen thiết yếu cho sự tương tác với môi trường vi mô trong vivo. Ở đây, chúng tôi báo cáo các nghiên cứu CRISPR toàn bộ bộ gen trong 2 mô hình chuột in vivo của bệnh ung thư bạch cầu cấp tính (AML) do sự hợp nhất KMT2A/MLLT3 gây ra hoặc do sự đồng biểu hiện của Hoxa9 và Meis1. Sự xác thực thứ cấp bằng cách sử dụng một thư viện tập trung đã xác định 72 gen thiết yếu đặc biệt cho sự phát triển bạch cầu trong vivo, bao gồm các thành phần của phức hợp chính thuộc lớp I về tính tương thích mô, Cd47, thụ thể bổ sung Cr1l, và con đường β-4-galactosylation. Quan trọng là, một số phát hiện đặc hiệu cho in vivo này có ảnh hưởng dự đoán hoặc được suy luận là các yếu tố điều hòa chính của hoạt động protein trong các trường hợp AML ở người. Chẳng hạn, chúng tôi đã xác định Fermt3, một yếu tố điều hòa chính của tín hiệu integrin, có sự phụ thuộc đặc hiệu in vivo với sự liên quan cao đến dự đoán. Tổng thể, chúng tôi cho thấy một quy trình thí nghiệm và tính toán cho các sàng lọc chức năng toàn bộ bộ gen trong vivo trong AML và cung cấp một nguồn lực toàn bộ bộ gen về các yếu tố thiết yếu thúc đẩy sự phát triển bạch cầu trong vivo.

Từ khóa


Tài liệu tham khảo

Papaemmanuil, 2016, Genomic classification and prognosis in acute myeloid leukemia, N Engl J Med., 374, 2209, 10.1056/NEJMoa1516192 Tyner, 2018, Functional genomic landscape of acute myeloid leukaemia, Nature., 562, 526, 10.1038/s41586-018-0623-z Ley, 2013, Genomic and epigenomic landscapes of adult de novo acute myeloid leukemia, N Engl J Med., 368, 2059, 10.1056/NEJMoa1301689 Shih, 2015, Mutational cooperativity linked to combinatorial epigenetic gain of function in acute myeloid leukemia, Cancer Cell., 27, 502, 10.1016/j.ccell.2015.03.009 Perl, 2019, Gilteritinib or chemotherapy for relapsed or refractory FLT3-mutated AML, N Engl J Med., 381, 1728, 10.1056/NEJMoa1902688 Stone, 2017, Midostaurin plus chemotherapy for acute myeloid leukemia with a FLT3 mutation, N Engl J Med., 377, 454, 10.1056/NEJMoa1614359 Borella, 2021, Targeting the plasticity of mesenchymal stromal cells to reroute the course of acute myeloid leukemia, Blood., 138, 557 Barbier, 2020, Endothelial E-selectin inhibition improves acute myeloid leukaemia therapy by disrupting vascular niche-mediated chemoresistance, Nat Commun., 11, 2042, 10.1038/s41467-020-15817-5 Kokkaliaris, 2020, Cell interactions in the bone marrow microenvironment affecting myeloid malignancies, Blood Adv., 4, 3795, 10.1182/bloodadvances.2020002127 van Gastel, 2020, Induction of a timed metabolic collapse to overcome cancer chemoresistance, Cell Metab., 32, 391, 10.1016/j.cmet.2020.07.009 Forte, 2020, Bone marrow mesenchymal stem cells support acute myeloid leukemia bioenergetics and enhance antioxidant defense and escape from chemotherapy, Cell Metab., 32, 829, 10.1016/j.cmet.2020.09.001 Ye, 2016, Leukemic stem cells evade chemotherapy by metabolic adaptation to an adipose tissue niche, Cell Stem Cell., 19, 23, 10.1016/j.stem.2016.06.001 Majeti, 2009, CD47 is an adverse prognostic factor and therapeutic antibody target on human acute myeloid leukemia stem cells, Cell., 138, 286, 10.1016/j.cell.2009.05.045 Herbrich, 2021, Overexpression of CD200 is a stem cell-specific mechanism of immune evasion in AML, J Immunother Cancer., 9, 1, 10.1136/jitc-2021-002968 Xu, 2021, Regulatory T cells promote the stemness of leukemia stem cells through IL10 cytokine-related signaling pathway, Leukemia., 36, 403, 10.1038/s41375-021-01375-2 Ramdas, 2020, Driver mutations in leukemia promote disease pathogenesis through a combination of cell-autonomous and niche modulation, Stem Cell Reports., 15, 95, 10.1016/j.stemcr.2020.05.002 Duarte, 2018, Inhibition of endosteal vascular niche remodeling rescues hematopoietic stem cell loss