Molecular pathogenesis and prognostication of "low‐grade'' and "high‐grade" endometrial stromal sarcoma

Abstract Endometrial stromal sarcomas (ESS) are a heterogeneous group of rare mesenchymal cancers. Considerable knowledge has been gained in recent years about the molecular characteristics of these cancers, which helps to classify them in a more meaningful manner leading to improved diagnosis, prognostication, and treatment. According to this classification, ESS is now grouped as low‐ or high‐grade. ESS may have overlapping clinical presentation, morphology, and immunohistochemical profile. Their genetic characteristics allow subdivision of many of them depending on which pathogenetically important fusion genes they carry, but clearly much more needs to be unraveled in this regard. We here provide an overview of the molecular pathogenetic knowledge gained so far on low‐ and high‐grade ESS.

balanced or unbalanced, the molecular product of such rearrangements is usually a so-called fusion gene. This is a hybrid formed from two previously independent genes. It has been known for more than 30 years that gene fusions play an important role in tumorigenesis. 4,5 Oncogenic fusions may lead to an abnormal gene product brought about by fusing elements from the two fusion partners. Alternatively, a proto-oncogene may be fused to a strong promoter leading to its upregulation. Oncogenic fusion transcripts may also be caused by trans-splicing or read-through events. Identification of an activated fusion gene improves diagnostic precision as well as prognostication, while at the same time providing pathogenetic information about the tumor. 6 Though different fusion genes have been identified in ESS, generally it seems that the presence of one fusion gene excludes the presence of another in the same tumor.

| LG-ESS
Since 1988, when the first cytogenetic abnormalities were reported in ESS, 7 8 The aberration leads at the molecular level to fusion of two zinc finger genes, JAZF1 (from 7p15) and SUZ12 (previously known as JJAZ1; from 17q21; Figure 1). 9 Many other chromosomal changes have also been described and their pattern of occurrence is clearly nonrandom ( Table 1). The molecular product behind each translocation has been identified for most of them. Ever improving methodological tools have facilitated the discoveries, especially the introduction of deep sequencing technologies allowing rapid screening of tumor genomes and transcriptomes. 10 The second most common rearrangement involves chromosome band 6p21 and the PHF1 gene. 11 PHF1 may recombine with several partners, not least JAZF1 trough an unbalanced 6;7-translocation. 11 Other partners are EPC1 through a 6;10-rearrangement 11 ; MEAF6 through a t(1;6)(p34;p21) 12 ; BRD8 via t(5;6)(q31;p21) 13 ; EPC2 through a 2;6-rearrangement 14 ; and recently a MBTD1/PHF1 was also reported. 15 A study by D'Angelo et al. 16 showed that tumors bearing PHF1 fusions, independently of which partner gene is involved, typically present sex cord-like differentiation, leading the authors to suggest that rearrangements of this gene preferentially induce such differentiation.
A less frequent chromosomal rearrangement is the t(X;17)(p11; q21) leading to the MBTD1/EZHIP (previously known as CXorf67) fusion. 17 Variants of the JAZF1/SUZ12 were recently identified in which JAZF1 recombines with BCORL1 18 and SUZ12 with MEAF6. 19 Another two novel chimeric fusions were reported by Dickson et al., 20 EPC1/SUZ12 and EPC1/BCOR. The identification of these transcripts underlines the promiscuous nature of EPC1, but also obfuscates the molecular distinction between high grade and low grade ESS. Both tumors were described as clinically aggressive and with morphological features compatible with HG-ESS. 20 The biological potential associated with these fusions remains to be fully characterized. Most likely, also other fusion gene products will emerge. Until a sufficient number of cases is studied and the clinical parameters correlated, it is likely to remain challenging to classify the observed molecular events as being fully specific for LG-or HG-ESS.
The above-mentioned fusion genes have so far not been seen in leiomyomas, leiomyosarcomas, and uterine tumor resembling ovarian sex cord stromal tumors (UTROSCT), all of which may on occasion be differential diagnoses. Nevertheless, none of the fusions is fully pathognomonic for LG-ESS as they have all been found also in other neoplasias. JAZF1/SUZ12 fusion is detected in 65% to 75% of ESN. 9,[21][22][23][24][25][26][27] This chimera has been found more frequently in classic LG-ESS than in LG-ESS exhibiting variant features. 28 Recently, an ESN with MEAF6/PHF1 was reported, providing further support for a continuum between these two tumor entities. 29 Interestingly, the ESN F I G U R E 1 Partial karyogram and chromatogram of the hallmarks for ESS. A, LG-ESS: partial karyogram showing the t(7;17)(p15;q11) (left), the ideograms for the rearranged chomosomes (center), and sequence chromatogram for the JAZF1/SUZ12 fusion gene (right). B, HG-ESS: partial karyogram showing the t(10;17)(q22;p13) (left), the ideograms for the rearranged chomosomes (center), and the sequence chromatogram for the YWHAE/NUTM2A/B fusion gene (right). Arrows point at breakpoints showed focal peripheral ossification, a rare feature of ESN and/or LG-ESS but a hallmark of ossifying fibromyxoid tumors with which they may share also other molecular events such as PHF1 fusions, that is, EP400/PHF1, MEAF6/PHF1, and EPC1/PHF1. It was therefore hypothesized that the MEAF6/PHF1 could be associated with metaplastic bone formation. 30 The aforementioned fusions occur in ossifying fibromyxoid tumors of soft parts, irrespective of whether the tumor is diagnosed as typical, atypical, or malignant, whereas JAZF1/PHF1 has been found in cardiac ossifying sarcomas. [31][32][33][34] Furthermore, two novel fusions, CREBBP/BCORL1 and KDM2A/WWTR1 have been reported in ossifying fibromyxoid tumors 35 showing additional overlap with LG-ESS as the genes CREBBP and KDM2B were previously found in a chimera in the latter tumor as well. 36 Recently, the JAZF1/BCORL1 fusion was identified in an adenosarcoma arising in the uterus. 37

