Update on Endometrial Stromal Tumours of the Uterus

Endometrial stromal tumours (ESTs) are rare, intriguing uterine mesenchymal neoplasms with variegated histopathological, immunohistochemical and molecular characteristics. Morphologically, ESTs resemble endometrial stromal cells in the proliferative phase of the menstrual cycle. In 1966 Norris and Taylor classified ESTs into benign and malignant categories according to the mitotic count. In the most recent classification by the WHO (2020), ESTs have been divided into four categories: Endometrial Stromal Nodules (ESNs), Low-Grade Endometrial Stromal Sarcomas (LG-ESSs), High-Grade Endometrial Stromal Sarcomas (HG-ESSs) and Undifferentiated Uterine Sarcomas (UUSs). ESNs are clinically benign. LG-ESSs are tumours of low malignant potential, often with indolent clinical behaviour, with some cases presented with a late recurrence after hysterectomy. HG-ESSs are tumours of high malignant potential with more aggressive clinical outcome. UUSs show high-grade morphological features with very aggressive clinical behavior. With the advent of molecular techniques, the morphological classification of ESTs can be integrated with molecular findings in enhanced classification of these tumours. In the future, the morphological and immunohistochemical features correlated with molecular categorisation of ESTs, will become a robust means to plan therapeutic decisions, especially in recurrences and metastatic disease. In this review, we summarise the morphological, immunohistochemical and molecular features of ESTs with particular reference to the most recent molecular findings.


Introduction
Endometrial stromal tumours (ESTs) are a rare, fascinating and complex subset of mesenchymal uterine neoplasms with heterogeneous morphological, immunohistochemical and genetic features. ESTs constitute ∼10% of uterine mesenchymal tumours [1].
Approximately 50% of endometrial stromal sarcomas (ESSs) occur in premenopausal women and the majority is detected at stage I of the International Federation of Gynecology and Obstetrics (FIGO) [2].
Morphologically, ESTs resemble endometrial stromal cells in the proliferative phase of the menstrual cycle. In 1966 Norris and Taylor attempted to classify ESTs in their seminal manuscript [3].
They divided the lesions into two groups; the first group with pushing margins was labelled as stromal nodules and the second group with infiltrating margins was defined as endolymphatic stromal myosis or stromal sarcoma according to the mitotic index: lesions with ≤ 10 mitoses per 10 HPF (high-power field) were classified as endolymphatic stromal myosis and neoplasms with ≥ 10 mitoses per 10 HPF were categorised as stromal sarcomas. In view of the clinical outcome (100% survival rate within five years), The aim of this review is to shed some light on the complex classification of ESTs, highlighting the varied histopathological, immunohistochemical and molecular features for each sub-group.

Endometrial Stromal Nodule (ESN) and Low-Grade Endometrial Stromal Sarcoma (LG-ESS)
ESN is a benign, whereas LG-ESS is a malignant neoplasm of the uterus (affecting the body of the uterus more than the cervix) and extra-uterine sites [8,9]. The mean age for LG-ESS is 52 years, ranging between 16 and 83 years [10]. The risk factors are pelvic radiation and prolonged use of tamoxifen or oestrogen. The most common findings are abnormal uterine bleeding and pelvic pain [11][12][13]. Other symptoms of patients with LG-ESS are uterine mass and metastases to the adnexae, lymph nodes and lungs [14].

ESN Morphology
ESNs are a proliferation of bland, uniform cells with oval nuclei and scanty cytoplasm, resembling endometrial stromal cells in the proliferative phase of the menstrual cycle ( Figure 1A). ESNs exhibit prominent arterioles and well-circumscribed expansive (noninfiltrative) margins ( Figure 1B). It may occasionally present with infiltrative margins, however these should be 3 mm or less in maximum dimension and not exceed three foci [15,16]. The mitotic rate is not high (less than 10 × 10 HPF). Areas of coagulative necrosis and sex-cord-like differentiation may be identified, but, by definition, lympho-vascular invasion (LVIS) is not present. Areas of smooth muscle metaplasia may be present and these should not mislead to an incorrect diagnosis of myometrial invasion. The differential diagnosis includes cellular leiomyoma and LG-ESS. The vascular pattern of ESN, composed of typical arterioles, is not a prominent feature of cellular leiomyoma. The presence of large blood vessels, one of the characteristics of cellular leiomyoma, may also be detected in ESN, but is not as conspicuous as in cellular leiomyoma. In addition, ESN usually does not contain the clefts frequently seen in cellular leiomyoma [17].
ESNs can be differentiated from LG-ESS exclusively by the presence of pushing margins and lack of LVIS [3,15,16], therefore, the definitive diagnosis of ESN can be rendered on resection specimens only and cannot be confidently established on biopsy or tissue removal systems (i.e., MyoSure resections). Nevertheless, even in resection specimens, the differential diagnosis between ESNs and LG-ESS may not be straightforward and extensive sampling may be required.

