Publications by authors named "Mathieu Lupien"

117 Publications

TFEB-mediated endolysosomal activity controls human hematopoietic stem cell fate.

Cell Stem Cell 2021 Jul 28. Epub 2021 Jul 28.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 1L7, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada; Ontario Institute for Cancer Research, Toronto, ON M5G 0A3, Canada. Electronic address:

It is critical to understand how human quiescent long-term hematopoietic stem cells (LT-HSCs) sense demand from daily and stress-mediated cues and then transition into bioenergetically active progeny to differentiate and meet these cellular needs. However, the demand-adapted regulatory circuits of these early steps of hematopoiesis are largely unknown. Here we show that lysosomes, sophisticated nutrient-sensing and signaling centers, are regulated dichotomously by transcription factor EB (TFEB) and MYC to balance catabolic and anabolic processes required for activating LT-HSCs and guiding their lineage fate. TFEB-mediated induction of the endolysosomal pathway causes membrane receptor degradation, limiting LT-HSC metabolic and mitogenic activation, promoting quiescence and self-renewal, and governing erythroid-myeloid commitment. In contrast, MYC engages biosynthetic processes while repressing lysosomal catabolism, driving LT-HSC activation. Our study identifies TFEB-mediated control of lysosomal activity as a central regulatory hub for proper and coordinated stem cell fate determination.
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http://dx.doi.org/10.1016/j.stem.2021.07.003DOI Listing
July 2021

CRISPRi screens reveal a DNA methylation-mediated 3D genome dependent causal mechanism in prostate cancer.

Nat Commun 2021 03 19;12(1):1781. Epub 2021 Mar 19.

Changhai Hospital, Shanghai, China.

Prostate cancer (PCa) risk-associated SNPs are enriched in noncoding cis-regulatory elements (rCREs), yet their modi operandi and clinical impact remain elusive. Here, we perform CRISPRi screens of 260 rCREs in PCa cell lines. We find that rCREs harboring high risk SNPs are more essential for cell proliferation and H3K27ac occupancy is a strong indicator of essentiality. We also show that cell-line-specific essential rCREs are enriched in the 8q24.21 region, with the rs11986220-containing rCRE regulating MYC and PVT1 expression, cell proliferation and tumorigenesis in a cell-line-specific manner, depending on DNA methylation-orchestrated occupancy of a CTCF binding site in between this rCRE and the MYC promoter. We demonstrate that CTCF deposition at this site as measured by DNA methylation level is highly variable in prostate specimens, and observe the MYC eQTL in the 8q24.21 locus in individuals with low CTCF binding. Together our findings highlight a causal mechanism synergistically driven by a risk SNP and DNA methylation-mediated 3D genome architecture, advocating for the integration of genetics and epigenetics in assessing risks conferred by genetic predispositions.
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http://dx.doi.org/10.1038/s41467-021-21867-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7979745PMC
March 2021

Biological and therapeutic implications of a unique subtype of NPM1 mutated AML.

Nat Commun 2021 02 16;12(1):1054. Epub 2021 Feb 16.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.

In acute myeloid leukemia (AML), molecular heterogeneity across patients constitutes a major challenge for prognosis and therapy. AML with NPM1 mutation is a distinct genetic entity in the revised World Health Organization classification. However, differing patterns of co-mutation and response to therapy within this group necessitate further stratification. Here we report two distinct subtypes within NPM1 mutated AML patients, which we label as primitive and committed based on the respective presence or absence of a stem cell signature. Using gene expression (RNA-seq), epigenomic (ATAC-seq) and immunophenotyping (CyToF) analysis, we associate each subtype with specific molecular characteristics, disease differentiation state and patient survival. Using ex vivo drug sensitivity profiling, we show a differential drug response of the subtypes to specific kinase inhibitors, irrespective of the FLT3-ITD status. Differential drug responses of the primitive and committed subtype are validated in an independent AML cohort. Our results highlight heterogeneity among NPM1 mutated AML patient samples based on stemness and suggest that the addition of kinase inhibitors to the treatment of cases with the primitive signature, lacking FLT3-ITD, could have therapeutic benefit.
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http://dx.doi.org/10.1038/s41467-021-21233-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7886883PMC
February 2021

PRMT5 inhibition disrupts splicing and stemness in glioblastoma.

Nat Commun 2021 02 12;12(1):979. Epub 2021 Feb 12.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.

