title: DNA methylation heterogeneity defines a disease spectrum in Ewing sarcoma.
abstract: Developmental tumors in children and young adults carry few genetic alterations, yet they have diverse clinical presentation. Focusing on Ewing sarcoma, we sought to establish the prevalence and characteristics of epigenetic heterogeneity in genetically homogeneous cancers. We performed genome-scale DNA methylation sequencing for a large cohort of Ewing sarcoma tumors and analyzed epigenetic heterogeneity on three levels: between cancers, between tumors, and within tumors. We observed consistent DNA hypomethylation at enhancers regulated by the disease-defining EWS-FLI1 fusion protein, thus establishing epigenomic enhancer reprogramming as a ubiquitous and characteristic feature of Ewing sarcoma. DNA methylation differences between tumors identified a continuous disease spectrum underlying Ewing sarcoma, which reflected the strength of an EWS-FLI1 regulatory signature and a continuum between mesenchymal and stem cell signatures. There was substantial epigenetic heterogeneity within tumors, particularly in patients with metastatic disease. In summary, our study provides a comprehensive assessment of epigenetic heterogeneity in Ewing sarcoma and thereby highlights the importance of considering nongenetic aspects of tumor heterogeneity in the context of cancer biology and personalized medicine.
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.
authors.forename: Nathan C
authors.initials: NC
authors.lastname: Sheffield
authors.affiliation: Institut Curie, PSL Research University, Service de Genetique, Pole de Medecine Diagnostique et Theranostique, Unité de Génétique Somatique, Paris, France.
authors.forename: Gaelle
authors.initials: G
authors.lastname: Pierron
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.
authors.forename: Johanna
authors.initials: J
authors.lastname: Klughammer
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.
authors.forename: Paul
authors.initials: P
authors.lastname: Datlinger
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.
authors.forename: Andreas
authors.initials: A
authors.lastname: Schönegger
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.
authors.forename: Michael
authors.initials: M
authors.lastname: Schuster
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.
authors.forename: Johanna
authors.initials: J
authors.lastname: Hadler
authors.affiliation: Institut Curie, PSL Research University, INSERM, U830, Paris, France.
authors.forename: Didier
authors.initials: D
authors.lastname: Surdez
authors.affiliation: Institut Curie, PSL Research University, Service de Genetique, Pole de Medecine Diagnostique et Theranostique, Unité de Génétique Somatique, Paris, France.
authors.forename: Delphine
authors.initials: D
authors.lastname: Guillemot
authors.affiliation: Institut Curie, PSL Research University, Service de Genetique, Pole de Medecine Diagnostique et Theranostique, Unité de Génétique Somatique, Paris, France.
authors.forename: Eve
authors.initials: E
authors.lastname: Lapouble
authors.affiliation: Institut Curie, PSL Research University, Service de Pathologie, Pole de Medecine Diagnostique et Theranostique, Paris, France.
authors.forename: Paul
authors.initials: P
authors.lastname: Freneaux
authors.affiliation: Service d'Anatomie et de Cytologie Pathologiques, Hopitaux de Brabois, Hopital d'Adultes, Nancy, France.
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Champigneulle
authors.affiliation: Centre de Pathologie du Pôle Est, Hopitaux de Lyon, Lyon, France.
authors.forename: Raymonde
authors.initials: R
authors.lastname: Bouvier
authors.affiliation: Children's Cancer Research Institute, St. Anna Kinderkrebsforschung, Vienna, Austria.
authors.forename: Diana
authors.initials: D
authors.lastname: Walder
authors.affiliation: Children's Cancer Research Institute, St. Anna Kinderkrebsforschung, Vienna, Austria.
authors.forename: Ingeborg M
authors.initials: IM
authors.lastname: Ambros
authors.affiliation: St. Anna Children's Hospital, St. Anna Kinderspital, Vienna, Austria.|Department of Pediatrics, Medical University of Vienna, Vienna, Austria.
authors.forename: Caroline
authors.initials: C
authors.lastname: Hutter
authors.affiliation: Children's Cancer Research Institute, St. Anna Kinderkrebsforschung, Vienna, Austria.
authors.forename: Eva
authors.initials: E
authors.lastname: Sorz
authors.affiliation: Department of Pathology, Institute of Biomedicine of Sevilla (IBiS), Virgen del Rocio University Hospital/CSIC/University of Sevilla, Seville, Spain.
authors.forename: Ana T
authors.initials: AT
authors.lastname: Amaral
authors.affiliation: Department of Pathology, Institute of Biomedicine of Sevilla (IBiS), Virgen del Rocio University Hospital/CSIC/University of Sevilla, Seville, Spain.
authors.forename: Enrique
authors.initials: E
authors.lastname: de Álava
authors.affiliation: Spinal Cord Injury and Tissue Regeneration Center Salzburg, Paracelsus Medical University, Salzburg, Austria.|Department of Blood Group Serology and Transfusion Medicine, Paracelsus Medical University, Salzburg, Austria.
authors.forename: Katharina
authors.initials: K
authors.lastname: Schallmoser
authors.affiliation: Spinal Cord Injury and Tissue Regeneration Center Salzburg, Paracelsus Medical University, Salzburg, Austria.|Institute for Experimental and Clinical Cell Therapy, Paracelsus Medical University, Salzburg, Austria.
