Machine Learning-Supported Diagnosis of Small Blue Round Cell Sarcomas Using Targeted RNA Sequencing.

Small blue round cell sarcomas (SBRCSs) are a heterogeneous group of tumors with overlapping morphologic features but markedly varying prognosis. They are characterized by distinct chromosomal alterations, particularly rearrangements leading to gene fusions, whose detection currently represents the most reliable diagnostic marker. Ewing sarcomas are the most common SBRCSs, defined by gene fusions involving EWSR1 and transcription factors of the ETS family, and the most frequent non-EWSR1-rearranged SBRCSs harbor a CIC rearrangement. Unfortunately, currently the identification of CIC::DUX4 translocation events, the most common CIC rearrangement, is challenging. Here, we present a machine-learning approach to support SBRCS diagnosis that relies on gene expression profiles measured via targeted sequencing. The analyses on a curated cohort of 69 soft-tissue tumors showed markedly distinct expression patterns for SBRCS subgroups. A random forest classifier trained on Ewing sarcoma and CIC-rearranged cases predicted probabilities of being CIC-rearranged >0.9 for CIC-rearranged-like sarcomas and <0.6 for other SBRCSs. Testing on a retrospective cohort of 1335 routine diagnostic cases identified 15 candidate CIC-rearranged tumors with a probability >0.75, all of which were supported by expert histopathologic reassessment. Furthermore, the multigene random forest classifier appeared advantageous over using high ETV4 expression alone, previously proposed as a surrogate to identify CIC rearrangement. Taken together, the expression-based classifier can offer valuable support for SBRCS pathologic diagnosis.

Renal CIC-LEUTX rearranged sarcoma with multiple pulmonary metastases: a case report and literature review.

BACKGROUND: CIC-rearranged sarcomas (CRS) are a group of heterogeneous tumors which mostly occur in the soft tissues of limbs and trunk, and are highly invasive with poor prognosis. Here, we describe a rare case of CRS that occurred in the left kidney with a CIC-LEUTX rearrangement. CASE PRESENTATION: A 45-year-old male was admitted to hospital with a dry cough for more than two months without obvious cause. Physical examination and laboratory tests revealed no notable abnormality. The CT scan demonstrated a mass in the left kidney and multiple nodules in both lungs. The percutaneous core needle biopsy showed similar histomorphology and immunophenotype of small round cell malignant tumors. Genetic test revealed a CIC-LEUTX gene fusion. CONCLUSIONS: We present a rare primary renal CRS with multiple pulmonary metastases, and LEUTX is confirmed as the fusion partner of CIC gene for the first time in a renal case.

A Practical Approach to Small Round Cell Tumors Involving the Gastrointestinal Tract and Abdomen.

Small round cell neoplasms are diagnostically challenging owing to their clinical and pathologic overlap, necessitating use of large immunopanels and molecular analysis. Ewing sarcomas (ES) are the most common, but EWSR1 is translocated in several diverse neoplasms, some with round cell morphology. Molecular advances enable classification of many tumors previously termed 'atypical ES'. The current WHO Classification includes two new undifferentiated round cell sarcomas (with CIC or BCOR alterations), and a group of sarcomas in which EWSR1 partners with non-Ewing family transcription factor genes. This article reviews the spectrum of small round cell sarcomas within the gastrointestinal tract and abdomen.

[Clinical and morphological characteristics of Ewing's sarcoma and the algorithm for diagnosing undifferentiated round cell sarcomas]. / Kliniko-morfologicheskaya kharakteristika sarkomy Yuinga i algoritm diagnostiki nedifferentsirovannykh kruglokletochnykh sarkom.

BACKGROUND: The group of undifferentiated round cell sarcomas, according to the World Health Organization Classification, in addition to Ewing's sarcoma (ES), includes round cell sarcoma with rearrangement of the EWSR1 gene with partners not from the ETS gene family, sarcoma with BCOR gene alterations, CIC -rearranged sarcoma. Despite the fact that all tumors have clear histological and immunological criteria, their diagnosis can be difficult, given the fact that there are overlapping variants of the morphological picture and immunophenotype both within the group and with other round cell tumors. OBJECTIVE: Present a comparative analysis of genetically verified ES, sarcoma with BCOR gene alterations and CIC-rearranged sarcoma. MATERIAL AND METHODS: A comparative study of biopsy specimens of bones, soft tissues and internal organs was carried out in 118 patients with ES, 10 with BCOR gene alterations and 8 with CIC-rearranged sarcomas. All cases were genetically verified. The following research methods were used: histological, immunohistochemical, RT-PCR, RNA sequencing and FISH. RESULTS: Within our cohort, it was shown that ES predominantly affects bones, while soft tissue localization is more typical for the other two undifferentiated round cell sarcomas. Histologically, in the overwhelming majority of cases, ES is characterized by a monomorphic round-cell structure; on the contrary, heterogeneous structure is typical for sarcoma with alterations of the BCOR gene, CIC-rearranged sarcoma. High sensitivity and specificity of CD99/NKX2.2 co-expression for ES, BCOR/SATB2/TLE1 for sarcoma with BCOR gene alterations, high specificity and low sensitivity of WT1/ETV4 co-expression for CIC-rearranged sarcoma was shown. CONCLUSION: For the differential diagnosis of undifferentiated round-cell sarcomas, it is necessary to take into account the clinical, morphology when compared with the data of the IHC study, and verification by molecular genetic methods is necessary to improve the accuracy of diagnosis.

