Publications by authors named "Charles W M Roberts"

69 Publications

A first-generation pediatric cancer dependency map.

Nat Genet 2021 04 22;53(4):529-538. Epub 2021 Mar 22.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, MA, USA.

Exciting therapeutic targets are emerging from CRISPR-based screens of high mutational-burden adult cancers. A key question, however, is whether functional genomic approaches will yield new targets in pediatric cancers, known for remarkably few mutations, which often encode proteins considered challenging drug targets. To address this, we created a first-generation pediatric cancer dependency map representing 13 pediatric solid and brain tumor types. Eighty-two pediatric cancer cell lines were subjected to genome-scale CRISPR-Cas9 loss-of-function screening to identify genes required for cell survival. In contrast to the finding that pediatric cancers harbor fewer somatic mutations, we found a similar complexity of genetic dependencies in pediatric cancer cell lines compared to that in adult models. Findings from the pediatric cancer dependency map provide preclinical support for ongoing precision medicine clinical trials. The vulnerabilities observed in pediatric cancers were often distinct from those in adult cancer, indicating that repurposing adult oncology drugs will be insufficient to address childhood cancers.
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http://dx.doi.org/10.1038/s41588-021-00819-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8049517PMC
April 2021

Partitioning of Chemotherapeutics into Nuclear Condensates-Opening the Door to New Approaches for Drug Development.

Mol Cell 2020 08;79(4):544-545

Comprehensive Cancer Center and Department of Oncology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA. Electronic address:

Klein et al. (2020) demonstrate for the first time that small-molecule cancer therapeutics are selectively partitioned and concentrated within phase-separated nuclear condensates, providing new insights to drug efficacy and creating the opportunity for enhanced control of therapeutic targeting.
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http://dx.doi.org/10.1016/j.molcel.2020.07.029DOI Listing
August 2020

Rhabdoid Tumors Are Sensitive to the Protein-Translation Inhibitor Homoharringtonine.

Clin Cancer Res 2020 09 6;26(18):4995-5006. Epub 2020 Jul 6.

Department of Pediatric Oncology, Dana-Farber Cancer Institute and Division of Hematology/Oncology, Boston Children's Hospital, Boston, Massachusetts.

Purpose: Rhabdoid tumors are devastating pediatric cancers in need of improved therapies. We sought to identify small molecules that exhibit and efficacy against preclinical models of rhabdoid tumor.

Experimental Design: We screened eight rhabdoid tumor cell lines with 481 small molecules and compared their sensitivity with that of 879 other cancer cell lines. Genome-scale CRISPR-Cas9 inactivation screens in rhabdoid tumors were analyzed to confirm target vulnerabilities. Gene expression and CRISPR-Cas9 data were queried across cell lines and primary rhabdoid tumors to discover biomarkers of small-molecule sensitivity. Molecular correlates were validated by manipulating gene expression. Subcutaneous rhabdoid tumor xenografts were treated with the most effective drug to confirm results.

Results: Small-molecule screening identified the protein-translation inhibitor homoharringtonine (HHT), an FDA-approved treatment for chronic myelogenous leukemia (CML), as the sole drug to which all rhabdoid tumor cell lines were selectively sensitive. Validation studies confirmed the sensitivity of rhabdoid tumor to HHT was comparable with that of CML cell lines. Low expression of the antiapoptotic gene , which encodes Bcl-XL, was the strongest predictor of HHT sensitivity, and HHT treatment consistently depleted Mcl-1, the synthetic-lethal antiapoptotic partner of Bcl-XL. Rhabdoid tumor cell lines and primary-tumor samples expressed low , and overexpression of induced resistance to HHT in rhabdoid tumor cells. Furthermore, HHT treatment inhibited rhabdoid tumor cell line and patient-derived xenograft growth .

Conclusions: Rhabdoid tumor cell lines and xenografts are highly sensitive to HHT, at least partially due to their low expression of . HHT may have therapeutic potential against rhabdoid tumors.
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http://dx.doi.org/10.1158/1078-0432.CCR-19-2717DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7501142PMC
September 2020

The SWI/SNF complex in cancer - biology, biomarkers and therapy.

Nat Rev Clin Oncol 2020 07 17;17(7):435-448. Epub 2020 Apr 17.

Department of Oncology, St. Jude Children's Research Hospital, Memphis, TN, USA.

Cancer genome-sequencing studies have revealed a remarkably high prevalence of mutations in genes encoding subunits of the SWI/SNF chromatin-remodelling complexes, with nearly 25% of all cancers harbouring aberrations in one or more of these genes. A role for such aberrations in tumorigenesis is evidenced by cancer predisposition in both carriers of germline loss-of-function mutations and genetically engineered mouse models with inactivation of any of several SWI/SNF subunits. Whereas many of the most frequently mutated oncogenes and tumour-suppressor genes have been studied for several decades, the cancer-promoting role of mutations in SWI/SNF genes has been recognized only more recently, and thus comparatively less is known about these alterations. Consequently, increasing research interest is being focused on understanding the prognostic and, in particular, the potential therapeutic implications of mutations in genes encoding SWI/SNF subunits. Herein, we review the burgeoning data on the mechanisms by which mutations affecting SWI/SNF complexes promote cancer and describe promising emerging opportunities for targeted therapy, including immunotherapy with immune-checkpoint inhibitors, presented by these mutations. We also highlight ongoing clinical trials open specifically to patients with cancers harbouring mutations in certain SWI/SNF genes.
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http://dx.doi.org/10.1038/s41571-020-0357-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8723792PMC
July 2020

Author Correction: BRD9 defines a SWI/SNF sub-complex and constitutes a specific vulnerability in malignant rhabdoid tumors.