authors.forename: Dirk
authors.initials: D
authors.lastname: Strunk
authors.affiliation: Division of Biomedical Research, Medical University of Graz, Graz, Austria.
authors.forename: Beate
authors.initials: B
authors.lastname: Rinner
authors.affiliation: Institute of Pathology, Medical University of Graz, Graz, Austria.
authors.forename: Bernadette
authors.initials: B
authors.lastname: Liegl-Atzwanger
authors.affiliation: Organizational Unit of Research Infrastructure, Biobank Graz, Medical University of Graz, Graz, Austria.
authors.forename: Berthold
authors.initials: B
authors.lastname: Huppertz
authors.affiliation: Department of Orthopedic Surgery, Medical University of Graz, Graz, Austria.
authors.forename: Andreas
authors.initials: A
authors.lastname: Leithner
authors.affiliation: Service d'Anatomie et de Cytologie Pathologiques, Hôpital Universitaire Trousseau, Tours, France.
authors.forename: Gonzague
authors.initials: G
authors.lastname: de Pinieux
authors.affiliation: Gustave Roussy, Département de Pathologie, Villejuif, France.
authors.forename: Philippe
authors.initials: P
authors.lastname: Terrier
authors.affiliation: Institut Curie, PSL Research University, Service de Genetique, Pole de Medecine Diagnostique et Theranostique, Unité de Génétique Somatique, Paris, France.|Institut Curie, PSL Research University, Departement d'Oncologie Pédiatrique Adolescent Jeunes Adultes, Paris, France.
authors.forename: Valérie
authors.initials: V
authors.lastname: Laurence
authors.affiliation: Institut Curie, PSL Research University, Departement d'Oncologie Pédiatrique Adolescent Jeunes Adultes, Paris, France.
authors.forename: Jean
authors.initials: J
authors.lastname: Michon
authors.affiliation: Children's Cancer Research Institute, St. Anna Kinderkrebsforschung, Vienna, Austria.|St. Anna Children's Hospital, St. Anna Kinderspital, Vienna, Austria.|Department of Pediatrics, Medical University of Vienna, Vienna, Austria.
authors.forename: Ruth
authors.initials: R
authors.lastname: Ladenstein
authors.affiliation: Children's Cancer Research Institute, St. Anna Kinderkrebsforschung, Vienna, Austria.|St. Anna Children's Hospital, St. Anna Kinderspital, Vienna, Austria.|Department of Pediatrics, Medical University of Vienna, Vienna, Austria.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Holter
authors.affiliation: Department of Orthopedics, Vienna General Hospital, Medical University of Vienna, Vienna, Austria.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Windhager
authors.affiliation: University Hospital Münster, Department of Pediatrics and Pediatric Hematology and Oncology, Münster, Germany.
authors.forename: Uta
authors.initials: U
authors.lastname: Dirksen
authors.affiliation: Children's Cancer Research Institute, St. Anna Kinderkrebsforschung, Vienna, Austria.|Department of Pediatrics, Medical University of Vienna, Vienna, Austria.
authors.forename: Peter F
authors.initials: PF
authors.lastname: Ambros
authors.affiliation: Institut Curie, PSL Research University, Service de Genetique, Pole de Medecine Diagnostique et Theranostique, Unité de Génétique Somatique, Paris, France.|Institut Curie, PSL Research University, INSERM, U830, Paris, France.
authors.forename: Olivier
authors.initials: O
authors.lastname: Delattre
authors.affiliation: Children's Cancer Research Institute, St. Anna Kinderkrebsforschung, Vienna, Austria.|Department of Pediatrics, Medical University of Vienna, Vienna, Austria.
authors.forename: Heinrich
authors.initials: H
authors.lastname: Kovar
authors.affiliation: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.|Department of Laboratory Medicine, Medical University of Vienna, Vienna, Austria.|Max Planck Institute for Informatics, Saarland Informatics Campus, Saarbrücken, Germany.|Ludwig Boltzmann Institute for Rare and Undiagnosed Diseases, Vienna, Austria.
authors.forename: Christoph
authors.identifier: http://orcid.org/0000-0001-6091-3088
authors.initials: C
authors.lastname: Bock
authors.affiliation: Children's Cancer Research Institute, St. Anna Kinderkrebsforschung, Vienna, Austria.
authors.forename: Eleni M
authors.initials: EM
authors.lastname: Tomazou
mesh_terms: D000293:Adolescent; D000328:Adult; D001859:Bone Neoplasms / Q000235:genetics*; D045744:Cell Line, Tumor; D002648:Child; D002675:Child, Preschool; D019175:DNA Methylation / Q000235:genetics*; D044127:Epigenesis, Genetic; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D018740:Genetic Heterogeneity; D006801:Humans; D008297:Male; D008875:Middle Aged; D015514:Oncogene Proteins, Fusion / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D051789:Proto-Oncogene Protein c-fli-1 / Q000235:genetics*; D034802:RNA-Binding Protein EWS / Q000235:genetics*; D012512:Sarcoma, Ewing / Q000235:genetics*; D055815:Young Adult
chemical_list: C106031:EWS-FLI fusion protein; D015514:Oncogene Proteins, Fusion; D051789:Proto-Oncogene Protein c-fli-1; D034802:RNA-Binding Protein EWS