Nature | 2021.01.26

Cancer research needs a better map

Jesse S. Boehm, Mathew J. Garnett, David J. Adams, Hayley E. Francies, Todd R. Golub, William C. Hahn, Francesco Iorio, James M. McFarland, Leopold Parts & Francisca Vazquez

Almost 15 years ago, scientists and clinicians set out to characterize genomes of tumours from thousands of patients. The result? The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC). Nearly every targeted cancer drug approved over the past decade has drawn from the data sets generated by these efforts. This information is now also providing clues to triangulate which individuals can benefit from new types of drug, such as pembrolizumab and nivolumab, which help the immune system to fight cancer. TCGA generated more than 2.5 petabytes of data measuring mutations, gene expression and protein levels across 33 cancer types. It catalysed innovation in DNA sequencing technology and genome analysis. It ultimately collected data from some 11,000 patients — data that thousands of researchers use. This work redefined cancers on the molecular level, and painted a picture of the mutations that occur in common tumour types.

bioRxiv | 2021.01.06

Integrated cross-study datasets of genetic dependencies in cancer

Clare Pacini, Joshua M. Dempster, Isabella Boyle, Emanuel Gonçalves, Hanna Najgebauer,... James M. McFarland, Mathew J. Garnett, Aviad Tsherniak & Francesco Iorio

CRISPR-Cas9 viability screens are increasingly performed at a genome-wide scale across large panels of cell lines to identify new therapeutic targets for precision cancer therapy. Integrating the datasets resulting from these studies is necessary to adequately represent the heterogeneity of human cancers and to assemble a comprehensive map of cancer genetic vulnerabilities. Here, we integrated the two largest public independent CRISPR-Cas9 screens performed to date (at the Broad and Sanger institutes) by assessing, comparing, and selecting methods for correcting biases due to heterogeneous single guide RNA efficiency, gene-independent responses to CRISPR-Cas9 targeting originated from copy number alterations, and experimental batch effects. Our integrated datasets recapitulate findings from the individual datasets, provide greater statistical power to cancer- and subtype-specific analyses, unveil additional biomarkers of gene dependency, and improve the detection of common essential genes. We provide the largest integrated resources of CRISPR-Cas9 screens to date and the basis for harmonizing existing and future functional genetics datasets.

Nature Communications | 2021.01.04

Global computational alignment of tumor and cell line transcriptional profiles

Allison Warren, Yejia Chen, Andrew Jones, Tsukasa Shibue, William C. Hahn, Jesse S. Boehm, Francisca Vazquez, Aviad Tsherniak & James M. McFarland

Cell lines are key tools for preclinical cancer research, but it remains unclear how well they represent patient tumor samples. Direct comparisons of tumor and cell line transcriptional profiles are complicated by several factors, including the variable presence of normal cells in tumor samples. We thus develop an unsupervised alignment method (Celligner) and apply it to integrate several large-scale cell line and tumor RNA-Seq datasets. Although our method aligns the majority of cell lines with tumor samples of the same cancer type, it also reveals large differences in tumor similarity across cell lines. Using this approach, we identify several hundred cell lines from diverse lineages that present a more mesenchymal and undifferentiated transcriptional state and that exhibit distinct chemical and genetic dependencies. Celligner could be used to guide the selection of cell lines that more closely resemble patient tumors and improve the clinical translation of insights gained from cell lines.

Cell Reports | 2020.12.15

Synthetic Lethal Interaction between the ESCRT Paralog Enzymes VPS4A and VPS4B in Cancers Harboring Loss of Chromosome 18q or 16q

Jasper E. Neggers, Brenton R. Paolella, Adhana Asfaw, Michael V. Rothberg... William C. Hahn, Kimberly Stegmaier, Todd R. Golub, Francisca Vazquez & Andrew J. Aguirre

Few therapies target the loss of tumor suppressor genes in cancer. We examine CRISPR-SpCas9 and RNA-interference loss-of-function screens to identify new therapeutic targets associated with genomic loss of tumor suppressor genes. The endosomal sorting complexes required for transport (ESCRT) ATPases VPS4A and VPS4B score as strong synthetic lethal dependencies. VPS4A is essential in cancers harboring loss of VPS4B adjacent to SMAD4 on chromosome 18q and VPS4B is required in tumors with co-deletion of VPS4A and CDH1 (E-cadherin) on chromosome 16q. We demonstrate that more than 30% of cancers selectively require VPS4A or VPS4B. VPS4A suppression in VPS4B-deficient cells selectively leads to ESCRT-III filament accumulation, cytokinesis defects, nuclear deformation, G2/M arrest, apoptosis, and potent tumor regression. CRISPR-SpCas9 screening and integrative genomic analysis reveal other ESCRT members, regulators of abscission, and interferon signaling as modifiers of VPS4A dependency. We describe a compendium of synthetic lethal vulnerabilities and nominate VPS4A and VPS4B as high-priority therapeutic targets for cancers with 18q or 16q loss.