Nat Commun 2019 Sep 26;10(1):4445. Epub 2019 Sep 26.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, MA, 02215, USA.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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http://dx.doi.org/10.1038/s41467-019-12524-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6763484PMC
September 2019

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

Cell Rep 2019 08;28(9):2331-2344.e8

Comprehensive Cancer Center and Department of Oncology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA. Electronic address:

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 tyrosine kinases (RTKs) as therapeutic targets, with RTK inhibition effective in suppressing RT cell growth in vitro and against a xenograft model in vivo. RT cell lines highly express and activate (phosphorylate) different RTKs, creating dependency without mutation or amplification. Downstream of RTK signaling, we identified PTPN11, encoding the pro-growth signaling protein SHP2, as a shared dependency across all RT cell lines. This study demonstrates that large-scale perturbational screening can uncover vulnerabilities in cancers with "quiet" genomes.
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http://dx.doi.org/10.1016/j.celrep.2019.07.021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7319190PMC
August 2019

BRD9 defines a SWI/SNF sub-complex and constitutes a specific vulnerability in malignant rhabdoid tumors.

Nat Commun 2019 04 23;10(1):1881. Epub 2019 Apr 23.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, MA, 02215, USA.

Bromodomain-containing protein 9 (BRD9) is a recently identified subunit of SWI/SNF(BAF) chromatin remodeling complexes, yet its function is poorly understood. Here, using a genome-wide CRISPR-Cas9 screen, we show that BRD9 is a specific vulnerability in pediatric malignant rhabdoid tumors (RTs), which are driven by inactivation of the SMARCB1 subunit of SWI/SNF. We find that BRD9 exists in a unique SWI/SNF sub-complex that lacks SMARCB1, which has been considered a core subunit. While SMARCB1-containing SWI/SNF complexes are bound preferentially at enhancers, we show that BRD9-containing complexes exist at both promoters and enhancers. Mechanistically, we show that SMARCB1 loss causes increased BRD9 incorporation into SWI/SNF thus providing insight into BRD9 vulnerability in RTs. Underlying the dependency, while its bromodomain is dispensable, the DUF3512 domain of BRD9 is essential for SWI/SNF integrity in the absence of SMARCB1. Collectively, our results reveal a BRD9-containing SWI/SNF subcomplex is required for the survival of SMARCB1-mutant RTs.
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http://dx.doi.org/10.1038/s41467-019-09891-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6479050PMC
April 2019

MDM2 and MDM4 Are Therapeutic Vulnerabilities in Malignant Rhabdoid Tumors.

Cancer Res 2019 05 12;79(9):2404-2414. Epub 2019 Feb 12.

Department of Pediatric Oncology, Dana-Farber Cancer Institute and Division of Hematology/Oncology, Boston Children's Hospital, Boston, Massachusetts.

Malignant rhabdoid tumors (MRT) are highly aggressive pediatric cancers that respond poorly to current therapies. In this study, we screened several MRT cell lines with large-scale RNAi, CRISPR-Cas9, and small-molecule libraries to identify potential drug targets specific for these cancers. We discovered and , the canonical negative regulators of p53, as significant vulnerabilities. Using two compounds currently in clinical development, idasanutlin (MDM2-specific) and ATSP-7041 (MDM2/4-dual), we show that MRT cells were more sensitive than other p53 wild-type cancer cell lines to inhibition of MDM2 alone as well as dual inhibition of MDM2/4. These compounds caused significant upregulation of the p53 pathway in MRT cells, and sensitivity was ablated by CRISPR-Cas9-mediated inactivation of . We show that loss of SMARCB1, a subunit of the SWI/SNF (BAF) complex mutated in nearly all MRTs, sensitized cells to MDM2 and MDM2/4 inhibition by enhancing p53-mediated apoptosis. Both MDM2 and MDM2/4 inhibition slowed MRT xenograft growth , with a 5-day idasanutlin pulse causing marked regression of all xenografts, including durable complete responses in 50% of mice. Together, these studies identify a genetic connection between mutations in the SWI/SNF chromatin-remodeling complex and the tumor suppressor gene and provide preclinical evidence to support the targeting of MDM2 and MDM4 in this often-fatal pediatric cancer. SIGNIFICANCE: This study identifies two targets, MDM2 and MDM4, as vulnerabilities in a deadly pediatric cancer and provides preclinical evidence that compounds inhibiting these proteins have therapeutic potential.
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http://dx.doi.org/10.1158/0008-5472.CAN-18-3066DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6497578PMC
May 2019

p53 Is a Master Regulator of Proteostasis in SMARCB1-Deficient Malignant Rhabdoid Tumors.