in AML, Cell Stem Cell., 22, 64, 10.1016/j.stem.2017.11.006 Basheer, 2019, Genome-scale drop-out screens to identify cancer cell vulnerabilities in AML, Curr Opin Genet Dev., 54, 83, 10.1016/j.gde.2019.04.004 Barbieri, 2017, Promoter-bound METTL3 maintains myeloid leukaemia by m6A-dependent translation control, Nature., 552, 126, 10.1038/nature24678 Yamauchi, 2018, Genome-wide CRISPR-Cas9 screen identifies leukemia-specific dependence on a pre-mRNA metabolic pathway regulated by DCPS, Cancer Cell., 33, 386, 10.1016/j.ccell.2018.01.012 Cao, 2021, ZMYND8-regulated IRF8 transcription axis is an acute myeloid leukemia dependency, Mol Cell., 81, 3604, 10.1016/j.molcel.2021.07.018 Zhang, 2020, Functional interrogation of HOXA9 regulome in MLLr leukemia via reporter-based CRISPR/Cas9 screen, eLife., 9, 1, 10.7554/eLife.57858 Seneviratne, 2019, The mitochondrial transacylase, tafazzin, regulates for AML stemness by modulating intracellular levels of phospholipids, Cell Stem Cell., 24, 621, 10.1016/j.stem.2019.02.020 Khan, 2020, Mitochondrial carrier homolog 2 is necessary for AML survival, Blood., 136, 81, 10.1182/blood.2019000106 Chen, 2020, Vitamin B6 addiction in acute myeloid leukemia, Cancer Cell., 37, 71, 10.1016/j.ccell.2019.12.002 Dai, 2021, In vivo genome-wide CRISPR screen reveals breast cancer vulnerabilities and synergistic mTOR/Hippo targeted combination therapy, Nat Commun., 12, 3055, 10.1038/s41467-021-23316-4 Ramakrishnan, 2020, CXCR4 signaling has a CXCL12-independent essential role in murine MLL-AF9-driven acute myeloid leukemia, Cell Rep., 31, 107684, 10.1016/j.celrep.2020.107684 Lin, 2022, An in vivo CRISPR screening platform for prioritizing therapeutic targets in AML, Cancer Discov, 12, 432, 10.1158/2159-8290.CD-20-1851 Platt, 2014, CRISPR-Cas9 knockin mice for genome editing and cancer modeling, Cell., 159, 440, 10.1016/j.cell.2014.09.014 Krivtsov, 2006, Transformation from committed progenitor to leukaemia stem cell initiated by MLL-AF9, Nature., 442, 818, 10.1038/nature04980 Kroon, 1998, Hoxa9 transforms primary bone marrow cells through specific collaboration with Meis1a but not Pbx1b, EMBO J., 17, 3714, 10.1093/emboj/17.13.3714 Naviaux, 1996, The pCL vector system: rapid production of helper-free, high-titer, recombinant retroviruses, J Virol., 70, 5701, 10.1128/jvi.70.8.5701-5705.1996 Sykes, 2016, Inhibition of dihydroorotate dehydrogenase overcomes differentiation blockade in acute myeloid leukemia, Cell., 167, 171, 10.1016/j.cell.2016.08.057 Sanjana, 2014, Improved vectors and genome-wide libraries for CRISPR screening, Nat Methods., 11, 783, 10.1038/nmeth.3047 Griffith, 2013, DGIdb: mining the druggable genome, Nat Methods., 10, 1209, 10.1038/nmeth.2689 Tzelepis, 2016, A CRISPR dropout screen identifies genetic vulnerabilities and therapeutic targets in acute myeloid leukemia, Cell Rep., 17, 1193, 10.1016/j.celrep.2016.09.079 Li, 2014, MAGeCK enables robust identification of essential genes from genome-scale CRISPR/Cas9 knockout screens, Genome Biol., 15, 554, 10.1186/s13059-014-0554-4 Margolin, 2006, ARACNE: an algorithm for the reconstruction of gene regulatory networks in a mammalian cellular context, BMC Bioinformatics., 7, S7, 10.1186/1471-2105-7-S1-S7 Alvarez, 2016, Functional characterization of somatic mutations in cancer using network-based inference of protein activity, Nat Genet., 48, 838, 10.1038/ng.3593 Ng, 2016, A 17-gene stemness score for rapid determination of risk in acute leukaemia, Nature., 540, 433, 10.1038/nature20598 Bolouri, 2018, The molecular landscape of pediatric acute myeloid leukemia reveals recurrent structural alterations and age-specific mutational interactions, Nat Med., 24, 103, 10.1038/nm.4439 Muntean, 2012, The pathogenesis of