| HG-ESS
The cytogenetic hallmark of HG-ESS is the balanced t(10;17)(q22;p13) translocation simultaneously reported in 2003 by two groups 38,39 ( Figure 1). The gene product it leads to was identified by Lee et al. 40 as an in-frame fusion between the YWHAE and NUTM2A/B genes (previously known as FAM22A/B; Figure 1). The fusion seems to be specific for HG-ESS as it was never identified in other gynecological tumors or neoplastic lesions, such as, uterine adenosarcomas, carcinosarcomas, leiomyosarcomas, leiomyomas, and polypoid endometriosis. 40 Kubo et al. 41 reported a low frequency of YWHAE and NUTM2A/B rearrangements in epithelioid leiomyosarcoma; admittedly, the immunostaining data of that study were suggestive of an unusual ESS. Splitting of probes for the YWHAE, FAM22A, and FAM22B genes has been reported in a uterine angiosarcoma. 42 Despite the fact that no fusion transcript involving the mentioned genes was discovered, the authors suggested that abnormalities of them may contribute to development of uterine angiosarcoma in much the same manner as they do in ESS. 42 Of further note in the context is the fact that no such rearrangement was identified in 21 angiosarcomas of extrauterine soft tissue. 40 However, the very same chromosomal translocation has been reported in clear cell sarcomas of the kidney by different groups [43][44][45] and shown to correspond to a YWHAE/NUTM2A/B fusion. 46 Kao et al. 47 identified it also in small round blue cell sarcomas of soft tissue, undifferentiated round cell sarcoma, and primitive myxoid mesenchymal tumor of infancy. 47,48 Recently, the first neonatal case of a round cell sarcoma bearing this chimera was described in a tumor with aggressive clinical behavior. 49 Sciallis et al. 50   BCOR-positive HG-ESS was identified at an early stage when an endocervical polypoid mass from the lower uterine segment was examined. 56  BCOR immunochemical staining has proved to be a highly sensi-  Lately, methylation profiles for different uterine tumors have been determined 91 showing different methylation clusters correlating with established diagnostic entities. The data obtained highlighted that the LG-ESS pattern differed from that of HG-ESS, and that, within the latter, distinct subgrouping of YWHAE-and BCOR-rearranged tumors was possible. 91 The copy number-profile was investigated by the same group in a series of uterine tumors that included LG-ESS, HG-ESS, UTROSCT, uterine leiomyomas, and uterine leiomyosarcomas. 69 The authors identified amplification of the MDM2 gene from chromosomal band 12q15 only in BCOR-rearranged HG-ESS. 69  Histone acetyltransferases (HAT) of the MYST family are known to be involved in vital cellular processes, such as, gene transcription, detection and repair of DNA damage, and DNA replication. They carry out a significant proportion of all nuclear acetylation, and their anomalous activity, or anomalous activity of complexes associated with them (these enzymes work in multisubunit protein complexes), can lead to different anomalies from cell death to uncontrolled growth, the latter leading to cancer formation. 93 There are different HATs in the MYST family, many of which are known to be involved in different types of cancer, for example, MOZ and MORF in acute myeloid leukemia.
Of all chimeric proteins associated with ESS, YWHAE/ NUTM2A/B is the only one that does not undergo epigenetic modification. The gene for YWHAE (14-3-3 ε) belongs to a broad family of proteins responsible for mediating signal transduction. 40 FAM22A/B was renamed NUTM2A/B due to its sequence homology with NUT (NUTM1), which is notable in NUT midline carcinoma. 94 The issue whether a linear tumor progression exists among the different EST was investigated by means of array based Comparative Genomic Hybridization (aCGH). 95 The fact that no chromosomal aberrations were common to the ESN, LG-ESS, and UES/UUS investigated led the authors to conclude that this proposition was unlikely. However, an increasing number of aberrations were registered from ESN to UES, correlating well with histological grading and worsening clinical behavior. 95

ACKNOWLEDGMENTS
The authors wish to thank the Norwegian Radium Hospital Foundation and the Anders Jahre's foundation through UNIFOR (University of Oslo) for their support.

DATA AVAILABILITY STATEMENT
Data sharing is not applicable to this article as no new data were created or analyzed in this study.