An Integrated Genome-wide CRISPRa Approach to Functionalize lncRNAs in Drug Resistance

Assaf C Bester; Jonathan D Lee; Alejandro Chavez; Yu-Ru Lee; Daphna Nachmani; Suhani Vora; Joshua Victor; Martin Sauvageau; Emanuele Monteleone; John L Rinn; Paolo Provero; George M Church; John G Clohessy; Pier Paolo Pandolfi

doi:10.1016/j.cell.2018.03.052

An Integrated Genome-wide CRISPRa Approach to Functionalize lncRNAs in Drug Resistance

Cell. 2018 Apr 19;173(3):649-664.e20. doi: 10.1016/j.cell.2018.03.052.

Authors

Affiliations

¹ Cancer Research Institute, Beth Israel Deaconess Cancer Center, Department of Medicine and Pathology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA; Ludwig Center at Harvard, Harvard Medical School, Boston, MA, USA.
² Wyss Institute for Biologically Inspired Engineering, Harvard University, Cambridge, MA, USA; Department of Pathology and Cell Biology, Columbia University College of Physicians and Surgeons, New York, NY 10032, USA; Taub Institute for Research on Alzheimer's Disease and the Aging Brain, Columbia University College of Physicians and Surgeons, New York, NY 10032, USA.
³ Wyss Institute for Biologically Inspired Engineering, Harvard University, Cambridge, MA, USA.
⁴ Department of Stem Cell and Regenerative Biology, Harvard University, The Broad Institute of MIT and Harvard, Cambridge, MA, USA; Functional Genomics and Noncoding RNAs Research Unit, Institut de Recherches Cliniques de Montréal (IRCM), Montréal, QC, Canada; Department of Biochemistry and Molecular Medicine, Université de Montréal, Montréal, QC, Canada.
⁵ Department of Molecular Biotechnology and Health Sciences, and GenoBiToUS, Genomics and Bioinformatics Service, University of Turin, Turin, Italy.
⁶ Department of Stem Cell and Regenerative Biology, Harvard University, The Broad Institute of MIT and Harvard, Cambridge, MA, USA.
⁷ Department of Molecular Biotechnology and Health Sciences, and GenoBiToUS, Genomics and Bioinformatics Service, University of Turin, Turin, Italy; Center for Translational Genomics and Bioinformatics, San Raffaele Scientific Institute IRCCS, Milan, Italy.
⁸ Cancer Research Institute, Beth Israel Deaconess Cancer Center, Department of Medicine and Pathology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA; Ludwig Center at Harvard, Harvard Medical School, Boston, MA, USA; Preclinical Murine Pharmacogenetics Facility and Mouse Hospital, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA.
⁹ Cancer Research Institute, Beth Israel Deaconess Cancer Center, Department of Medicine and Pathology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA; Ludwig Center at Harvard, Harvard Medical School, Boston, MA, USA. Electronic address: ppandolf@bidmc.harvard.edu.

Abstract

Resistance to chemotherapy plays a significant role in cancer mortality. To identify genetic units affecting sensitivity to cytarabine, the mainstay of treatment for acute myeloid leukemia (AML), we developed a comprehensive and integrated genome-wide platform based on a dual protein-coding and non-coding integrated CRISPRa screening (DICaS). Putative resistance genes were initially identified using pharmacogenetic data from 760 human pan-cancer cell lines. Subsequently, genome scale functional characterization of both coding and long non-coding RNA (lncRNA) genes by CRISPR activation was performed. For lncRNA functional assessment, we developed a CRISPR activation of lncRNA (CaLR) strategy, targeting 14,701 lncRNA genes. Computational and functional analysis identified novel cell-cycle, survival/apoptosis, and cancer signaling genes. Furthermore, transcriptional activation of the GAS6-AS2 lncRNA, identified in our analysis, leads to hyperactivation of the GAS6/TAM pathway, a resistance mechanism in multiple cancers including AML. Thus, DICaS represents a novel and powerful approach to identify integrated coding and non-coding pathways of therapeutic relevance.

Keywords: AML; AXL/GAS6; CRISPR; CRISPRa; TEM; cancer; cytarabine; drug-resistance; leukemia; lncRNA.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Animals
CRISPR-Cas Systems*
Cytarabine / pharmacology
Drug Resistance, Neoplasm*
Female
Gene Expression Profiling
Gene Regulatory Networks
Genome, Human*
HEK293 Cells
HL-60 Cells
Humans
K562 Cells
Leukemia, Myeloid, Acute / drug therapy
Leukemia, Myeloid, Acute / genetics
Male
Mice
Pharmacogenetics
Proteins / genetics
RNA / analysis
RNA, Long Noncoding / genetics*
RNA, Messenger / genetics
Signal Transduction

Substances

Proteins
RNA, Long Noncoding
RNA, Messenger
Cytarabine
RNA

Abstract

Publication types

MeSH terms

Substances

Grants and funding