LG-ESS Morphology
LG-ESS shows the same histopathological features as ESN, except, as has already been mentioned, for the presence of infiltrative/permeative, tongue-like margins ( Figure  2A) and LVIS that may also be observed in the parametrial veins [15].
From the purely morphological point of view, some low-grade ESTs show "hybrid" features between ESN and LG-ESS. These lesions have been labelled as ESTs with limited infiltration (EST-LI). The extent of myometrial invasion in this entity is less than in LG-ESS but more than in ESN (occasional finger-like projections into the myometrium of up to 3 mm are allowable) [17]. Obviously, the diagnosis of this lesion is highly subjective and depends on the extent of the sampling; therefore, the diagnostic reproduceable value is limited. In addition, the clinical outcome among patients with this sub-category of morphologically low-grade EST is not clear [17,18] and a small percentage of EST-LI can show malignant behaviour with distant metastases. Based on their findings Moore & McCluggage recommended in a recent paper that these neoplasms should be regarded as LG-ESS [19].
In some instances, the histological diagnosis of EST could be problematic, especially on biopsy. Some samples may contain foamy histiocytes, foci of hyalinisation [15,16,20], smooth muscle differentiation [17,[21][22][23][24][25][26][27][28][29][30] where radiated collagen fibres of the smooth The mitotic rate is not high (less than 10 × 10 HPF). Areas of coagulative necrosis and sex-cord-like differentiation may be identified, but, by definition, lympho-vascular invasion (LVIS) is not present. Areas of smooth muscle metaplasia may be present and these should not mislead to an incorrect diagnosis of myometrial invasion. The differential diagnosis includes cellular leiomyoma and LG-ESS. The vascular pattern of ESN, composed of typical arterioles, is not a prominent feature of cellular leiomyoma. The presence of large blood vessels, one of the characteristics of cellular leiomyoma, may also be detected in ESN, but is not as conspicuous as in cellular leiomyoma. In addition, ESN usually does not contain the clefts frequently seen in cellular leiomyoma [17].
ESNs can be differentiated from LG-ESS exclusively by the presence of pushing margins and lack of LVIS [3,15,16], therefore, the definitive diagnosis of ESN can be rendered on resection specimens only and cannot be confidently established on biopsy or tissue removal systems (i.e., MyoSure resections). Nevertheless, even in resection specimens, the differential diagnosis between ESNs and LG-ESS may not be straightforward and extensive sampling may be required.

