The clear cell sarcoma functional genomic landscape
Emanuele Panza, Benjamin B. Ozenberger, Krystal M. Straessler, Jared J. Barrott, Li Li, Yanliang Wang, Mingchao Xie, Anne Boulet, Simon W.A. Titen, Clinton C. Mason, Alexander J. Lazar, Li Ding, Mario R. Capecchi, Kevin B. Jones
Emanuele Panza, Benjamin B. Ozenberger, Krystal M. Straessler, Jared J. Barrott, Li Li, Yanliang Wang, Mingchao Xie, Anne Boulet, Simon W.A. Titen, Clinton C. Mason, Alexander J. Lazar, Li Ding, Mario R. Capecchi, Kevin B. Jones
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Research Article Genetics Oncology

The clear cell sarcoma functional genomic landscape

Abstract

Clear cell sarcoma (CCS) is a deadly malignancy affecting adolescents and young adults. It is characterized by reciprocal translocations resulting in expression of the chimeric EWSR1-ATF1 or EWSR1-CREB1 fusion proteins, driving sarcomagenesis. Besides these characteristics, CCS has remained genomically uncharacterized. Copy number analysis of human CCSs showed frequent amplifications of the MITF locus and chromosomes 7 and 8. Few alterations were shared with Ewing sarcoma or desmoplastic, small round cell tumors, which are other EWSR1-rearranged tumors. Exome sequencing in mouse tumors generated by expression of EWSR1-ATF1 from the Rosa26 locus demonstrated no other repeated pathogenic variants. Additionally, we generated a new CCS mouse by Cre-loxP–induced chromosomal translocation between Ewsr1 and Atf1, resulting in copy number loss of chromosome 6 and chromosome 15 instability, including amplification of a portion syntenic to human chromosome 8, surrounding Myc. Additional experiments in the Rosa26 conditional model demonstrated that Mitf or Myc can contribute to sarcomagenesis. Copy number observations in human tumors and genetic experiments in mice rendered, for the first time to our knowledge, a functional landscape of the CCS genome. These data advance efforts to understand the biology of CCS using innovative models that will eventually allow us to validate preclinical therapies necessary to achieve longer and better survival for young patients with this disease.

Authors

Emanuele Panza, Benjamin B. Ozenberger, Krystal M. Straessler, Jared J. Barrott, Li Li, Yanliang Wang, Mingchao Xie, Anne Boulet, Simon W.A. Titen, Clinton C. Mason, Alexander J. Lazar, Li Ding, Mario R. Capecchi, Kevin B. Jones

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Figure 7

MYC stabilization enhances EWSR1-ATF1–induced sarcomagenesis but alters tumor phenotypes.

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MYC stabilization enhances EWSR1-ATF1–induced sarcomagenesis but alters...
(A) BioDiscovery data on CNAs in regions of human chromosome 8 that are syntenic to amplified regions of mouse chromosome 15 and that flank MYC. (B) CNAs in regions of mouse chromosome 15 syntenic to human chromosome 8 and surrounding Myc. (C) Schematic showing the floxed stop cassette that allows for Cre-inducible expression of MycT58A at the Igs2 locus. (D) Breeding strategy to generate mice that express both EWSR1-ATF1 (EA1) and MycT58A upon TATCre injection, as well as Igs2WT/WT littermate controls. (E) Tumor growth curves for Rosa26EA1/WT mice with either Igs2LSL-Myc/WT (red) or Igs2WT/WT (black), following TATCre injection at 28 days of age. (F) Graph of the blinded quantitation of H&E-stained slides for histologic features distinguishing tumors expressing EA1 alone and tumors expressing EA1 plus MycT58A (EA1+Myc, red), with, for reference, the prevalence of the nested morphology observed in human CCSs on a tissue microarray (n = 20). (G) Photomicrographs of H&E-stained tissue sections demonstrating the 2 variants on myxoid features, 1 in each genotype, as well as the nested histomorphology observed focally in 7 human tumors (n = 20) and all Myc-activating mouse tumors (each photomicrograph is a 100 μm square obtained with a 60× original magnification objective lens).