Cancer Cell 2019 02;35(2):204-220.e9

Institute for Applied Cancer Science, The University of Texas MD Anderson Cancer Center, 1515 Holcombe Boulevard, Houston, TX 77030, USA.

Alterations in chromatin remodeling genes have been increasingly implicated in human oncogenesis. Specifically, the biallelic inactivation of the SWI/SNF subunit SMARCB1 results in the emergence of extremely aggressive pediatric malignancies. Here, we developed embryonic mosaic mouse models of malignant rhabdoid tumors (MRTs) that faithfully recapitulate the clinical-pathological features of the human disease. We demonstrated that SMARCB1-deficient malignancies exhibit dramatic activation of the unfolded protein response (UPR) and ER stress response via a genetically intact MYC-p19-p53 axis. As a consequence, these tumors display an exquisite sensitivity to agents inducing proteotoxic stress and inhibition of the autophagic machinery. In conclusion, our findings provide a rationale for drug repositioning trials investigating combinations of agents targeting the UPR and autophagy in SMARCB1-deficient MRTs.
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http://dx.doi.org/10.1016/j.ccell.2019.01.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7876656PMC
February 2019

Comprehensive Analysis of Chromatin States in Atypical Teratoid/Rhabdoid Tumor Identifies Diverging Roles for SWI/SNF and Polycomb in Gene Regulation.

Cancer Cell 2019 01 27;35(1):95-110.e8. Epub 2018 Dec 27.

Hopp-Children's Cancer Center at the NCT Heidelberg (KiTZ), 69120 Heidelberg, Germany; Division of Pediatric Neurooncology, German Cancer Research Center (DKFZ) and German Cancer Consortium (DKTK), Im Neuenheimer Feld 280, 69120 Heidelberg, Germany. Electronic address:

Biallelic inactivation of SMARCB1, encoding a member of the SWI/SNF chromatin remodeling complex, is the hallmark genetic aberration of atypical teratoid rhabdoid tumors (ATRT). Here, we report how loss of SMARCB1 affects the epigenome in these tumors. Using chromatin immunoprecipitation sequencing (ChIP-seq) on primary tumors for a series of active and repressive histone marks, we identified the chromatin states differentially represented in ATRTs compared with other brain tumors and non-neoplastic brain. Re-expression of SMARCB1 in ATRT cell lines enabled confirmation of our genome-wide findings for the chromatin states. Additional generation of ChIP-seq data for SWI/SNF and Polycomb group proteins and the transcriptional repressor protein REST determined differential dependencies of SWI/SNF and Polycomb complexes in regulation of diverse gene sets in ATRTs.
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http://dx.doi.org/10.1016/j.ccell.2018.11.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6341227PMC
January 2019

Mutational processes shape the landscape of TP53 mutations in human cancer.

Nat Genet 2018 10 17;50(10):1381-1387. Epub 2018 Sep 17.

Dana-Farber Cancer Institute, Boston, MA, USA.

Unlike most tumor suppressor genes, the most common genetic alterations in tumor protein p53 (TP53) are missense mutations. Mutant p53 protein is often abundantly expressed in cancers and specific allelic variants exhibit dominant-negative or gain-of-function activities in experimental models. To gain a systematic view of p53 function, we interrogated loss-of-function screens conducted in hundreds of human cancer cell lines and performed TP53 saturation mutagenesis screens in an isogenic pair of TP53 wild-type and null cell lines. We found that loss or dominant-negative inhibition of wild-type p53 function reliably enhanced cellular fitness. By integrating these data with the Catalog of Somatic Mutations in Cancer (COSMIC) mutational signatures database, we developed a statistical model that describes the TP53 mutational spectrum as a function of the baseline probability of acquiring each mutation and the fitness advantage conferred by attenuation of p53 activity. Collectively, these observations show that widely-acting and tissue-specific mutational processes combine with phenotypic selection to dictate the frequencies of recurrent TP53 mutations.
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http://dx.doi.org/10.1038/s41588-018-0204-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6168352PMC
October 2018

High Frequency of Ovarian Cyst Development in Vhl;Snf5 Mice.

Am J Pathol 2018 07 22;188(7):1510-1516. Epub 2018 Apr 22.

Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina; Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, North Carolina. Electronic address:

The new paradigm of mutations in chromatin-modifying genes as driver events in the development of cancers has proved challenging to resolve the complex influences over disease phenotypes. In particular, impaired activities of members of the SWI/SNF chromatin remodeling complex have appeared in an increasing variety of tumors. Mutations in SNF5, a member of this ubiquitously expressed complex, arise in almost all cases of malignant rhabdoid tumor in the absence of additional genetic alterations. Therefore, we studied how activation of additional oncogenic pathways might shift the phenotype of disease driven by SNF5 loss. With the use of a genetically engineered mouse model, we examined the effects of a hypomorphic Vhl allele on disease phenotype, with a modest up-regulation of the hypoxia response pathway. Snf5;Vhl mice did not demonstrate a substantial difference in overall survival or a change in malignant rhabdoid tumor development. However, a high percentage of female mice showed complex hemorrhagic ovarian cysts, a phenotype rarely found in either parental mouse strain. These lesions also showed mosaic expression of SNF5 by immunohistochemistry. Therefore, our studies implicate that modest changes in angiogenic regulation interact with perturbations of SWI/SNF complex activity to modulate disease phenotypes.
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http://dx.doi.org/10.1016/j.ajpath.2018.03.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6024179PMC
July 2018

AP-1 Transcription Factors and the BAF Complex Mediate Signal-Dependent Enhancer Selection.