LG-ESS Morphology
LG-ESS shows the same histopathological features as ESN, except, as has already been mentioned, for the presence of infiltrative/permeative, tongue-like margins ( Figure 2A) and LVIS that may also be observed in the parametrial veins [15].
CD10 antibody is routinely used for the diagnosis of ESTs, ( Figure 3A) and is the most popular antibody used to differentiate LG-ESS from HG-ESSs [54][55][56][57][58]. However, it is well known that CD10 is not specific for the diagnosis of ESTs [59,60] and some ESTs may show negative immunostaining with CD10 [61,62]. In addition, CD10 can also be strongly positive in undifferentiated uterine sarcoma [63].
CD10 antibody is routinely used for the diagnosis of ESTs, ( Figure 3A) and is the most popular antibody used to differentiate LG-ESS from HG-ESSs [54][55][56][57][58]. However, it is well known that CD10 is not specific for the diagnosis of ESTs [59,60] and some ESTs may show negative immunostaining with CD10 [61,62]. In addition, CD10 can also be strongly positive in undifferentiated uterine sarcoma [63].
A diffuse or focal positivity for SMA [56,58,66] ( Figure 4A) and desmin ( Figure 4B) is present [54,58,65,70]. H-caldesmon can be focally positive [56] and, typically, strong positivity is seen in areas of smooth muscle differentiation with a 'starburst' appearance [31]. A weak expression of gonadotropin-releasing hormone receptor (GnRH-R) has been reported in ~95% of cases [64]. Aromatase (CYP19A1) expression has been observed in ~85% of cases [64]. Interferon-induced transmembrane protein-1 (IFITM1), a novel marker for endometrial stromal cells, is positive in ~80% of cases. IFITM1 is superior to CD10 in differentiating ESTs from smooth muscle neoplasm (~30% in cases of leiomyomas and leiomyosarcomas) [71]. However, ~90% of carcinosarcomas demonstrate positivity with this marker, but in this case the differential diagnosis with ESTs would be based on pure morphological grounds.  The nuclear expression of β-catenin was reported in 50% of cases, with 90% demonstrating nuclear expression of Lymphoid Enhancer-binding Factor 1 (LEF1). This finding is suggestive of activation of the Wnt pathway in LG-ESS [72].
LG-ESSs are mainly reported to be negative with Cyclin D1 and BCOR [73] and both markers are commonly implemented in diagnostic differential panel between LG-ESS and HG-ESS.
However, in some cases, Cyclin D1 can be positive in the LG-ESS, but with concomitant strong ER and PR positivity and focal CD10 expression, which is unusual in high-grade endometrial stromal sarcoma [68,74]. The Ki67 proliferation index is usually low and has been reported to be positive in~5-20% of the lesional cells [54,68,73,74]. It is worth mentioning that Ki67 can vary between tumours found in the same uterine specimen. Fujiishi et al. found the difference in the expression of Ki67 between the right anterior, right posterior, and fundal tumours to be 10%, 10%, and 3%, respectively [56].

Molecular Biology
LG-ESSs are genetically heterogeneous with relatively numerous identified chromosomal translocations resulting in gene fusions. However, approximately one third of these tumours do not harbour genetic fusions [7]. JAZF1-SUZ12 is the most common gene fusion, present in approximately half of the cases [31,78,79]  The nuclear expression of β-catenin was reported in 50% of cases, with 90% demonstrating nuclear expression of Lymphoid Enhancer-binding Factor 1 (LEF1). This finding is suggestive of activation of the Wnt pathway in LG-ESS [72].
LG-ESSs are mainly reported to be negative with Cyclin D1 and BCOR [73] and both markers are commonly implemented in diagnostic differential panel between LG-ESS and HG-ESS.
However, in some cases, Cyclin D1 can be positive in the LG-ESS, but with concomitant strong ER and PR positivity and focal CD10 expression, which is unusual in highgrade endometrial stromal sarcoma [68,74]. The Ki67 proliferation index is usually low and has been reported to be positive in ~5-20% of the lesional cells [54,68,73,74]. It is worth mentioning that Ki67 can vary between tumours found in the same uterine specimen. Fujiishi et al. found the difference in the expression of Ki67 between the right anterior, right posterior, and fundal tumours to be 10%, 10%, and 3%, respectively [56].

High-Grade Endometrial Stromal Sarcoma (HG-ESS)
On gross examination, these tumours usually show haemorrhage and necrosis [5,[91][92][93]. The morphological spectrum varies according to the genetic aberrations. These neoplasms [5,[90][91][92]94] show necrosis with a permeative pattern of myometrium invasion. LVIS is a constant finding. The cellular population is a mixture of round and spindle cells. The round cell component displays high cellularity with a vague nested architecture ( Figure 6A), composed of cells with a scanty/moderate eosinophilic cytoplasm ( Figure 6B). The nuclei exhibit a finely granular to slightly vesicular chromatin with an irregular nuclear membrane, no prominent nucleoli and no significant nuclear atypia/pleomorphism. Sex-cord-like differentiation and pseudo-glandular/pseudo-papillary pattern may rarely be observed. The mitotic rate within the round cells component is high.