Glioblastoma (GBM) is a deadly cancer in which cancer stem cells (CSCs) sustain tumor growth and contribute to therapeutic resistance. Protein arginine methyltransferase 5 (PRMT5) has recently emerged as a promising target in GBM. Using two orthogonal-acting inhibitors of PRMT5 (GSK591 or LLY-283), we show that pharmacological inhibition of PRMT5 suppresses the growth of a cohort of 46 patient-derived GBM stem cell cultures, with the proneural subtype showing greater sensitivity. We show that PRMT5 inhibition causes widespread disruption of splicing across the transcriptome, particularly affecting cell cycle gene products. We identify a GBM splicing signature that correlates with the degree of response to PRMT5 inhibition. Importantly, we demonstrate that LLY-283 is brain-penetrant and significantly prolongs the survival of mice with orthotopic patient-derived xenografts. Collectively, our findings provide a rationale for the clinical development of brain penetrant PRMT5 inhibitors as treatment for GBM.
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http://dx.doi.org/10.1038/s41467-021-21204-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7881162PMC
February 2021

Large organized chromatin lysine domains help distinguish primitive from differentiated cell populations.

Nat Commun 2021 01 21;12(1):499. Epub 2021 Jan 21.

Princess Margaret Cancer Centre, Toronto, ON, M5G 1L7, Canada.

The human genome is partitioned into a collection of genomic features, inclusive of genes, transposable elements, lamina interacting regions, early replicating control elements and cis-regulatory elements, such as promoters, enhancers, and anchors of chromatin interactions. Uneven distribution of these features within chromosomes gives rise to clusters, such as topologically associating domains (TADs), lamina-associated domains, clusters of cis-regulatory elements or large organized chromatin lysine (K) domains (LOCKs). Here we show that LOCKs from diverse histone modifications discriminate primitive from differentiated cell types. Active LOCKs (H3K4me1, H3K4me3 and H3K27ac) cover a higher fraction of the genome in primitive compared to differentiated cell types while repressive LOCKs (H3K9me3, H3K27me3 and H3K36me3) do not. Active LOCKs in differentiated cells lie proximal to highly expressed genes while active LOCKs in primitive cells tend to be bivalent. Genes proximal to bivalent LOCKs are minimally expressed in primitive cells. Furthermore, bivalent LOCKs populate TAD boundaries and are preferentially bound by regulators of chromatin interactions, including CTCF, RAD21 and ZNF143. Together, our results argue that LOCKs discriminate primitive from differentiated cell populations.
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http://dx.doi.org/10.1038/s41467-020-20830-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7820432PMC
January 2021

Single-cell chromatin accessibility profiling of glioblastoma identifies an invasive cancer stem cell population associated with lower survival.

Elife 2021 Jan 11;10. Epub 2021 Jan 11.

Princess Margaret Cancer Centre, University Health Network, Toronto, Canada.

Chromatin accessibility discriminates stem from mature cell populations, enabling the identification of primitive stem-like cells in primary tumors, such as glioblastoma (GBM) where self-renewing cells driving cancer progression and recurrence are prime targets for therapeutic intervention. We show, using single-cell chromatin accessibility, that primary human GBMs harbor a heterogeneous self-renewing population whose diversity is captured in patient-derived glioblastoma stem cells (GSCs). In-depth characterization of chromatin accessibility in GSCs identifies three GSC states: Reactive, Constructive, and Invasive, each governed by uniquely essential transcription factors and present within GBMs in varying proportions. Orthotopic xenografts reveal that GSC states associate with survival, and identify an invasive GSC signature predictive of low patient survival, in line with the higher invasive properties of Invasive state GSCs compared to Reactive and Constructive GSCs as shown by in vitro and in vivo assays. Our chromatin-driven characterization of GSC states improves prognostic precision and identifies dependencies to guide combination therapies.
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http://dx.doi.org/10.7554/eLife.64090DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7847307PMC
January 2021

The Transition from Quiescent to Activated States in Human Hematopoietic Stem Cells Is Governed by Dynamic 3D Genome Reorganization.

Cell Stem Cell 2021 03 25;28(3):488-501.e10. Epub 2020 Nov 25.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 1L7, Canada; Department of Medical Biophysics, University of Toronto, Toronto, ON M5G 1L7, Canada; Ontario Institute for Cancer Research, Toronto, ON M5G 0A3, Canada. Electronic address:

Lifelong blood production requires long-term hematopoietic stem cells (LT-HSCs), marked by stemness states involving quiescence and self-renewal, to transition into activated short-term HSCs (ST-HSCs) with reduced stemness. As few transcriptional changes underlie this transition, we used single-cell and bulk assay for transposase-accessible chromatin sequencing (ATAC-seq) on human HSCs and hematopoietic stem and progenitor cell (HSPC) subsets to uncover chromatin accessibility signatures, one including LT-HSCs (LT/HSPC signature) and another excluding LT-HSCs (activated HSPC [Act/HSPC] signature). These signatures inversely correlated during early hematopoietic commitment and differentiation. The Act/HSPC signature contains CCCTC-binding factor (CTCF) binding sites mediating 351 chromatin interactions engaged in ST-HSCs, but not LT-HSCs, enclosing multiple stemness pathway genes active in LT-HSCs and repressed in ST-HSCs. CTCF silencing derepressed stemness genes, restraining quiescent LT-HSCs from transitioning to activated ST-HSCs. Hence, 3D chromatin interactions centrally mediated by CTCF endow a gatekeeper function that governs the earliest fate transitions HSCs make by coordinating disparate stemness pathways linked to quiescence and self-renewal.
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http://dx.doi.org/10.1016/j.stem.2020.11.001DOI Listing
March 2021

Pioneer of prostate cancer: past, present and the future of FOXA1.