Mol Cell 2017 12;68(6):1067-1082.e12

Department of Neurobiology, Harvard Medical School, 220 Longwood Avenue, Boston, MA 02115, USA.

Enhancer elements are genomic regulatory sequences that direct the selective expression of genes so that genetically identical cells can differentiate and acquire the highly specialized forms and functions required to build a functioning animal. To differentiate, cells must select from among the ∼10 enhancers encoded in the genome the thousands of enhancers that drive the gene programs that impart their distinct features. We used a genetic approach to identify transcription factors (TFs) required for enhancer selection in fibroblasts. This revealed that the broadly expressed, growth-factor-inducible TFs FOS/JUN (AP-1) play a central role in enhancer selection. FOS/JUN selects enhancers together with cell-type-specific TFs by collaboratively binding to nucleosomal enhancers and recruiting the SWI/SNF (BAF) chromatin remodeling complex to establish accessible chromatin. These experiments demonstrate how environmental signals acting via FOS/JUN and BAF coordinate with cell-type-specific TFs to select enhancer repertoires that enable differentiation during development.
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http://dx.doi.org/10.1016/j.molcel.2017.11.026DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5744881PMC
December 2017

Renal Medullary Carcinoma: Establishing Standards in Practice.

J Oncol Pract 2017 07;13(7):414-421

Vanderbilt University Medical Center, Nashville; St Jude Children's Research Hospital, Memphis, TN; University of Texas MD Anderson Cancer Center; Baylor College of Medicine, Houston, TX; American University of Beirut Medical Center, Beirut, Lebanon; Université Paris-Est Créteil, Créteil; Institut Curie; University Pierre and Marie Curie, Paris, France; University of North Carolina at Chapel Hill, Chapel Hill, NC; Dana-Farber/Boston Children's Cancer and Blood Disorders Center, Boston; Broad Institute of Harvard and Massachusetts Institute of Technology, Cambridge, MA; Johns Hopkins University, Baltimore, MD; Memorial Sloan Kettering Cancer Center, New York, NY; and University Hospitals Leuven, Leuven, Belgium.

Although renal medullary carcinoma (RMC) is a rare subtype of kidney cancer, it is particularly devastating in that it is nearly uniformly lethal. No established guidelines exist for the diagnosis and management of RMC. In April 2016, a panel of experts developed clinical guidelines on the basis of a literature review and consensus statements. The goal was to propose recommendations for standardized diagnostic and management approaches and to establish an international clinical registry and biorepository for RMC. Published data are limited to case reports and small retrospective reviews. The RMC Working Group prepared recommendations to inform providers and patients faced with a low level of medical evidence. The diagnosis of RMC should be considered in all patients younger than 50 years with poorly differentiated carcinoma that arises from the renal medulla. These patients should be tested for sickle cell hemoglobinopathies, and if positive, SMARCB1/INI1 loss should be confirmed by immunohistochemistry. The majority of patients with RMC are diagnosed with metastatic disease. Upfront radical nephrectomy should be considered in patients with good performance status and low metastatic burden or after response to systemic therapy. Currently, cytotoxic, platinum-based chemotherapy provides the best, albeit brief, palliative clinical benefit. Vascular endothelial growth factor-directed therapies and mammalian target of rapamycin inhibitors are ineffective in RMC as monotherapy. Therapeutic trials of novel agents are now available for RMC, and every effort should be made to enroll patients in clinical studies.
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http://dx.doi.org/10.1200/JOP.2017.020909DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5508447PMC
July 2017

PGBD5 promotes site-specific oncogenic mutations in human tumors.

Nat Genet 2017 Jul 15;49(7):1005-1014. Epub 2017 May 15.

Molecular Pharmacology Program, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, New York, USA.

Genomic rearrangements are a hallmark of human cancers. Here, we identify the piggyBac transposable element derived 5 (PGBD5) gene as encoding an active DNA transposase expressed in the majority of childhood solid tumors, including lethal rhabdoid tumors. Using assembly-based whole-genome DNA sequencing, we found previously undefined genomic rearrangements in human rhabdoid tumors. These rearrangements involved PGBD5-specific signal (PSS) sequences at their breakpoints and recurrently inactivated tumor-suppressor genes. PGBD5 was physically associated with genomic PSS sequences that were also sufficient to mediate PGBD5-induced DNA rearrangements in rhabdoid tumor cells. Ectopic expression of PGBD5 in primary immortalized human cells was sufficient to promote cell transformation in vivo. This activity required specific catalytic residues in the PGBD5 transposase domain as well as end-joining DNA repair and induced structural rearrangements with PSS breakpoints. These results define PGBD5 as an oncogenic mutator and provide a plausible mechanism for site-specific DNA rearrangements in childhood and adult solid tumors.
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http://dx.doi.org/10.1038/ng.3866DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5489359PMC
July 2017

The SWI/SNF chromatin remodelling complex is required for maintenance of lineage specific enhancers.

