Expanding the spectrum of translocations in sclerosing epitheloid fibrosarcoma: A new case with EWSR1‐CREB3L3 fusion

To the Editor: Dewaele et al. recently reported in this journal a novel EWSR1-CREB3L3 gene fusion in a mesenteric sclerosing epithelioid fibrosarcoma (SEF). The authors performed fluorescence in situ hybridization (FISH), array comparative hybridization (aCGH), reverse transcriptasePCR, and Sanger sequencing to analyze an intraabdominal SEF in an 18-year-old male. Moreover, their FISH and aCGH findings were consistent with unbalanced EWSR1 gene rearrangement. The authors concluded that it would also be worthwhile to examine if CREB3L3 or other members of the CREB3 family of transcription factors are involved in SEF, as well as in the biologically related entity low-grade fibromyxoid sarcoma (LGFMS), given that translocation partners had not been explored in the previously reported EWSR1-rearranged SEFs. Herein, we describe a case of “pure” SEF, presenting as a pelvic tumor in which EWSR1-CREB3L3 fusion was detected using our nextgeneration sequencing assay, OncoPanel, which was developed at the Center for Advanced Molecular Diagnostics at Brigham and Women's Hospital and the Center for Cancer Genome Discovery at Dana Farber Cancer Institute, Boston, Massachusetts, and is performed in a Clinical Laboratory Improvement Amendments-certified laboratory to detect somatic mutations, copy-number variations, and structural variants in all new cancers samples arriving at our institution. The patient was a 40-year-old woman who presented with a pelvic mass measuring 22.0 cm that invaded into the bladder, myometrium, cervix, and right adnexa. Microscopically, the tumor showed an infiltrative growth of epithelioid and round cells arranged in cords, nests, and sheets, within a densely sclerotic stroma (Figure 1A). The tumor cells had hyperchromatic nuclei and small-to-inconspicuous nucleoli and varying amounts of cytoplasm. Mitoses numbered 13 per 10 high power fields (2 mm), and necrosis was present. By immunohistochemistry, tumor cells expressed MUC4 (Figure 1B). Other immunohistochemical results included positivity for HMGA2, TLE1, focal/ weak expression of S-100, inhibin, and IFITM1, and negativity for SOX10, GFAP, keratin, EMA, synaptophysin, SALL4, cyclin D1, Claudin-4, calretinin, OCT3/4, CD138, CD10, SMA, desmin, and STAT6. The morphologic and immunohistochemical features, in conjunction with the molecular genetic results (see below), supported the diagnosis of SEF. DNA was extracted from formalin-fixed paraffin-embedded tissue. Targeted next-generation sequencing was performed as previously described. Sequencing identified a rearrangement with breakpoints involving EWSR1 intron 8 and CREB3L3 intron 3 (Figure 1C). The directionality of the observed sequences is consistent with a gene fusion with EWSR1 as the 50 partner and CREB3L3 as the 30 partner. In addition, copy-number analysis of chromosome 22 showed copy imbalances involving the EWSR1 gene, with relative gain of EWSR1 exons 1-8, and relative loss of EWSR1 exons 9-18 (Figure 1D). The overall sequencing findings are consistent with an unbalanced EWSR1-CREB3L3 rearrangement. Chromosome arm 19p also showed relative copy-number loss (data not shown). On the basis of the sequencing findings, FISH analysis for EWSR1 was performed, and EWSR1 rearrangement observed in 47 of 50 nuclei (94%). However, all abnormal nuclei contained 2 copies of isolated 5’ EWSR1 red signal and zero copies of isolated 3’ EWSR1 green signal, consistent with an unbalanced rearrangement (Figure 1E). The unbalanced nature of the EWSR1 rearrangement was consistent with the gain and loss findings on chromosome 22 at the EWSR1 locus detected by our Oncopanel (eg, gain of exons 1-8 and loss of exon 9-18 of the EWSR1 gene). Our reported case is the second example of “pure” SEF with EWSR1-CREB3L3 gene fusion. The genetic features of SEF have been better characterized over recent years, with EWSR1-CREB3L1 and EWSR1-CREB3L2 present in most cases of “pure” SEF showing MUC4 expression, and FUS-CREB3L2 fusions in most hybrid LGFMS-SEF tumors and only rarely in “pure” SEFs. Recently, Arbajian et al. reported recurrent aberrations in SEF and hybrid LGFMS-SEF, including DMD microdeletions, 11p deletions, and CD24 upregulation, as well as single examples of PAX5-CREB3L1 and FUS-CREM fusions in pure SEFs. The authors suggested that these additional genomic imbalances may drive the more aggressive behavior of SEF and LGFMS-SEF, in comparison to the more prolonged course that LGFMS follows. The identification of CREB3L3 as a translocation partner in SEF is not entirely surprising given the homology between members of the CREB3 transcription factor family, and it is likely that the oncogenic roles of EWSR1-CREB3L1/2/3 are similar in SEF. It remains Received: 12 July 2018 Revised: 16 August 2018 Accepted: 17 August 2018 DOI: 10.1002/gcc.22677