Protein Cell 2021 01 18;12(1):29-38. Epub 2020 Sep 18.

Princess Margaret Cancer Centre, University Health Network, Toronto, Canada.

Prostate cancer is the most commonly diagnosed non-cutaneous cancers in North American men. While androgen deprivation has remained as the cornerstone of prostate cancer treatment, resistance ensues leading to lethal disease. Forkhead box A1 (FOXA1) encodes a pioneer factor that induces open chromatin conformation to allow the binding of other transcription factors. Through direct interactions with the Androgen Receptor (AR), FOXA1 helps to shape AR signaling that drives the growth and survival of normal prostate and prostate cancer cells. FOXA1 also possesses an AR-independent role of regulating epithelial-to-mesenchymal transition (EMT). In prostate cancer, mutations converge onto the coding sequence and cis-regulatory elements (CREs) of FOXA1, leading to functional alterations. In addition, FOXA1 activity in prostate cancer can be modulated post-translationally through various mechanisms such as LSD1-mediated protein demethylation. In this review, we describe the latest discoveries related to the function and regulation of FOXA1 in prostate cancer, pointing to their relevance to guide future clinical interventions.
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http://dx.doi.org/10.1007/s13238-020-00786-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7815845PMC
January 2021

GLUT1 inhibition blocks growth of RB1-positive triple negative breast cancer.

Nat Commun 2020 08 21;11(1):4205. Epub 2020 Aug 21.

Structural Genomics Consortium, University of Toronto, Toronto, ON, M5G 1L7, Canada.

Triple negative breast cancer (TNBC) is a deadly form of breast cancer due to the development of resistance to chemotherapy affecting over 30% of patients. New therapeutics and companion biomarkers are urgently needed. Recognizing the elevated expression of glucose transporter 1 (GLUT1, encoded by SLC2A1) and associated metabolic dependencies in TNBC, we investigated the vulnerability of TNBC cell lines and patient-derived samples to GLUT1 inhibition. We report that genetic or pharmacological inhibition of GLUT1 with BAY-876 impairs the growth of a subset of TNBC cells displaying high glycolytic and lower oxidative phosphorylation (OXPHOS) rates. Pathway enrichment analysis of gene expression data suggests that the functionality of the E2F pathway may reflect to some extent OXPHOS activity. Furthermore, the protein levels of retinoblastoma tumor suppressor (RB1) strongly correlate with the degree of sensitivity to GLUT1 inhibition in TNBC, where RB1-negative cells are insensitive to GLUT1 inhibition. Collectively, our results highlight a strong and targetable RB1-GLUT1 metabolic axis in TNBC and warrant clinical evaluation of GLUT1 inhibition in TNBC patients stratified according to RB1 protein expression levels.
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http://dx.doi.org/10.1038/s41467-020-18020-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7442809PMC
August 2020

Epigenetic Switch-Induced Viral Mimicry Evasion in Chemotherapy-Resistant Breast Cancer.

Cancer Discov 2020 09 16;10(9):1312-1329. Epub 2020 Jun 16.

Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario, Canada.

Tumor progression upon treatment arises from preexisting resistant cancer cells and/or adaptation of persister cancer cells committing to an expansion phase. Here, we show that evasion from viral mimicry response allows the growth of taxane-resistant triple-negative breast cancer (TNBC). This is enabled by an epigenetic state adapted to taxane-induced metabolic stress, where DNA hypomethylation over loci enriched in transposable elements (TE) is compensated by large chromatin domains of H3K27me3 to warrant TE repression. This epigenetic state creates a vulnerability to epigenetic therapy against EZH2, the H3K27me3 methyltransferase, which alleviates TE repression in taxane-resistant TNBC, leading to double-stranded RNA production and growth inhibition through viral mimicry response. Collectively, our results illustrate how epigenetic states over TEs promote cancer progression under treatment and can inform about vulnerabilities to epigenetic therapy. SIGNIFICANCE: Drug-resistant cancer cells represent a major barrier to remission for patients with cancer. Here we show that drug-induced metabolic perturbation and epigenetic states enable evasion from the viral mimicry response induced by chemotherapy in TNBC. These epigenetic states define a vulnerability to epigenetic therapy using EZH2 inhibitors in taxane-resistant TNBC...
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http://dx.doi.org/10.1158/2159-8290.CD-19-1493DOI Listing
September 2020

Metabolic Regulation of the Epigenome Drives Lethal Infantile Ependymoma.