bioRxiv | 2020.10.17

Phosphate dysregulation via the XPR1:KIDINS220 protein complex is a therapeutic vulnerability in ovarian cancer

Daniel P Bondeson, Brenton R Paolella, Adhana Asfaw, Michael Rothberg... Michael Mannstadt, James M McFarland, Francisca Vazquez & Todd R Golub

Clinical outcomes for patients with ovarian and uterine cancers have not improved greatly in the past twenty years. To identify ovarian and uterine cancer vulnerabilities, we analyzed genome-scale CRISPR/ Cas9 loss-of-function screens across 739 human cancer cell lines. We found that many ovarian cancer cell lines overexpress the phosphate importer SLC34A2, which renders them sensitive to loss of the phosphate exporter XPR1. We extensively validated the XPR1 vulnerability in cancer cell lines and found that the XPR1 dependency was retained in vivo. Overexpression of SLC34A2 is frequently observed in tumor samples and is regulated by PAX8 - a transcription factor required for ovarian cancer survival. XPR1 overexpression and copy number amplifications are also frequently observed. Mechanistically, SLC34A2 overexpression and impaired phosphate efflux leads to the accumulation of intracellular phosphate and cell death. We further show that proper localization and phosphate efflux by XPR1 requires a novel binding partner, KIDINS220. Loss of either XPR1 or KIDINS220 results in acidic vacuolar structures which precede cell death. These data point to the XPR1:KIDINS220 complex - and phosphate dysregulation more broadly - as a therapeutic vulnerability in ovarian cancer.

Nature Communications | 2020.08.27

Multiplexed single-cell transcriptional response profiling to define cancer vulnerabilities and therapeutic mechanism of action

James M. McFarland, Brenton R. Paolella, Allison Warren, Kathryn Geiger-Schuller,... Aviv Regev, Andrew J. Aguirre, Francisca Vazquez, & Aviad Tsherniak

Assays to study cancer cell responses to pharmacologic or genetic perturbations are typically restricted to using simple phenotypic readouts such as proliferation rate. Information-rich assays, such as gene-expression profiling, have generally not permitted efficient profiling of a given perturbation across multiple cellular contexts. Here, we develop MIX-Seq, a method for multiplexed transcriptional profiling of post-perturbation responses across a mixture of samples with single-cell resolution, using SNP-based computational demultiplexing of single-cell RNA-sequencing data. We show that MIX-Seq can be used to profile responses to chemical or genetic perturbations across pools of 100 or more cancer cell lines. We combine it with Cell Hashing to further multiplex additional experimental conditions, such as post-treatment time points or drug doses. Analyzing the high-content readout of scRNA-seq reveals both shared and context-specific transcriptional response components that can identify drug mechanism of action and enable prediction of long-term cell viability from short-term transcriptional responses to treatment.

Molecular Systems Biology | 2020.07.21

The Cancer Dependency Map enables drug mechanism‐of‐action investigations

Francisca Vazquez & Jesse Boehm

How do small molecules exert their effects in mammalian cells? This seemingly simple question continues to represent one of the fundamental challenges of modern translational science and as such has long been the subject of intense scientific scrutiny. In their recent study, Garnett and colleagues (Gonçalves et al, 2020) demonstrate proof‐of‐concept for a new way to attack this problem systematically for Oncology drugs, by identifying correlated CRISPR‐ and drug‐killing profiles in the Cancer Dependency Map dataset.

Nature Cancer | 2020.01.20

Discovering the anticancer potential of non-oncology drugs by systematic viability profiling

Steven M. Corsello, Rohith T. Nagari, Ryan D. Spangler, Jordan Rossen, Mustafa Kocak,... Jesse S. Boehm, Christopher C. Mader, Aviad Tsherniak & Todd R. Golub

Anticancer uses of non-oncology drugs have occasionally been found, but such discoveries have been serendipitous. We sought to create a public resource containing the growth-inhibitory activity of 4,518 drugs tested across 578 human cancer cell lines. We used PRISM (profiling relative inhibition simultaneously in mixtures), a molecular barcoding method, to screen drugs against cell lines in pools. An unexpectedly large number of non-oncology drugs selectively inhibited subsets of cancer cell lines in a manner predictable from...