Nat Commun 2017 03 6;8:14648. Epub 2017 Mar 6.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts 02215, USA.

Genes encoding subunits of SWI/SNF (BAF) chromatin remodelling complexes are collectively altered in over 20% of human malignancies, but the mechanisms by which these complexes alter chromatin to modulate transcription and cell fate are poorly understood. Utilizing mouse embryonic fibroblast and cancer cell line models, here we show via ChIP-seq and biochemical assays that SWI/SNF complexes are preferentially targeted to distal lineage specific enhancers and interact with p300 to modulate histone H3 lysine 27 acetylation. We identify a greater requirement for SWI/SNF at typical enhancers than at most super-enhancers and at enhancers in untranscribed regions than in transcribed regions. Our data further demonstrate that SWI/SNF-dependent distal enhancers are essential for controlling expression of genes linked to developmental processes. Our findings thus establish SWI/SNF complexes as regulators of the enhancer landscape and provide insight into the roles of SWI/SNF in cellular fate control.
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http://dx.doi.org/10.1038/ncomms14648DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5343482PMC
March 2017

Synthetic vulnerabilities of mesenchymal subpopulations in pancreatic cancer.

Nature 2017 02 8;542(7641):362-366. Epub 2017 Feb 8.

Institute for Applied Cancer Science, The University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA.

Malignant neoplasms evolve in response to changes in oncogenic signalling. Cancer cell plasticity in response to evolutionary pressures is fundamental to tumour progression and the development of therapeutic resistance. Here we determine the molecular and cellular mechanisms of cancer cell plasticity in a conditional oncogenic Kras mouse model of pancreatic ductal adenocarcinoma (PDAC), a malignancy that displays considerable phenotypic diversity and morphological heterogeneity. In this model, stochastic extinction of oncogenic Kras signalling and emergence of Kras-independent escaper populations (cells that acquire oncogenic properties) are associated with de-differentiation and aggressive biological behaviour. Transcriptomic and functional analyses of Kras-independent escapers reveal the presence of Smarcb1-Myc-network-driven mesenchymal reprogramming and independence from MAPK signalling. A somatic mosaic model of PDAC, which allows time-restricted perturbation of cell fate, shows that depletion of Smarcb1 activates the Myc network, driving an anabolic switch that increases protein metabolism and adaptive activation of endoplasmic-reticulum-stress-induced survival pathways. Increased protein turnover renders mesenchymal sub-populations highly susceptible to pharmacological and genetic perturbation of the cellular proteostatic machinery and the IRE1-α-MKK4 arm of the endoplasmic-reticulum-stress-response pathway. Specifically, combination regimens that impair the unfolded protein responses block the emergence of aggressive mesenchymal subpopulations in mouse and patient-derived PDAC models. These molecular and biological insights inform a potential therapeutic strategy for targeting aggressive mesenchymal features of PDAC.
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http://dx.doi.org/10.1038/nature21064DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7609022PMC
February 2017

ARID1A loss impairs enhancer-mediated gene regulation and drives colon cancer in mice.

Nat Genet 2017 Feb 12;49(2):296-302. Epub 2016 Dec 12.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.

Genes encoding subunits of SWI/SNF (BAF) chromatin-remodeling complexes are collectively mutated in ∼20% of all human cancers. Although ARID1A is the most frequent target of mutations, the mechanism by which its inactivation promotes tumorigenesis is unclear. Here we demonstrate that Arid1a functions as a tumor suppressor in the mouse colon, but not the small intestine, and that invasive ARID1A-deficient adenocarcinomas resemble human colorectal cancer (CRC). These tumors lack deregulation of APC/β-catenin signaling components, which are crucial gatekeepers in common forms of intestinal cancer. We find that ARID1A normally targets SWI/SNF complexes to enhancers, where they function in coordination with transcription factors to facilitate gene activation. ARID1B preserves SWI/SNF function in ARID1A-deficient cells, but defects in SWI/SNF targeting and control of enhancer activity cause extensive dysregulation of gene expression. These findings represent an advance in colon cancer modeling and implicate enhancer-mediated gene regulation as a principal tumor-suppressor function of ARID1A.
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http://dx.doi.org/10.1038/ng.3744DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5285448PMC
February 2017

SMARCB1-mediated SWI/SNF complex function is essential for enhancer regulation.

Nat Genet 2017 Feb 12;49(2):289-295. Epub 2016 Dec 12.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.

SMARCB1 (also known as SNF5, INI1, and BAF47), a core subunit of the SWI/SNF (BAF) chromatin-remodeling complex, is inactivated in nearly all pediatric rhabdoid tumors. These aggressive cancers are among the most genomically stable, suggesting an epigenetic mechanism by which SMARCB1 loss drives transformation. Here we show that, despite having indistinguishable mutational landscapes, human rhabdoid tumors exhibit distinct enhancer H3K27ac signatures, which identify remnants of differentiation programs. We show that SMARCB1 is required for the integrity of SWI/SNF complexes and that its loss alters enhancer targeting-markedly impairing SWI/SNF binding to typical enhancers, particularly those required for differentiation, while maintaining SWI/SNF binding at super-enhancers. We show that these retained super-enhancers are essential for rhabdoid tumor survival, including some that are shared by all subtypes, such as SPRY1, and other lineage-specific super-enhancers, such as SOX2 in brain-derived rhabdoid tumors. Taken together, our findings identify a new chromatin-based epigenetic mechanism underlying the tumor-suppressive activity of SMARCB1.
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http://dx.doi.org/10.1038/ng.3746DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5285474PMC
February 2017

Integrated genetic and pharmacologic interrogation of rare cancers.