Cell 2020 06 22;181(6):1329-1345.e24. Epub 2020 May 22.

The Arthur and Sonia Labatt Brain Tumor Research Center, The Hospital for Sick Children, Toronto, ON M5G 0A4, Canada; Developmental & Stem Cell Biology Program, The Hospital for Sick Children, Toronto, ON M5G 0A4, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON M5G 1L7, Canada.

Posterior fossa A (PFA) ependymomas are lethal malignancies of the hindbrain in infants and toddlers. Lacking highly recurrent somatic mutations, PFA ependymomas are proposed to be epigenetically driven tumors for which model systems are lacking. Here we demonstrate that PFA ependymomas are maintained under hypoxia, associated with restricted availability of specific metabolites to diminish histone methylation, and increase histone demethylation and acetylation at histone 3 lysine 27 (H3K27). PFA ependymomas initiate from a cell lineage in the first trimester of human development that resides in restricted oxygen. Unlike other ependymomas, transient exposure of PFA cells to ambient oxygen induces irreversible cellular toxicity. PFA tumors exhibit a low basal level of H3K27me3, and, paradoxically, inhibition of H3K27 methylation specifically disrupts PFA tumor growth. Targeting metabolism and/or the epigenome presents a unique opportunity for rational therapy for infants with PFA ependymoma.
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http://dx.doi.org/10.1016/j.cell.2020.04.047DOI Listing
June 2020

Disrupting Mitochondrial Copper Distribution Inhibits Leukemic Stem Cell Self-Renewal.

Cell Stem Cell 2020 06 15;26(6):926-937.e10. Epub 2020 May 15.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada. Electronic address:

Leukemic stem cells (LSCs) rely on oxidative metabolism and are differentially sensitive to targeting mitochondrial pathways, which spares normal hematopoietic cells. A subset of mitochondrial proteins is folded in the intermembrane space via the mitochondrial intermembrane assembly (MIA) pathway. We found increased mRNA expression of MIA pathway substrates in acute myeloid leukemia (AML) stem cells. Therefore, we evaluated the effects of inhibiting this pathway in AML. Genetic and chemical inhibition of ALR reduces AML growth and viability, disrupts LSC self-renewal, and induces their differentiation. ALR inhibition preferentially decreases its substrate COX17, a mitochondrial copper chaperone, and knockdown of COX17 phenocopies ALR loss. Inhibiting ALR and COX17 increases mitochondrial copper levels which in turn inhibit S-adenosylhomocysteine hydrolase (SAHH) and lower levels of S-adenosylmethionine (SAM), DNA methylation, and chromatin accessibility to lower LSC viability. These results provide insight into mechanisms through which mitochondrial copper controls epigenetic status and viability of LSCs.
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http://dx.doi.org/10.1016/j.stem.2020.04.010DOI Listing
June 2020

Telomere dysfunction cooperates with epigenetic alterations to impair murine embryonic stem cell fate commitment.

Elife 2020 04 16;9. Epub 2020 Apr 16.

Institut de Recherche en Immunologie et Cancérologie (IRIC), Département de biologie moléculaire, Faculté de Médecine, Université de Montréal, Montréal, Canada.

The precise relationship between epigenetic alterations and telomere dysfunction is still an extant question. Previously, we showed that eroded telomeres lead to differentiation instability in murine embryonic stem cells (mESCs) via DNA hypomethylation at pluripotency-factor promoters. Here, we uncovered that telomerase reverse transcriptase null () mESCs exhibit genome-wide alterations in chromatin accessibility and gene expression during differentiation. These changes were accompanied by an increase of H3K27me3 globally, an altered chromatin landscape at the promoter, and a refractory response to differentiation cues. Inhibition of the Polycomb Repressive Complex 2 (PRC2), an H3K27 tri-methyltransferase, exacerbated the impairment in differentiation and pluripotency gene repression in mESCs but not wild-type mESCs, whereas inhibition of H3K27me3 demethylation led to a partial rescue of the phenotype. These data reveal a new interdependent relationship between H3K27me3 and telomere integrity in stem cell lineage commitment that may have implications in aging and cancer.
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http://dx.doi.org/10.7554/eLife.47333DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7192583PMC
April 2020

Author Correction: Human somatic cell mutagenesis creates genetically tractable sarcomas.

Nat Genet 2020 Apr;52(4):464

Department of Medical Biophysics, Ontario Cancer Institute, University of Toronto, Toronto, Ontario, Canada.

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/s41588-020-0589-2DOI Listing
April 2020

Noncoding mutations target cis-regulatory elements of the FOXA1 plexus in prostate cancer.

