Genomic Copy Number Dictates a Gene-Independent Cell Response to CRISPR/Cas9 Targeting.
The CRISPR/Cas9 system enables genome editing and somatic cell genetic screens in mammalian cells. We performed genome-scale loss-of-function screens in 33 cancer cell lines to identify genes essential for proliferation/survival and found a strong correlation between increased gene copy number and decreased cell viability after genome editing. Within regions of copy-number gain, CRISPR/Cas9 targeting of both expressed and unexpressed genes, as well as intergenic loci, led to significantly decreased cell proliferation through induction of a G2 cell-cycle arrest. By examining single-guide RNAs that map to multiple genomic sites, we found that this cell response to CRISPR/Cas9 editing correlated strongly with the number of target loci. These observations indicate that genome targeting by CRISPR/Cas9 elicits a gene-independent antiproliferative cell response. This effect has important practical implications for the interpretation of CRISPR/Cas9 screening data and confounds the use of this technology for the identification of essential genes in amplified regions.



We found that the number of CRISPR/Cas9-induced DNA breaks dictates a gene-independent antiproliferative response in cells. These observations have practical implications for using CRISPR/Cas9 to interrogate cancer gene function and illustrate that cancer cells are highly sensitive to site-specific DNA damage, which may provide a path to novel therapeutic strategies. Cancer Discov; 6(8); 914-29. ©2016 AACR.See related commentary by Sheel and Xue, p. 824See related article by Munoz et al., p. 900This article is highlighted in the In This Issue feature, p. 803.
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts. Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts. Harvard Medical School, Boston, Massachusetts.
Andrew J
AJ
Aguirre
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Robin M
RM
Meyers
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Barbara A
BA
Weir
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Francisca
F
Vazquez
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Cheng-Zhong
CZ
Zhang
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Uri
U
Ben-David
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
April
A
Cook
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Gavin
G
Ha
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
William F
WF
Harrington
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Mihir B
MB
Doshi
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Maria
M
Kost-Alimova
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Stanley
S
Gill
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Han
H
Xu
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Levi D
LD
Ali
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Guozhi
G
Jiang
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Sasha
S
Pantel
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Yenarae
Y
Lee
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Amy
A
Goodale
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Andrew D
AD
Cherniack
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Coyin
C
Oh
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Gregory
G
Kryukov
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
Glenn S
GS
Cowley
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts. Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts. Harvard Medical School, Boston, Massachusetts. Howard Hughes Medical Institute, Chevy Chase, Maryland.
Levi A
LA
Garraway
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts. Harvard Medical School, Boston, Massachusetts. Boston Children's Hospital, Boston, Massachusetts.
Kimberly
K
Stegmaier
Broad Institute of Harvard and MIT, Cambridge, Massachusetts. St. Jude Children's Research Hospital, Memphis, Tennessee.
Charles W
CW
Roberts
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts. Harvard Medical School, Boston, Massachusetts. Howard Hughes Medical Institute, Chevy Chase, Maryland.
Todd R
TR
Golub
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts. Harvard Medical School, Boston, Massachusetts. Department of Pathology, Harvard Medical School, Boston, Massachusetts. Center for Cancer Genome Discovery, Dana-Farber Cancer Institute, Boston, Massachusetts.
Matthew
M
Meyerson
Broad Institute of Harvard and MIT, Cambridge, Massachusetts.
David E
DE
Root
Broad Institute of Harvard and MIT, Cambridge, Massachusetts. william_hahn@dfci.harvard.edu aviad@broadinstitute.org.
Aviad
A
Tsherniak
Dana-Farber Cancer Institute, Boston, Massachusetts. Broad Institute of Harvard and MIT, Cambridge, Massachusetts. Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts. Harvard Medical School, Boston, Massachusetts. Center for Cancer Genome Discovery, Dana-Farber Cancer Institute, Boston, Massachusetts. william_hahn@dfci.harvard.edu aviad@broadinstitute.org.
William C
WC
Hahn
D064113:CRISPR-Cas Systems; D045744:Cell Line, Tumor; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D053837:DNA Cleavage; D056915:DNA Copy Number Variations; D004249:DNA Damage; D059565:G2 Phase Cell Cycle Checkpoints; D005784:Gene Amplification; D018628:Gene Dosage; D000072669:Gene Editing; D015870:Gene Expression; D055786:Gene Knockout Techniques; D018390:Gene Targeting / Q000379:methods; D020043:Genes, Essential; D023281:Genomics / Q000379:methods; D057166:High-Throughput Screening Assays; D006801:Humans; D000094704:RNA, Guide, CRISPR-Cas Systems
D000094704:RNA, Guide, CRISPR-Cas Systems