Nat Commun 2016 06 22;7:11987. Epub 2016 Jun 22.

Broad Institute of Harvard and MIT, 415 Main Street, Cambridge, Massachusetts 02142, USA.

Identifying therapeutic targets in rare cancers remains challenging due to the paucity of established models to perform preclinical studies. As a proof-of-concept, we developed a patient-derived cancer cell line, CLF-PED-015-T, from a paediatric patient with a rare undifferentiated sarcoma. Here, we confirm that this cell line recapitulates the histology and harbours the majority of the somatic genetic alterations found in a metastatic lesion isolated at first relapse. We then perform pooled CRISPR-Cas9 and RNAi loss-of-function screens and a small-molecule screen focused on druggable cancer targets. Integrating these three complementary and orthogonal methods, we identify CDK4 and XPO1 as potential therapeutic targets in this cancer, which has no known alterations in these genes. These observations establish an approach that integrates new patient-derived models, functional genomics and chemical screens to facilitate the discovery of targets in rare cancers.
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http://dx.doi.org/10.1038/ncomms11987DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4917959PMC
June 2016

Targeting EZH2 in cancer.

Nat Med 2016 Feb;22(2):128-34

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.

Recent genomic studies have resulted in an emerging understanding of the role of chromatin regulators in the development of cancer. EZH2, a histone methyl transferase subunit of a Polycomb repressor complex, is recurrently mutated in several forms of cancer and is highly expressed in numerous others. Notably, both gain-of-function and loss-of-function mutations occur in cancers but are associated with distinct cancer types. Here we review the spectrum of EZH2-associated mutations, discuss the mechanisms underlying EZH2 function, and synthesize a unifying perspective that the promotion of cancer arises from disruption of the role of EZH2 as a master regulator of transcription. We further discuss EZH2 inhibitors that are now showing early signs of promise in clinical trials and also additional strategies to combat roles of EZH2 in cancer.
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http://dx.doi.org/10.1038/nm.4036DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4918227PMC
February 2016

Multicenter Feasibility Study of Tumor Molecular Profiling to Inform Therapeutic Decisions in Advanced Pediatric Solid Tumors: The Individualized Cancer Therapy (iCat) Study.

JAMA Oncol 2016 May;2(5):608-615

Dana-Farber/Boston Children's Cancer and Blood Disorders Center, Boston, Massachusetts6Harvard Medical School, Boston, Massachusetts.

Importance: Pediatric cancers represent a unique case with respect to cancer genomics and precision medicine, as the mutation frequency is low, and targeted therapies are less available. Consequently, it is unknown whether clinical sequencing can be of benefit.

Objective: To assess the feasibility of identifying actionable alterations and making individualized cancer therapy (iCat) recommendations in pediatric patients with extracranial solid tumors.

Design, Setting, And Participants: Clinical sequencing study at 4 academic medical centers enrolling patients between September 5, 2012, and November 19, 2013, with 1 year of clinical follow-up. Participants were 30 years or younger with high-risk, recurrent, or refractory extracranial solid tumors. The data analysis was performed October 28, 2014.

Interventions: Tumor profiling performed on archived clinically acquired specimens consisted of mutation detection by a Sequenom assay or targeted next-generation sequencing and copy number assessment by array comparative genomic hybridization. Results were reviewed by a multidisciplinary expert panel, and iCat recommendations were made if an actionable alteration was present, and an appropriate drug was available.

Main Outcomes And Measures: Feasibility was assessed using a 2-stage design based on the proportion of patients with recommendations.

Results: Of 100 participants (60 male; median [range] age, 13.4 [0.8-29.8] years), profiling was technically successful in 89 (89% [95% CI, 83%-95%]). Median (range) follow-up was 6.8 (2.0-23.6) months. Overall, 31 (31% [95% CI, 23%-41%]) patients received an iCat recommendation and 3 received matched therapy. The most common actionable alterations leading to an iCat recommendation were cancer-associated signaling pathway gene mutations (n = 10) and copy number alterations in MYC/MYCN (n = 6) and cell cycle genes (n = 11). Additional alterations with implications for clinical care but not resulting in iCat recommendations were identified, including mutations indicating the possible presence of a cancer predisposition syndrome and translocations suggesting a change in diagnosis. In total, 43 (43% [95% CI, 33%-53%]) participants had results with potential clinical significance.

Conclusions And Relevance: A multi-institution clinical genomics study in pediatric oncology is feasible and a substantial proportion of relapsed or refractory pediatric solid tumors have actionable alterations.