Nat Commun 2020 01 23;11(1):441. Epub 2020 Jan 23.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.

Prostate cancer is the second most commonly diagnosed malignancy among men worldwide. Recurrently mutated in primary and metastatic prostate tumors, FOXA1 encodes a pioneer transcription factor involved in disease onset and progression through both androgen receptor-dependent and androgen receptor-independent mechanisms. Despite its oncogenic properties however, the regulation of FOXA1 expression remains unknown. Here, we identify a set of six cis-regulatory elements in the FOXA1 regulatory plexus harboring somatic single-nucleotide variants in primary prostate tumors. We find that deletion and repression of these cis-regulatory elements significantly decreases FOXA1 expression and prostate cancer cell growth. Six of the ten single-nucleotide variants mapping to FOXA1 regulatory plexus significantly alter the transactivation potential of cis-regulatory elements by modulating the binding of transcription factors. Collectively, our results identify cis-regulatory elements within the FOXA1 plexus mutated in primary prostate tumors as potential targets for therapeutic intervention.
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http://dx.doi.org/10.1038/s41467-020-14318-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6978390PMC
January 2020

Candidate Cancer Driver Mutations in Distal Regulatory Elements and Long-Range Chromatin Interaction Networks.

Mol Cell 2020 03 17;77(6):1307-1321.e10. Epub 2020 Jan 17.

Computational Biology Program, Ontario Institute for Cancer Research, 661 University Avenue Suite 510, Toronto, ON M5G 0A3, Canada; Department of Medical Biophysics, University of Toronto, 101 College Street Suite 15-701, Toronto, ON M5G 1L7, Canada. Electronic address:

A comprehensive catalog of cancer driver mutations is essential for understanding tumorigenesis and developing therapies. Exome-sequencing studies have mapped many protein-coding drivers, yet few non-coding drivers are known because genome-wide discovery is challenging. We developed a driver discovery method, ActiveDriverWGS, and analyzed 120,788 cis-regulatory modules (CRMs) across 1,844 whole tumor genomes from the ICGC-TCGA PCAWG project. We found 30 CRMs with enriched SNVs and indels (FDR < 0.05). These frequently mutated regulatory elements (FMREs) were ubiquitously active in human tissues, showed long-range chromatin interactions and mRNA abundance associations with target genes, and were enriched in motif-rewiring mutations and structural variants. Genomic deletion of one FMRE in human cells caused proliferative deficiencies and transcriptional deregulation of cancer genes CCNB1IP1, CDH1, and CDKN2B, validating observations in FMRE-mutated tumors. Pathway analysis revealed further sub-significant FMREs at cancer genes and processes, indicating an unexplored landscape of infrequent driver mutations in the non-coding genome.
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http://dx.doi.org/10.1016/j.molcel.2019.12.027DOI Listing
March 2020

Functional Enhancers Shape Extrachromosomal Oncogene Amplifications.

Cell 2019 11 21;179(6):1330-1341.e13. Epub 2019 Nov 21.

Department of Genetics and Genome Sciences, Case Western Reserve University School of Medicine, Case Comprehensive Cancer Center, Cleveland, OH 44106, USA. Electronic address:

Non-coding regions amplified beyond oncogene borders have largely been ignored. Using a computational approach, we find signatures of significant co-amplification of non-coding DNA beyond the boundaries of amplified oncogenes across five cancer types. In glioblastoma, EGFR is preferentially co-amplified with its two endogenous enhancer elements active in the cell type of origin. These regulatory elements, their contacts, and their contribution to cell fitness are preserved on high-level circular extrachromosomal DNA amplifications. Interrogating the locus with a CRISPR interference screening approach reveals a diversity of additional elements that impact cell fitness. The pattern of fitness dependencies mirrors the rearrangement of regulatory elements and accompanying rewiring of the chromatin topology on the extrachromosomal amplicon. Our studies indicate that oncogene amplifications are shaped by regulatory dependencies in the non-coding genome.
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http://dx.doi.org/10.1016/j.cell.2019.10.039DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7241652PMC
November 2019

Cistrome Partitioning Reveals Convergence of Somatic Mutations and Risk Variants on Master Transcription Regulators in Primary Prostate Tumors.

Cancer Cell 2019 12 14;36(6):674-689.e6. Epub 2019 Nov 14.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON M5G 1L7, Canada; Department of Medical Biophysics, University of Toronto, Toronto, ON M5G 1L7, Canada; Ontario Institute for Cancer Research, Toronto, ON M5G 0A3, Canada. Electronic address:

Thousands of noncoding somatic single-nucleotide variants (SNVs) of unknown function are reported in tumors. Partitioning the genome according to cistromes reveals the enrichment of somatic SNVs in prostate tumors as opposed to adjacent normal tissue cistromes of master transcription regulators, including AR, FOXA1, and HOXB13. This parallels enrichment of prostate cancer genetic predispositions over these transcription regulators' tumor cistromes, exemplified at the 8q24 locus harboring both risk variants and somatic SNVs in cis-regulatory elements upregulating MYC expression. However, Massively Parallel Reporter Assays reveal that few SNVs can alter the transactivation potential of individual cis-regulatory elements. Instead, similar to inherited risk variants, SNVs accumulate in cistromes of master transcription regulators required for prostate cancer development.
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http://dx.doi.org/10.1016/j.ccell.2019.10.005DOI Listing
December 2019

Genome-wide germline correlates of the epigenetic landscape of prostate cancer.

