Original ArticlesFUS-NFATC2 or EWSR1-NFATC2 Fusions Are Present in a Large Proportion of Simple Bone CystsPižem, Jože MD, PhD*; Šekoranja, Daja MD*; Zupan, Andrej PhD*; Boštjančič, Emanuela PhD*; Matjašič, Alenka PhD*; Mavčič, Blaž MD, PhD†,‡; Contreras, Juan A. PhD§; Gazič, Barbara MD, PhD§; Martinčič, David MD, PhD†,‡; Snoj, Žiga MD, PhD‡,∥; Limpel Novak, Katarina A. MD∥; Salapura, Vladka MD, PhD‡,∥Author Information *Institute of Pathology, Faculty of Medicine ‡Faculty of Medicine, University of Ljubljana †Department of Orthopedic Surgery ∥Institute of Radiology, University Medical Center Ljubljana §Department of Pathology, Institute of Oncology, Ljubljana, Slovenia Conflicts of Interest and Source of Funding: Supported by the Slovenian Research Agency (research core funding no. P3-0054). The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article. Correspondence: Jože Pižem, MD, PhD, Institute of Pathology, Faculty of Medicine, University of Ljubljana, Korytkova 2, Ljubljana 1000, Slovenia (e-mail: [email protected]). The American Journal of Surgical Pathology: December 2020 - Volume 44 - Issue 12 - p 1623-1634 doi: 10.1097/PAS.0000000000001584 Buy Metrics Abstract A simple bone cyst (SBC) is a benign bone lesion of unknown etiology. It can be differentiated from an aneurysmal bone cyst (ABC) by radiologic and histopathologic features, as well as by the absence of fusions of the USP6 gene characteristic of an ABC. In an attempt to differentiate between ABC and SBC in a recurrent bone cyst, we performed targeted RNA sequencing and found an EWSR1-NFATC2 fusion and no fusion of the USP6 gene. We subsequently analyzed additional 10 cysts, consistent with SBCs after radiologic-pathologic correlation, for the presence of an NFATC2 gene fusion, by targeted RNA sequencing, reverse-transcription polymerase chain reaction (RT-PCR) and Sanger sequencing, and fluorescent in situ hybridization. Targeted RNA sequencing showed a FUS-NFATC2 fusion in 4 of 11 SBCs and an EWSR1-NFATC2 fusion in 2 of 11 SBCs. No fusion was identified in 3 SBCs and the analysis was not successful in 2 SBCs because of the low quantity or poor quality of isolated RNA. All the 6 fusions detected by targeted RNA sequencing were confirmed by RT-PCR and Sanger sequencing, and 5 of the 6 fusions by fluorescent in situ hybridization. An additional FUS-NFATC2 fusion was identified by RT-PCR, Sanger sequencing, and fluorescent in situ hybridization in 1 of the 3 cases negative for fusions by targeted RNA sequencing. At least a large subset of SBCs represents cystic neoplasms characterized by FUS-NFATC2 or EWSR1-NFATC2 fusions, which also define a group of distinct, rare “Ewing-like” sarcomas that predominantly arise in long bones. Our results provide additional evidence of the existence of benign lesions with FUS-NFATC2 or EWSR1-NFATC2 fusions. Although they can recur locally in a nondestructive manner, their clinical course and possible relation to sarcoma with EWSR1-NFATC2 or FUS-NFATC2 fusion remains to be elucidated. Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.