Nature Communications | 2019.12.20

Agreement between two large pan-cancer CRISPR-Cas9 gene dependency data sets

Joshua M. Dempster, Clare Pacini, Sasha Pantel, Fiona M. Behan,... David E. Root, Mathew J. Garnett, Aviad Tsherniak & Francesco Iorio

Genome-scale CRISPR-Cas9 viability screens performed in cancer cell lines provide a systematic approach to identify cancer dependencies and new therapeutic targets. As multiple large-scale screens become available, a formal assessment of the reproducibility of these experiments becomes necessary. We analyze data from recently published pan-cancer CRISPR-Cas9 screens performed at the Broad and Sanger Institutes. Despite significant differences in experimental protocols and reagents, we find that the screen results are highly...

Cell | 2019.10.17

Optical Pooled Screens in Human Cells

David Feldman, Avtar Singh, Jonathan L. Schmid-Burgk, Rebecca J Carlson,... Anja Mezger, Anthony J. Garrity, Feng Zhang & Paul C. Blainey

Genetic screens are critical for the systematic identification of genes underlying cellular phenotypes. Pooling gene perturbations greatly improves scalability but is not compatible with imaging of complex and dynamic cellular phenotypes. Here, we introduce a pooled approach for optical genetic screens in mammalian cells. We use targeted in situ sequencing to demultiplex a library of genetic perturbations following image-based phenotyping. We screened a set of 952 genes across millions of cells for involvement in nuclear factor κB (NF-κB)...

Cell Reports | 2019.08.27

Small-Molecule and CRISPR Screening Converge to Reveal Receptor Tyrosine Kinase Dependencies in Pediatric Rhabdoid Tumors

Elaine M. Oberlick, Matthew G. Rees, Brinton Seashore-Ludlow, Francisca Vazquez,... William C. Hahn, Elizabeth A. Stewart, Stuart L. Schreiber & Charles W.M. Roberts

Cancer is often seen as a disease of mutations and chromosomal abnormalities. However, some cancers, including pediatric rhabdoid tumors (RTs), lack recurrent alterations targetable by current drugs and need alternative, informed therapeutic options. To nominate potential targets, we performed a high-throughput small-molecule screen complemented by a genome-scale CRISPR-Cas9 gene-knockout screen in a large number of RT and control cell lines. These approaches converged to reveal several receptor...

Nature | 2019.05.08

Next-generation characterization of the Cancer Cell Line Encyclopedia

Mahmoud Ghandi, Franklin W. Huang, Judit Jané-Valbuena, Gregory V. Kryukov,... Todd R. Golub, Levi A. Garraway & William R. Sellers

Large panels of comprehensively characterized human cancer models, including the Cancer Cell Line Encyclopedia (CCLE), have provided a rigorous framework with which to study genetic variants, candidate targets, and small-molecule and biological therapeutics and to identify new marker-driven cancer dependencies. To improve our understanding of the molecular features that contribute to cancer phenotypes, including drug responses, here we have expanded the characterizations of cancer cell lines to include genetic,...

Nature Medicine | 2019.05.08

The landscape of cancer cell line metabolism

Haoxin Li, Shaoyang Ning, Mahmoud Ghandi, Gregory V. Kryukov,... Stuart L. Schreiber, Clary B. Clish, Levi A. Garraway & William R. Sellers

Despite considerable efforts to identify cancer metabolic alterations that might unveil druggable vulnerabilities, systematic characterizations of metabolism as it relates to functional genomic features and associated dependencies remain uncommon. To further understand the metabolic diversity of cancer, we profiled 225 metabolites in 928 cell lines from more than 20 cancer types in the Cancer Cell Line Encyclopedia (CCLE) using liquid chromatography–mass spectrometry (LC-MS). This resource enables unbiased association analysis...

Nature | 2019.04.10

WRN helicase is a synthetic lethal target in microsatellite unstable cancers

Edmond M. Chan, Tsukasa Shibue, James M. McFarland, Benjamin Gaeta,... Todd R. Golub, Aviad Tsherniak, Francisca Vazquez & Adam J. Bass

Synthetic lethality—an interaction between two genetic events through which the co-occurrence of these two genetic events leads to cell death, but each event alone does not—can be exploited for cancer therapeutics. DNA repair processes represent attractive synthetic lethal targets, because many cancers exhibit an impairment of a DNA repair pathway, which can lead to dependence on specific repair proteins. The success of poly(ADP-ribose) polymerase 1 (PARP-1) inhibitors in cancers with deficiencies in homologous recombination highlights the potential of this approach. Hypothesizing that other DNA repair defects,...