Nat Med 2019 10 7;25(10):1615-1626. Epub 2019 Oct 7.

Ontario Institute for Cancer Research, Toronto, Ontario, Canada.

Oncogenesis is driven by germline, environmental and stochastic factors. It is unknown how these interact to produce the molecular phenotypes of tumors. We therefore quantified the influence of germline polymorphisms on the somatic epigenome of 589 localized prostate tumors. Predisposition risk loci influence a tumor's epigenome, uncovering a mechanism for cancer susceptibility. We identified and validated 1,178 loci associated with altered methylation in tumoral but not nonmalignant tissue. These tumor methylation quantitative trait loci influence chromatin structure, as well as RNA and protein abundance. One prominent tumor methylation quantitative trait locus is associated with AKT1 expression and is predictive of relapse after definitive local therapy in both discovery and validation cohorts. These data reveal intricate crosstalk between the germ line and the epigenome of primary tumors, which may help identify germline biomarkers of aggressive disease to aid patient triage and optimize the use of more invasive or expensive diagnostic assays.
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http://dx.doi.org/10.1038/s41591-019-0579-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7418214PMC
October 2019

Identifying clusters of -regulatory elements underpinning TAD structures and lineage-specific regulatory networks.

Genome Res 2019 10 18;29(10):1733-1743. Epub 2019 Sep 18.

Princess Margaret Cancer Centre, Toronto, Ontario M5G 1L7, Canada.

Cellular identity relies on cell-type-specific gene expression controlled at the transcriptional level by -regulatory elements (CREs). CREs are unevenly distributed across the genome, giving rise to individual CREs and clusters of CREs (COREs). Technical and biological features hinder CORE identification. We addressed these issues by developing an unsupervised machine learning approach termed clustering of genomic regions analysis method (CREAM). CREAM automates CORE detection from chromatin accessibility profiles that are enriched in CREs strongly bound by master transcription regulators, proximal to highly expressed and essential genes, and discriminating cell identity. Although COREs share similarities with super-enhancers, we highlight differences in terms of the genomic distribution and structure of these -regulatory units. We further show the enhanced value of COREs over super-enhancers to identify master transcription regulators, highly expressed and essential genes defining cell identity. COREs enrich at topologically associated domain (TAD) boundaries. They are also preferentially bound by the chromatin looping factors CTCF and cohesin, in contrast to super-enhancers, forming clusters of CTCF and cohesin binding regions and defining homotypic clusters of transcription regulator binding regions (HCTs). Finally, we show the clinical utility of CREAM to identify COREs across chromatin accessibility profiles to stratify more than 400 tumor samples according to their cancer type and to delineate cancer type-specific active biological pathways. Collectively, our results support the utility of CREAM to delineate COREs underlying, with greater accuracy than individual CREs or super-enhancers, the cell-type-specific biological underpinning across a wide range of normal and cancer cell types.
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http://dx.doi.org/10.1101/gr.248658.119DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6771399PMC
October 2019

High-resolution structural genomics reveals new therapeutic vulnerabilities in glioblastoma.

Genome Res 2019 08 27;29(8):1211-1222. Epub 2019 Jun 27.

Clark Smith Brain Tumour Centre, Charbonneau Cancer Institute, University of Calgary, Calgary, Alberta T2N 4N1, Canada.

We investigated the role of 3D genome architecture in instructing functional properties of glioblastoma stem cells (GSCs) by generating sub-5-kb resolution 3D genome maps by in situ Hi-C. Contact maps at sub-5-kb resolution allow identification of individual DNA loops, domain organization, and large-scale genome compartmentalization. We observed differences in looping architectures among GSCs from different patients, suggesting that 3D genome architecture is a further layer of inter-patient heterogeneity for glioblastoma. Integration of DNA contact maps with chromatin and transcriptional profiles identified specific mechanisms of gene regulation, including the convergence of multiple super enhancers to individual stemness genes within individual cells. We show that the number of loops contacting a gene correlates with elevated transcription. These results indicate that stemness genes are hubs of interaction between multiple regulatory regions, likely to ensure their sustained expression. Regions of open chromatin common among the GSCs tested were poised for expression of immune-related genes, including We demonstrate that this gene is co-expressed with stemness genes in GSCs and that CD276 can be targeted with an antibody-drug conjugate to eliminate self-renewing cells. Our results demonstrate that integrated structural genomics data sets can be employed to rationally identify therapeutic vulnerabilities in self-renewing cells.
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http://dx.doi.org/10.1101/gr.246520.118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6673710PMC
August 2019

Pervasive H3K27 Acetylation Leads to ERV Expression and a Therapeutic Vulnerability in H3K27M Gliomas.