Trial Registration: clinicaltrials.gov Identifier: NCT01853345.
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http://dx.doi.org/10.1001/jamaoncol.2015.5689DOI Listing
May 2016

Atypical teratoid/rhabdoid tumors-current concepts, advances in biology, and potential future therapies.

Neuro Oncol 2016 06 10;18(6):764-78. Epub 2016 Jan 10.

Children's Hospital and Swabian Children's Cancer Center, Augsburg, Germany (M.C.F.); Department of Pathology and Laboratory Medicine, Children's Hospital Los Angeles, Los Angeles, California (J.A.B.); INSERM U830, Laboratory of Genetics and Biology of Cancers, and Department of Pediatric Oncology, Curie Institute, Paris, France (F.B.); Comprehensive Cancer Center and Department of Oncology, St Jude Children's Research Hospital, Memphis, Tennessee (C.W.M.R.); Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts (S.N.C.); Division of Hematology/Oncology, Boston Children's Hospital, Boston, Massachusetts (S.N.C.); Department of Pediatrics, Harvard Medical School, Boston, Massachusetts (S.N.C.).

Atypical teratoid/rhabdoid tumor (AT/RT) is the most common malignant CNS tumor of children below 6 months of age. The majority of AT/RTs demonstrate genomic alterations in SMARCB1 (INI1, SNF5, BAF47) or, to a lesser extent, SMARCA4 (BRG1) of the SWItch/sucrose nonfermentable chromatin remodeling complex. Recent transcription and methylation profiling studies suggest the existence of molecular subgroups. Thus, at the root of these seemingly enigmatic tumors lies a network of factors related to epigenetic regulation, which is not yet completely understood. While conventional-type chemotherapy may have significant survival benefit for certain patients, it remains to be determined which patients will eventually prove resistant to chemotherapy and thus need novel therapeutic strategies. Elucidation of the molecular consequences of a disturbed epigenome has led to the identification of a series of transduction cascades, which may be targeted for therapy. Among these are the pathways of cyclin D1/cyclin-dependent kinases 4 and 6, Hedgehog/GLI1, Wnt/ß-catenin, enhancer of zeste homolog 2, and aurora kinase A, among others. Compounds specifically targeting these pathways or agents that alter the epigenetic state of the cell are currently being evaluated in preclinical settings and in experimental clinical trials for AT/RT.
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http://dx.doi.org/10.1093/neuonc/nov264DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4864253PMC
June 2016

Ezh2 regulates differentiation and function of natural killer cells through histone methyltransferase activity.

Proc Natl Acad Sci U S A 2015 Dec 14;112(52):15988-93. Epub 2015 Dec 14.

Department of Cell Biology, 2011 Collaborative Innovation Center of Tianjin for Medical Epigenetics, Laboratory of Epigenetics in Development and Tumorigenesis, Tianjin Research Center of Basic Medical Sciences, Tianjin Key Laboratory of Medical Epigenetics, Tianjin Medical University, Tianjin 300070, China;

Changes of histone modification status at critical lineage-specifying gene loci in multipotent precursors can influence cell fate commitment. The contribution of these epigenetic mechanisms to natural killer (NK) cell lineage determination from common lymphoid precursors is not understood. Here we investigate the impact of histone methylation repressive marks (H3 Lys27 trimethylation; H3K27(me3)) on early NK cell differentiation. We demonstrate that selective loss of the histone-lysine N-methyltransferase Ezh2 (enhancer of zeste homolog 2) or inhibition of its enzymatic activity with small molecules unexpectedly increased generation of the IL-15 receptor (IL-15R) CD122(+) NK precursors and mature NK progeny from both mouse and human hematopoietic stem and progenitor cells. Mechanistic studies revealed that enhanced NK cell expansion and cytotoxicity against tumor cells were associated with up-regulation of CD122 and the C-type lectin receptor NKG2D. Moreover, NKG2D deficiency diminished the positive effects of Ezh2 inhibitors on NK cell commitment. Identification of the contribution of Ezh2 to NK lineage specification and function reveals an epigenetic-based mechanism that regulates NK cell development and provides insight into the clinical application of Ezh2 inhibitors in NK-based cancer immunotherapies.
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http://dx.doi.org/10.1073/pnas.1521740112DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4702963PMC
December 2015

Molecular analyses reveal close similarities between small cell carcinoma of the ovary, hypercalcemic type and atypical teratoid/rhabdoid tumor.

Oncotarget 2016 Jan;7(2):1732-40

Department of Human Genetics, McGill University, Montreal, Quebec, Canada.

Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) is the most common undifferentiated ovarian malignancy diagnosed in women under age 40. We and others recently determined that germline and/or somatic deleterious mutations in SMARCA4 characterize SCCOHT. Alterations in this gene, or the related SWI/SNF chromatin remodeling gene SMARCB1, have been previously reported in atypical teratoid/rhabdoid tumors (ATRTs) and malignant rhabdoid tumors (MRTs). To further describe the somatic landscape of SCCOHT, we performed whole exome sequencing on 14 tumors and their matched normal tissues and compared their genomic alterations with those in ATRT and ovarian high grade serous carcinoma (HGSC). We confirmed that SMARCA4 is the only recurrently mutated gene in SCCOHT, and show that recurrent allelic imbalance is observed exclusively on chromosome 19p, where SMARCA4 resides. By comparing genomic alterations between SCCOHT, ATRT and HGSC, we demonstrate that SCCOHTs, like ATRTs, have a remarkably simple genome and harbor significantly fewer somatic protein-coding mutations and chromosomal alterations than HGSC. Furthermore, a comparison of global DNA methylation profiles of 45 SCCOHTs, 65 ATRTs, and 92 HGSCs demonstrates a strong epigenetic correlation between SCCOHT and ATRT. Our results further confirm that the genomic and epigenomic signatures of SCCOHT are more similar to those of ATRT than HGSC, supporting our previous hypothesis that SCCOHT is a rhabdoid tumor and should be renamed MRT of the ovary. Furthermore, we conclude that SMARCA4 inactivation is the main cause of SCCOHT, and that new distinct therapeutic approaches should be developed to specifically target this devastating tumor.
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http://dx.doi.org/10.18632/oncotarget.6459DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4811493PMC
January 2016

Functionally distinct patterns of nucleosome remodeling at enhancers in glucocorticoid-treated acute lymphoblastic leukemia.

Epigenetics Chromatin 2015 2;8:53. Epub 2015 Dec 2.

Department of Oncology, St. Jude Children's Research Hospital, 262 Danny Thomas Place, MS 281, Memphis, TN 38105 USA.

Background: Precise nucleosome positioning is an increasingly recognized feature of promoters and enhancers, reflecting complex contributions of DNA sequence, nucleosome positioning, histone modification and transcription factor binding to enhancer activity and regulation of gene expression. Changes in nucleosome position and occupancy, histone variants and modifications, and chromatin remodeling are also critical elements of dynamic transcriptional regulation, but poorly understood at enhancers. We investigated glucocorticoid receptor-associated (GR) nucleosome dynamics at enhancers in acute lymphoblastic leukemia.

Results: For the first time, we demonstrate functionally distinct modes of nucleosome remodeling upon chromatin binding by GR, which we term central, non-central, phased, and minimal. Central and non-central remodeling reflect nucleosome eviction by GR and cofactors, respectively. Phased remodeling involves nucleosome repositioning and is associated with rapidly activated enhancers and induction of gene expression. Minimal remodeling sites initially have low levels of enhancer-associated histone modification, but the majority of these regions gain H3K4me2 or H3K27Ac to become de novo enhancers. Minimal remodeling regions are associated with gene ontologies specific to decreased B cell number and mTOR inhibition and may make unique contributions to glucocorticoid-induced leukemia cell death.

Conclusions: Our findings form a novel framework for understanding the dynamic interplay between transcription factor binding, nucleosome remodeling, enhancer function, and gene expression in the leukemia response to glucocorticoids.
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http://dx.doi.org/10.1186/s13072-015-0046-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4667523PMC
December 2015

SWI/SNF-mutant cancers depend on catalytic and non-catalytic activity of EZH2.

Nat Med 2015 Dec 9;21(12):1491-6. Epub 2015 Nov 9.

Department of Pediatric Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts, USA.

Human cancer genome sequencing has recently revealed that genes that encode subunits of SWI/SNF chromatin remodeling complexes are frequently mutated across a wide variety of cancers, and several subunits of the complex have been shown to have bona fide tumor suppressor activity. However, whether mutations in SWI/SNF subunits result in shared dependencies is unknown. Here we show that EZH2, a catalytic subunit of the polycomb repressive complex 2 (PRC2), is essential in all tested cancer cell lines and xenografts harboring mutations of the SWI/SNF subunits ARID1A, PBRM1, and SMARCA4, which are several of the most frequently mutated SWI/SNF subunits in human cancer, but that co-occurrence of a Ras pathway mutation is correlated with abrogation of this dependence. Notably, we demonstrate that SWI/SNF-mutant cancer cells are primarily dependent on a non-catalytic role of EZH2 in the stabilization of the PRC2 complex, and that they are only partially dependent on EZH2 histone methyltransferase activity. These results not only reveal a shared dependency of cancers with genetic alterations in SWI/SNF subunits, but also suggest that EZH2 enzymatic inhibitors now in clinical development may not fully suppress the oncogenic activity of EZH2.
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http://dx.doi.org/10.1038/nm.3968DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4886303PMC
December 2015

Toward understanding and exploiting tumor heterogeneity.

Nat Med 2015 Aug;21(8):846-53

Inserm, UMR-1162, Génomique fonctionnelle des tumeurs solides, Institut Universitaire d'Hématologie (IUH), Paris, France.

The extent of tumor heterogeneity is an emerging theme that researchers are only beginning to understand. How genetic and epigenetic heterogeneity affects tumor evolution and clinical progression is unknown. The precise nature of the environmental factors that influence this heterogeneity is also yet to be characterized. Nature Medicine, Nature Biotechnology and the Volkswagen Foundation organized a meeting focused on identifying the obstacles that need to be overcome to advance translational research in and tumor heterogeneity. Once these key questions were established, the attendees devised potential solutions. Their ideas are presented here.
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http://dx.doi.org/10.1038/nm.3915DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4785013PMC
August 2015
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