Morphology
These neoplasms [5,[90][91][92]94] show necrosis with a permeative pattern of myometrium invasion. LVIS is a constant finding. The cellular population is a mixture of round and spindle cells. The round cell component displays high cellularity with a vague nested architecture ( Figure 6A), composed of cells with a scanty/moderate eosinophilic cytoplasm ( Figure 6B). The nuclei exhibit a finely granular to slightly vesicular chromatin with an irregular nuclear membrane, no prominent nucleoli and no significant nuclear atypia/pleomorphism. Sex-cord-like differentiation and pseudo-glandular/pseudo-papillary pattern may rarely be observed. The mitotic rate within the round cells component is high.
Approximately 50% of cases show a low-grade spindle cell component ( Figure 7A  Approximately 50% of cases show a low-grade spindle cell component ( Figure 7A) with low/intermediate cellularity, a fascicular pattern of growth consisting of a proliferation of bland spindle cells set in a fibro-collagenous to fibro-myxoid matrix, juxtaposed to the round cell component. The nuclei show even chromatin with no prominent nucleoli. The mitotic index in this component is low.

Molecular Biology
These neoplasms demonstrate rearrangement of YWHAE ( Figure 8) and commonly harbour YWHAE-NUTM2 fusion [5,7,57]. NUTM2 has been previously named as FAM22 by HUGO Gene Nomenclature Committee (HGNC) Symbols [98]. There are several members of paralog genes in this family, of which NUTM2A, NYTM2B and NUTM2E protein coding genes have been associated with HG-ESSs. Mainly NUTM2A and NUTM2B have been reported to form fusion with YWHAE as alternative gene fusion partners [7,97]. In a more recent report, NUTM2E (paralog of NUTM2B) has also been detected as an alternative gene fusion partner [89].
The low-grade spindle cell component may display positive staining with ER, PR, and CD10 [63,74] or show diffuse positivity with Cyclin D1 and PR and focal positivity for CD10 and p16 [74], or demonstrate positive staining with Cyclin D1 and negative staining with CD10 [96].

Molecular Biology
These neoplasms demonstrate rearrangement of YWHAE ( Figure 8) and commonly harbour YWHAE-NUTM2 fusion [5,7,57]. NUTM2 has been previously named as FAM22 by HUGO Gene Nomenclature Committee (HGNC) Symbols [98]. There are several members of paralog genes in this family, of which NUTM2A, NYTM2B and NUTM2E protein coding genes have been associated with HG-ESSs. Mainly NUTM2A and NUTM2B have been reported to form fusion with YWHAE as alternative gene fusion partners [7,97]. In a more recent report, NUTM2E (paralog of NUTM2B) has also been detected as an alternative gene fusion partner [89].
WT1 gene expression is often absent or shows low expression levels [89]. Notably, it has been previously highlighted that high immunohistochemical expression of CD117 (c-kit) can be frequently found in tumours with YWHAE genetic rearrangement, but c-kitimmunoreactive YWHAE-NUTM2A/B sarcomas have not demonstrated known mutations in KIT gene [99].
WT1 gene expression is often absent or shows low expression levels [89]. Notably, it has been previously highlighted that high immunohistochemical expression of CD117 (ckit) can be frequently found in tumours with YWHAE genetic rearrangement, but c-kitimmunoreactive YWHAE-NUTM2A/B sarcomas have not demonstrated known mutations in KIT gene [99].

Morphology
These tumours often show neoplastic-type or infarct-type necrosis [92]. The pattern of myometrial invasion could be infiltrative and tongue-like, similar to LG-ESS, or may display a broad front of invasion with irregular borders. A mixed pattern may also be present. LVIS is a common finding. The neoplasm shows a fascicular pattern and is composed of cells with eosinophilic cytoplasm (scanty/moderate or abundant), and spindle/oval, occasionally round, nuclei with a finely dispersed chromatin and no discernible nucleoli. Although infrequent, severe nuclear atypia/pleomorphism has been described. The stroma is myxoid in the majority of cases and collagen plaques may be identified. The intra-tumoural vessels may be large-sized or arterioles; a hemangiopericytoma-like vascular pattern may be seen. Occasionally, tumours may contain benign-appearing endometrioid glands. The proliferation rate may be very low (1 × 10 HFP) or moderate/high (50 × 10 HFP). The accompanying LG-ESS component has not been reported.
A rare novel sub-type of HG-ESS with ZC3H7B-BCOR fusion has been described that shares significant histopathological overlap with myxoid leiomyosarcoma [100].