Cancer Cell 2019 05;35(5):782-797.e8

Department of Pediatrics, Division of Hematology and Oncology, Texas Children's Cancer and Hematology Centers, Dan L Duncan Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA. Electronic address:

High-grade gliomas defined by histone 3 K27M driver mutations exhibit global loss of H3K27 trimethylation and reciprocal gain of H3K27 acetylation, respectively shaping repressive and active chromatin landscapes. We generated tumor-derived isogenic models bearing this mutation and show that it leads to pervasive H3K27ac deposition across the genome. In turn, active enhancers and promoters are not created de novo and instead reflect the epigenomic landscape of the cell of origin. H3K27ac is enriched at repeat elements, resulting in their increased expression, which in turn can be further amplified by DNA demethylation and histone deacetylase inhibitors providing an exquisite therapeutic vulnerability. These agents may therefore modulate anti-tumor immune responses as a therapeutic modality for this untreatable disease.
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http://dx.doi.org/10.1016/j.ccell.2019.04.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6521975PMC
May 2019

A chemical toolbox for the study of bromodomains and epigenetic signaling.

Nat Commun 2019 04 23;10(1):1915. Epub 2019 Apr 23.

Structural Genomics Consortium, Buchmann Institute for Life Sciences, Goethe-University Frankfurt, 60438, Frankfurt, Germany.

Bromodomains (BRDs) are conserved protein interaction modules which recognize (read) acetyl-lysine modifications, however their role(s) in regulating cellular states and their potential as targets for the development of targeted treatment strategies is poorly understood. Here we present a set of 25 chemical probes, selective small molecule inhibitors, covering 29 human bromodomain targets. We comprehensively evaluate the selectivity of this probe-set using BROMOscan and demonstrate the utility of the set identifying roles of BRDs in cellular processes and potential translational applications. For instance, we discovered crosstalk between histone acetylation and the glycolytic pathway resulting in a vulnerability of breast cancer cell lines under conditions of glucose deprivation or GLUT1 inhibition to inhibition of BRPF2/3 BRDs. This chemical probe-set will serve as a resource for future applications in the discovery of new physiological roles of bromodomain proteins in normal and disease states, and as a toolset for bromodomain target validation.
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http://dx.doi.org/10.1038/s41467-019-09672-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6478789PMC
April 2019

The Mitochondrial Transacylase, Tafazzin, Regulates for AML Stemness by Modulating Intracellular Levels of Phospholipids.

Cell Stem Cell 2019 04 28;24(4):621-636.e16. Epub 2019 Mar 28.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada; Department of Medical Biophysics, University of Toronto, Toronto, ON, Canada.

Tafazzin (TAZ) is a mitochondrial transacylase that remodels the mitochondrial cardiolipin into its mature form. Through a CRISPR screen, we identified TAZ as necessary for the growth and viability of acute myeloid leukemia (AML) cells. Genetic inhibition of TAZ reduced stemness and increased differentiation of AML cells both in vitro and in vivo. In contrast, knockdown of TAZ did not impair normal hematopoiesis under basal conditions. Mechanistically, inhibition of TAZ decreased levels of cardiolipin but also altered global levels of intracellular phospholipids, including phosphatidylserine, which controlled AML stemness and differentiation by modulating toll-like receptor (TLR) signaling.
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http://dx.doi.org/10.1016/j.stem.2019.02.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7137093PMC
April 2019

Targeting bivalency de-represses Indian Hedgehog and inhibits self-renewal of colorectal cancer-initiating cells.

Nat Commun 2019 03 29;10(1):1436. Epub 2019 Mar 29.

Princess Margaret Cancer Centre, University Health Network, Toronto, ON, M5G1L7, Canada.

In embryonic stem cells, promoters of key lineage-specific differentiation genes are found in a bivalent state, having both activating H3K4me3 and repressive H3K27me3 histone marks, making them poised for transcription upon loss of H3K27me3. Whether cancer-initiating cells (C-ICs) have similar epigenetic mechanisms that prevent lineage commitment is unknown. Here we show that colorectal C-ICs (CC-ICs) are maintained in a stem-like state through a bivalent epigenetic mechanism. Disruption of the bivalent state through inhibition of the H3K27 methyltransferase EZH2, resulted in decreased self-renewal of patient-derived C-ICs. Epigenomic analyses revealed that the promoter of Indian Hedgehog (IHH), a canonical driver of normal colonocyte differentiation, exists in a bivalent chromatin state. Inhibition of EZH2 resulted in de-repression of IHH, decreased self-renewal, and increased sensitivity to chemotherapy in vivo. Our results reveal an epigenetic block to differentiation in CC-ICs and demonstrate the potential for epigenetic differentiation therapy of a solid tumour through EZH2 inhibition.
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http://dx.doi.org/10.1038/s41467-019-09309-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6441108PMC
March 2019

The Proteogenomic Landscape of Curable Prostate Cancer.