Nature | 2018.12.17

Loss of ADAR1 in tumours overcomes resistance to immune checkpoint blockade

Jeffrey J. Ishizuka, Robert T. Manguso, Collins K. Cheruiyot, Kevin Bi,... John G. Doench, David Kozono, Erez Y. Levanon & W. Nicholas Haining

Most patients with cancer either do not respond to immune checkpoint blockade or develop resistance to it, often because of acquired mutations that impair antigen presentation. Here we show that loss of function of the RNA-editing enzyme ADAR1 in tumour cells profoundly sensitizes tumours to immunotherapy and overcomes resistance to checkpoint blockade. In the absence of ADAR1, A-to-I editing of interferon-inducible RNA species is reduced, leading to double-stranded RNA ligand sensing by PKR and MDA5; this results in growth inhibition...

Nature Communications | 2018.11.02

Improved estimation of cancer dependencies from large-scale RNAi screens using model-based normalization and data integration

James M. McFarland, Zandra V. Ho, Guillaume Kugener, Joshua M. Dempster,... Todd R. Golub, William C. Hahn, David E. Root & Aviad Tsherniak

The availability of multiple datasets comprising genome-scale RNAi viability screens in hundreds of diverse cancer cell lines presents new opportunities for understanding cancer vulnerabilities. Integrated analyses of these data to assess differential dependency across genes and cell lines are challenging due to confounding factors such as batch effects and variable screen quality, as well as difficulty assessing gene dependency on an absolute scale. To address these issues, we incorporated cell line screen-quality parameters and hierarchical Bayesian inference into DEMETER2, an analytical framework...

Nature Genetics | 2017.10.30

Computational correction of copy number effect improves specificity of CRISPR-Cas9 essentiality screens in cancer cells

Robin M. Meyers, Jordan G. Bryan, James M. McFarland, Barbara A. Weir,... David E. Root, William C. Hahn, & Aviad Tsherniak

The CRISPR–Cas9 system has revolutionized gene editing both at single genes and in multiplexed loss-of-function screens, thus enabling precise genome-scale identification of genes essential for proliferation and survival of cancer cells1,2. However, previous studies have reported that a gene-independent antiproliferative effect of Cas9-mediated DNA cleavage confounds such measurement of genetic dependency, thereby leading to false-positive results in copy number–amplified regions3,4. We developed CERES, a computational method...

Cell | 2017.07.27

Defining a Cancer Dependency Map

Aviad Tsherniak, Francisca Vazquez, Phillip G. Montgomery, Barbara A. Weir,... Todd R. Golub, Jesse S. Boehm, & William C. Hahn

Most human epithelial tumors harbor numerous alterations, making it difficult to predict which genes are required for tumor survival. To systematically identify cancer dependencies, we analyzed 501 genome-scale loss-of-function screens performed in diverse human cancer cell lines. We developed DEMETER, an analytical framework that segregates on- from off-target effects of RNAi. 769 genes were differentially required in subsets of these cell lines at a threshold of six SDs from the mean. We found predictive models for 426 dependencies....

Nature | 2017.07.19

In vivo CRISPR screening identifies Ptpn2 as a cancer immunotherapy target

Robert T. Manguso, Hans W. Pope, Margaret D. Zimmer, Flavian D. Brown,... David E. Root, Arlene H. Sharpe, John G. Doench & W. Nicholas Haining

Immunotherapy with PD-1 checkpoint blockade is effective in only a minority of patients with cancer, suggesting that additional treatment strategies are needed. Here we use a pooled in vivo genetic screening approach using CRISPR–Cas9 genome editing in transplantable tumours in mice treated with immunotherapy to discover previously undescribed immunotherapy targets. We tested 2,368 genes expressed by melanoma cells to identify those that synergize with or cause resistance to checkpoint blockade. We recovered the known immune...

Nature Medicine | 2017.04.07

The Drug Repurposing Hub: a next-generation drug library and information resource

Steven M. Corsello, Joshua A. Bittker, Zihan Liu, Joshua Gould,... Christopher C. Mader, Aravind Subramanian, & Todd R. Golub

Drug repurposing, the application of an existing therapeutic to a new disease indication, holds the promise of rapid clinical impact at a lower cost than de novo drug development. To date there has not been a systematic effort to identify such opportunities, limited in part by the lack of a comprehensive library of clinical compounds suitable for testing. To address this challenge, we hand-curated a collection of 4,707 compounds, experimentally confirmed their identity, and annotated them with literature-reported targets...