Morphology
These tumours often show neoplastic-type or infarct-type necrosis [92]. The pattern of myometrial invasion could be infiltrative and tongue-like, similar to LG-ESS, or may display a broad front of invasion with irregular borders. A mixed pattern may also be present. LVIS is a common finding. The neoplasm shows a fascicular pattern and is composed of cells with eosinophilic cytoplasm (scanty/moderate or abundant), and spindle/oval, occasionally round, nuclei with a finely dispersed chromatin and no discernible nucleoli. Although infrequent, severe nuclear atypia/pleomorphism has been described. The stroma is myxoid in the majority of cases and collagen plaques may be identified. The intra-tumoural vessels may be large-sized or arterioles; a hemangiopericytoma-like vascular pattern may be seen. Occasionally, tumours may contain benign-appearing endometrioid glands. The proliferation rate may be very low (1 × 10 HFP) or moderate/high (50 × 10 HFP). The accompanying LG-ESS component has not been reported.
A rare novel sub-type of HG-ESS with ZC3H7B-BCOR fusion has been described that shares significant histopathological overlap with myxoid leiomyosarcoma [100].

Immunohistochemistry
There is positive staining with Cyclin D1 and CD10; immunoreactivity with BCOR is seen in~50% of cases, but in a recent paper, the BCOR positivity was up to~80% of the lesional cells [77,101,102]. Expression of ER and PR is variable. Focal positive staining with SMA and caldesmon can be identified, but positivity with desmin is usually negative [7]. Some recent reports also demonstrated immunoreactivity with TLE1 (Transducin-like enhancer protein 1), CD99 and BCL2 [101][102][103].

Molecular Biology
ZC3H7B-BCOR and its reciprocal fusion are mainly associated with and reported in these neoplasms [77,93,105]. Recent reports showed MDM2, FRS2 and CDK4 amplification and loss of CDKN2A in some cases [89,105,106].
Yoshida et al. showed elevated expression of BCOR and significant upregulation of ZIC2, HOXA13 and NTRK3 in an extra-uterine case (chest wall) of HG-ESS with ZC3H7B-BCOR fusion [107].

Immunohistochemistry
Clinical experience with regard to these tumours is very limited, with only a few reported cases. They show a different immunoprofile with respect to ZC3H7B-BCOR neoplasms. They display diffuse positive immunoreactivity with BCOR and Cyclin D1, less positive staining for CD10 and mostly negative staining with ER and PR. In addition, they may show immunoreactivity with desmin. SMA and caldesmon seem to be negative [77,93,108]. There is strong and diffuse cytoplasmic expression of pan-Trk [109].

Molecular Biology
Juckett et al. reported that BCOR-ITDs occurred most frequently in exon 15 and near C-terminus, and were present in 52.4% cases of uterine sarcomas. Interestingly, the tested cases did not carry any of the simultaneous gene fusions typically associated with ESSs [93].
Lin et al. reported no amplification of CDK4 and/or MDM2; however, a homozygous deletion of CDKN2A and CDKN2B was present in 20% of cases. Mutations in STAG2, PASK, SMARCB1, ATRX, CTNNB1 and ARID1A were also seen in the minority of the tested BCOR-ITDs cases [105]. Upregulation of the expression of NTRK3, FGFR3, RET, BCOR, GLI1 and PTCH1 genes has also been reported [109].