Cancer Cell 2019 03;35(3):414-427.e6

Department of Medical Biophysics, University of Toronto, Toronto, ON M5G 1L7, Canada; Ontario Institute for Cancer Research, Toronto, ON M5G 0A3, Canada; Department of Pharmacology and Toxicology, University of Toronto, Toronto, ON M5S 1A8, Canada; Department of Human Genetics, University of California, 12-109 CHS, 10833 Le Conte Avenue, Los Angeles, CA 90095, USA; Department of Urology, University of California, Los Angeles, CA 90024, USA; Jonsson Comprehensive Cancer Centre, University of California, Los Angeles, CA 90024, USA; Institute for Precision Health, University of California, Los Angeles, CA 90024, USA. Electronic address:

DNA sequencing has identified recurrent mutations that drive the aggressiveness of prostate cancers. Surprisingly, the influence of genomic, epigenomic, and transcriptomic dysregulation on the tumor proteome remains poorly understood. We profiled the genomes, epigenomes, transcriptomes, and proteomes of 76 localized, intermediate-risk prostate cancers. We discovered that the genomic subtypes of prostate cancer converge on five proteomic subtypes, with distinct clinical trajectories. ETS fusions, the most common alteration in prostate tumors, affect different genes and pathways in the proteome and transcriptome. Globally, mRNA abundance changes explain only ∼10% of protein abundance variability. As a result, prognostic biomarkers combining genomic or epigenomic features with proteomic ones significantly outperform biomarkers comprised of a single data type.
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http://dx.doi.org/10.1016/j.ccell.2019.02.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6511374PMC
March 2019

C3D: a tool to predict 3D genomic interactions between cis-regulatory elements.

Bioinformatics 2019 03;35(5):877-879

Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.

Motivation: The 3D genome architecture influences the regulation of genes by facilitating chromatin interactions between distal cis-regulatory elements and gene promoters. We implement Cross Cell-type Correlation based on DNA accessibility (C3D), a customizable computational tool that predicts chromatin interactions using an unsupervised algorithm that utilizes correlations in chromatin measurements, such as DNaseI hypersensitivity signals.

Results: C3D accurately predicts 32.7%, 18.3% and 24.1% of interactions, validated by ChIA-PET assays, between promoters and distal regions that overlie DNaseI hypersensitive sites in K562, MCF-7 and GM12878 cells, respectively.

Availability And Implementation: Source code is open-source and freely available on GitHub (https://github.com/LupienLabOrganization/C3D) under the GNU GPLv3 license. C3D is implemented in Bash and R; it runs on any platform with Bash (≥4.0), R (≥3.1.1) and BEDTools (≥2.19.0). It requires the following R packages: GenomicRanges, Sushi, data.table, preprocessCore and dynamicTreeCut.

Supplementary Information: Supplementary data are available at Bioinformatics online.
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http://dx.doi.org/10.1093/bioinformatics/bty717DOI Listing
March 2019

SMuRF: a novel tool to identify regulatory elements enriched for somatic point mutations.

BMC Bioinformatics 2018 Nov 26;19(1):454. Epub 2018 Nov 26.

Princess Margaret Cancer Centre, The MaRS Center, University Health Network, 101 College Street, Toronto, ON, M5G 1L7, Canada.

Background: Single Nucleotide Variants (SNVs), including somatic point mutations and Single Nucleotide Polymorphisms (SNPs), in noncoding cis-regulatory elements (CREs) can affect gene regulation and lead to disease development. Several approaches have been developed to identify highly mutated regions, but these do not take into account the specific genomic context, and thus likelihood of mutation, of CREs.

Results: Here, we present SMuRF (Significantly Mutated Region Finder), a user-friendly command-line tool to identify these significantly mutated regions from user-defined genomic intervals and SNVs. We demonstrate this using publicly available datasets in which SMuRF identifies 72 significantly mutated CREs in liver cancer, including known mutated gene promoters as well as previously unreported regions.

Conclusions: SMuRF is a helpful tool to allow the simple identification of significantly mutated regulatory elements. It is open-source and freely available on GitHub ( https://github.com/LupienLab/SMURF ).
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http://dx.doi.org/10.1186/s12859-018-2501-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6258448PMC
November 2018
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