mixed-lineage leukemia, Annu Rev Pathol., 7, 283, 10.1146/annurev-pathol-011811-132434 Golub, 1999, Molecular classification of cancer: class discovery and class prediction by gene expression monitoring, 286, 531 Collins, 2016, Role of HOXA9 in leukemia: dysregulation, cofactors and essential targets, Oncogene., 35, 1090, 10.1038/onc.2015.174 Miller, 2013, In vivo RNAi screening identifies a leukemia-specific dependence on integrin beta 3 signaling, Cancer Cell., 24, 45, 10.1016/j.ccr.2013.05.004 Wang, 2017, Gene essentiality profiling reveals gene networks and synthetic lethal interactions with oncogenic Ras, Cell., 168, 890, 10.1016/j.cell.2017.01.013 Bernt, 2011, MLL-rearranged leukemia is dependent on aberrant H3K79 methylation by DOT1L, Cancer Cell., 20, 66, 10.1016/j.ccr.2011.06.010 Chen, 2006, The tumor suppressor menin regulates hematopoiesis and myeloid transformation by influencing Hox gene expression, Proc Natl Acad Sci USA., 103, 1018, 10.1073/pnas.0510347103 Pillinger, 2016, Targeting PI3Kδ and PI3Kγ signalling disrupts human AML survival and bone marrow stromal cell mediated protection, Oncotarget., 7, 39784, 10.18632/oncotarget.9289 Miwa, 2009, Complement-dependent T-cell lymphopenia caused by thymocyte deletion of the membrane complement regulator Crry, Blood., 113, 2684, 10.1182/blood-2008-05-157966 Giannini, 2020, β4GALT1 controls β1 integrin function to govern thrombopoiesis and hematopoietic stem cell homeostasis, Nat Commun., 11, 356, 10.1038/s41467-019-14178-y Jan, 2012, Clonal evolution of preleukemic hematopoietic stem cells precedes human acute myeloid leukemia, Sci Transl Med., 4, 149ra118, 10.1126/scitranslmed.3004315 Corces-Zimmerman, 2014, Preleukemic mutations in human acute myeloid leukemia affect epigenetic regulators and persist in remission, Proc Natl Acad Sci USA., 111, 2548, 10.1073/pnas.1324297111 Bowman, 2018, Clonal hematopoiesis and evolution to hematopoietic malignancies, Cell Stem Cell., 22, 157, 10.1016/j.stem.2018.01.011 Eriksson, 2015, Epigenetic aberrations in acute myeloid leukemia: early key events during leukemogenesis, Exp Hematol., 43, 609, 10.1016/j.exphem.2015.05.009 Krönke, 2013, Clonal evolution in relapsed NPM1-mutated acute myeloid leukemia, Blood., 122, 100, 10.1182/blood-2013-01-479188 Miwa, 2002, Crry, but not CD59 and DAF, is indispensable for murine erythrocyte protection in vivo from spontaneous complement attack, Blood., 99, 3707, 10.1182/blood.V99.10.3707 Laverdière, 2016, Complement cascade gene expression defines novel prognostic subgroups of acute myeloid leukemia, Exp Hematol., 44, 1039, 10.1016/j.exphem.2016.07.012 Sugahara, 2012, Large-scale identification of target proteins of a glycosyltransferase isozyme by lectin-IGOT-LC/MS, an LC/MS-based glycoproteomic approach, Sci Rep., 2, 680, 10.1038/srep00680 Asano, 2003, Impaired selectin-ligand biosynthesis and reduced inflammatory responses in β-1,4-galactosyltransferase-I- deficient mice, Blood., 102, 1678, 10.1182/blood-2003-03-0836 Pang, 2018, Multiple roles of glycans in hematological malignancies, Front Oncol., 8, 364, 10.3389/fonc.2018.00364 Wilkinson, 2013, RUNX1 is a key target in t(4;11) leukemias that contributes to gene activation through an AF4-MLL complex interaction, Cell Rep., 3, 116, 10.1016/j.celrep.2012.12.016 Docking, 2021, A clinical transcriptome approach to patient stratification and therapy selection in acute myeloid leukemia, Nat Commun., 12, 2474, 10.1038/s41467-021-22625-y Bromberger, 2018, Direct Rap1/Talin1 interaction regulates platelet and neutrophil integrin activity in mice, Blood., 132, 2754, 10.1182/blood-2018-04-846766 Kuijpers, 2009, LAD-1/variant syndrome is caused by mutations in FERMT3, Blood., 113, 4740, 10.1182/blood-2008-10-182154