NTRK-Uterine Tumours
Morphology, Immunohistochemistry and Molecular Biology These sarcomas show no definite endometrial stromal origin. Chiang et al. [110] describes a few cases of a sub-type of sarcomas with novel NTRK fusion. These tumours demonstrate RBPMS-NTRK3, TPR-NTRK1, LMNA-NTRK1 and TPM3-NTRK1 gene fusions and seem to affect premenopausal women with frequent cervical involvement (three in cervix uteri and one in corpus uteri). They show infiltrative or expansive myometrial invasion and are composed of fascicles of cells with spindle nuclei, small nucleoli and abundant eosinophilic cytoplasm. Severe nuclear atypia/pleomorphism and necrosis may be present. The stroma is myxoid/edematous. The vascular pattern may be either delicate with thin-walled vessels or composed of thick-walled blood vessels. LVIS is not identified. The proliferation rate is relatively high. Immunohistochemistry shows positive immunostaining with CD10 (not all cases), focal positive staining with SMA and very focal positivity (<10% of lesional cells) with S100. There is positive immunoreactivity with TrkA and pan-Trk. H3K27me3 expression is retained. AE1/3, desmin, ER, PR, CD34 and SOX-10 are negative [110].
The fascinating finding is that COL1A1-PDGFB rearrangement is a characteristic of dermatofibrosarcoma protuberans (DFS) and has not been reported in uterine sarcomas.
The authors classified the S100-positive sarcomas as malignant peripheral nerve sheath tumours. This group showed diffuse positive staining with Cyclin D1 and focal positivity with BCOR. CD34, HMB45, Melan A, ER, PR, desmin, Trk and H3K27me3 were negative [111].
Grindstaff et al. recently described an additional COL1A1-PDGFB fusion-positive uterine case with diffuse CD34 immunopositivity. There was focal positive staining with CD10, SMA and positive aberrant expression of p53. The Ki67 proliferation index was high. Desmin, caldesmon and p16 were negative [112].
Michal et al. described a STRN-NTRK3-rearranged uterine sarcoma with a peculiar morphology. The neoplasm contained bland epithelioid/plasmacytoid cells embedded in a myxoid stroma with a complex, arborising vascular pattern and focal peri-vascular hyalinisation. There was infarct-type necrosis but frank tumoural necrosis, significant nuclear pleomorphism and mitoses were not seen. The neoplastic cells displayed strong positive immunostaining with S100, CD34 and nuclear and cytoplasmic Pan-Trk. The extensive immunohistochemistry with keratins, CD10, myogenic, peri-neural/neural, melanocytic, neuroendocrine and vascular markers showed negative staining. The Ki67 proliferation rate was low [114].

High-Grade Sarcoma Not Otherwise Specified (NOS)
The 2020 WHO classification of EST contains a vague sub-category that seems to be associated with a LG-ESS component [7].

Undifferentiated Uterine Sarcomas (UUS) UUS
Morphology, Immunohistochemistry and Molecular Biology UUS is a rare uterine sarcoma [115][116][117] that includes a variegated group of neoplasms with no specific line of differentiation and, by definition, is a diagnosis of exclusion. The patients with UUS are postmenopausal and present with uterine bleeding and pelvic pain. Macroscopically, UUS usually displays necrosis and haemorrhage. Morphologically, UUS can be sub-divided into monomorphic and pleomorphic sub-types [118]. It consists of epithelioid and/or spindled cells with high mitotic rate and a destructive pattern of myometrial invasion. LVIS and necrosis are common findings [119]. When UUS consists of a uniform cellular population, HG-ESS harbouring YWHAE-NUTM2 (FAM22) fusion should be ruled out. Conversely, some pleomorphic sarcomas may exhibit YWHAE, JAZF1 and NTRK rearrangements. Therefore, these neoplasms should not be classified as UUS [77,78,110,111]. Tumours displaying high-grade nuclear atypia/pleomorphism, but associated with a LG-ESS component, should be classified under endometrial stromal sarcoma NOS category [118].
UUSs can harbour simultaneous numerous gene fusions. Brahmi et al. [89] have reported cases with multiple detected (>3) in-frame fusions and a case with novel translocation CREBBP-BCOR. Moreover, UUS cases demonstrated complex genomic profiles with numerous gene fusions and/or had mutations involving TP53, KRAS, NRAS or BRAF genes.
Novel YWHAE gene rearrangement with no known partner has been reported in two UUSs with marked nuclear pleomorphism. This novel rearrangement may represent novel fusions in this sarcoma subtype [63]. A single case (1/23) harbouring HMGA2-RAD51B fusion has been described, which also demonstrated high expression of NTRK3, FGFR3, RET, BCOR, GLI1 and PTCH1, and low expression of ESR1. [109]. Other cases have demonstrated low expression of these genes along with low ESR1 (17/23) expression; however, no fusions have been observed [109].

Conclusions
In the era of Next Generation Sequencing (NGS) there is a huge effort to integrate the morphological and immunohistochemical classification of ESTs with molecular subcategorisation. The aim of the molecular classification of ESTs, as in other neoplasms, is not a purely academic exercise, but rather an understanding of the molecular bases in order to develop a specific target therapy for each sub-category. This will be of paramount importance for planning a therapeutic strategy in the metastatic disease. Therefore, in the future, we will be witnessing discoveries of new entities and, gradually, the histopathological classification will be replaced by molecular classification. Genetic profile of ESSs with the most frequently reported and novel molecular alterations are presented in Table 2.

Acknowledgments:
The authors would like to thank Paul Roberts for providing the photo in Figures  5 and 8.

Conflicts of Interest:
The authors declare no conflict of interest.