Cell | 2016.12.15

Perturb-Seq: Dissecting Molecular Circuits with Scalable Single-Cell RNA Profiling of Pooled Genetic Screens

Atray Dixit, Oren Parnas, Biyu Li, Jenny Chen,... Eric S. Lander, Jonathan S. Weissman, Nir Friedman, & Aviv Regev

Genetic screens help infer gene function in mammalian cells, but it has remained difficult to assay complex phenotypes-such as transcriptional profiles-at scale. Here, we develop Perturb-seq, combining single-cell RNA sequencing (RNA-seq) and clustered regularly interspaced short palindromic repeats (CRISPR)-based perturbations to perform many such assays in a pool. We demonstrate Perturb-seq by analyzing 200,000 cells in immune cells and cell lines, focusing on transcription factors regulating the response of dendritic cells...

Science | 2016.03.11

MTAP deletion confers enhanced dependency on the PRMT5 arginine methyltransferase in cancer cells

Gregory V. Kryukov, Frederick H. Wilson, Jason R. Ruth, Joshiawa Paulk,... Clary B. Clish, James E. Bradner, William C. Hahn, Levi A. Garraway

The discovery of cancer dependencies has the potential to inform therapeutic strategies and to identify putative drug targets. Integrating data from comprehensive genomic profiling of cancer cell lines and from functional characterization of cancer cell dependencies, we discovered that loss of the enzyme methylthioadenosine phosphorylase (MTAP) confers a selective dependence on protein arginine methyltransferase 5 (PRMT5) and its binding partner WDR77. MTAP is frequently lost due to its proximity to the commonly deleted tumor...

Nature Biotechnology | 2016.02.29

High-throughput identification of genotype-specific cancer vulnerabilities in mixtures of barcoded tumor cell lines

Channing Yu, Aristotle M. Mannan, Griselda Metta Yvone, Kenneth N. Ross,... Stuart L. Schreiber, Andrew L. Kung, & Todd R. Golub

Hundreds of genetically characterized cell lines are available for the discovery of genotype-specific cancer vulnerabilities. However, screening large numbers of compounds against large numbers of cell lines is currently impractical, and such experiments are often difficult to control1,2,3,4. Here we report a method called PRISM that allows pooled screening of mixtures of cancer cell lines by labeling each cell line with 24-nucleotide barcodes. PRISM revealed the expected patterns of cell killing seen in conventional (unpooled) assays...

Nature Chemical Biology | 2015.12.14

Correlating chemical sensitivity and basal gene expression reveals mechanism of action

Matthew G. Rees, Brinton Seashore-Ludlow, Jaime H. Cheah, Drew J. Adams,... Paul A. Clemons, Alykhan F. Shamji, & Stuart L. Schreiber

Changes in cellular gene expression in response to small-molecule or genetic perturbations have yielded signatures that can connect unknown mechanisms of action (MoA) to ones previously established. We hypothesized that differential basal gene expression could be correlated with patterns of small-molecule sensitivity across many cell lines to illuminate the actions of compounds whose MoA are unknown. To test this idea, we correlated the sensitivity patterns of 481 compounds with ∼19,000 basal transcript levels across 823 different...

Nature | 2015.11.16

Pharmacogenomic agreement between two cancer cell line data sets

The Cancer Cell Line Encyclopedia Consortium & The Genomics of Drug Sensitivity in Cancer Consortium

Large cancer cell line collections broadly capture the genomic diversity of human cancers and provide valuable insight into anti-cancer drug response. Here we show substantial agreement and biological consilience between drug sensitivity measurements and their associated genomic predictors from two publicly available large-scale pharmacogenomics resources: The Cancer Cell Line Encyclopedia and the Genomics of Drug Sensitivity in Cancer databases....

Nature | 2012.03.28

The Cancer Cell Line Encyclopedia enables predictive modelling of anticancer drug sensitivity

TJordi Barretina, Giordano Caponigro, Nicolas Stransky, Kavitha Venkatesan,... William R. Sellers, Robert Schlegel, & Levi A. Garraway

The systematic translation of cancer genomic data into knowledge of tumour biology and therapeutic possibilities remains challenging. Such efforts should be greatly aided by robust preclinical model systems that reflect the genomic diversity of human cancers and for which detailed genetic and pharmacological annotation is available1. Here we describe the Cancer Cell Line Encyclopedia (CCLE): a compilation of gene expression, chromosomal copy number and massively parallel sequencing data from 947 human cancer cell lines. When coupled with pharmacological profiles for 24 anticancer drugs across 479 of the cell lines....