[CIC-rearranged sarcoma: a case report and literature review]. / CIC-perestroennaya sarkoma: klinicheskoe nablyudenie i obzor literatury.

CIC-rearranged sarcoma is a rare and extremely aggressive tumor that occurs mainly in soft tissues. Despite the fact that identification of a characteristic genetic rearrangement is necessary to verify the diagnosis, in most cases, the correct diagnosis can be made by comparing histological signs and a characteristic immunophenotype, which greatly speeds up the diagnosis. The article describes a case of CIC-rearranged sarcoma in a 14-year girl with the successful application of the CWS-2009 treatment protocol.

CIC-rearranged sarcoma presenting with superior vena cava syndrome: case report.

CIC-rearranged sarcomas are rare mesenchymal neoplasms belonging to the family of undifferentiated small round cell sarcomas. This report details the case of a 45-year-old man presenting with symptoms of mediastinal compression, radiological diagnosis of a mediastinal mass and rapid evolution to full-blown superior vena cava syndrome. The emergency was successfully managed with a pharmacological approach. Formulation of a pathological diagnosis of CIC-rearranged sarcoma was initially supported by fluorescence in situ hybridisation findings and later validated by next-generation sequencing, which showed CIC-DUX4 gene fusion. A chemotherapy regimen was started with immediate benefits for the patient. The spectrum of pathological entities able to cause superior vena cava syndrome is wide, and recognition of rare causes is important to tailor the therapeutic approach to the specific disease. This is, to the best of our knowledge, the first report of CIC-rearranged sarcoma presenting with superior vena cava syndrome.

Approach to Primary Vertebral Tumors in the Light of the 2020 Updated World Health Organization Classification of Bone Tumors.

AIM: To define a new approach for primary vertebral tumors by discussing them in the light of 2020 World Health Organization (WHO) classification of bone tumors. MATERIAL AND METHODS: In this study, we have discussed primary vertebral tumors in light of the 2020 Updated WHO Classification of Soft Tissue and Bone Tumors. RESULTS: Chondroblastoma and chondromyxoid fibroma has been classified in the benign category, while synovial chondromatosis has been moved from the benign category to the intermediate category. In the updated classification, grade I chondrosarcoma has been classified in the malignant category with grade II?III chondrosarcomas. Minor amendments have been made for osteosarcoma subtypes. Neoplasms of undetermined origin, such as aneurysmal cysts, simple bone cysts, fibrosis dysplasia, and osteofibrous dysplasia, have been classified as neoplastic lesions. Chordomas have been classified into ?not otherwise stated,? poorly differentiated chordomas, and dedifferentiated chordomas. Ewing?s sarcomas have been classified in a separate section for undifferentiated, small, round cell sarcomas of the bone and soft tissue. In this section, three distinct subsets different from Ewing?s sarcoma have been discussed. CIC-rearranged sarcoma, BCOR-rearranged sarcoma, and round cell sarcomas with EWSR1 gene fusion with non-ETS family members. CONCLUSION: In this study, we have reviewed the new classifications and discussed their effect on decision making in spinal oncologic surgery.

Patterns of care and outcome of CIC-rearranged sarcoma patients: A nationwide study of the French sarcoma group.

BACKGROUND: CIC-rearranged sarcomas (CIC-RS) represent the most frequent subset of "Ewing-like" undifferentiated small round cell sarcomas. These tumors tend to be more aggressive than Ewing sarcomas. Moreover, treatment strategy can differ according to teams. The primary aim of this retrospective study was to describe the characteristics, treatments, and outcome for patients with CIC-RS included in the French NETSARC+ database. METHODS: Pediatric and adult patients from 13 French centers with a diagnosis of CIC-RS were registered from October 2008 to March 2021. Patients and tumors characteristics were collected from the national network NETSARC+ database (http://netsarc.sarcomabcb.org). CIC-RS diagnosis was pathologically and molecularly confirmed with a central review by expert pathologists. Two groups of patients were studied: those treated as classical Ewing sarcomas (cohort EwS) and those treated as high-grade soft tissue sarcomas (cohort STS) according to ESMO and/or EpSSG guidelines. Survival was calculated using the Kaplan-Meier method and the log-rank test was used to compare survival. RESULTS: Among 79 patients, the male/female sex ratio was 0.7 and the median age at diagnosis was 27 years (range 2-87). With a median follow-up of 37 months, 39 patients died of the disease. Median overall survival from diagnosis was 18 months, with no significant difference between both cohorts (p = 0.9). Nevertheless, when focusing on patients with metastatic disease at diagnosis (N = 21), all patients from cohort STS died of disease while some patients from cohort EwS were still alive and in complete remission. CONCLUSION: FSG experience confirms the aggressive clinical course of CDS patients regardless of chemotherapy regimen.

Ewing and Ewing-like sarcomas: A morphological guide through genetically-defined entities.

The fifth edition of the World Health Organization classification of soft tissue and bone tumors redefined Ewing sarcoma by fusions between EWSR1/FUS and ETS family of transcription factors, and recognized three tumor groups among Ewing-like sarcoma: CIC-rearranged sarcoma, sarcoma with BCOR genetic alterations, and round cell sarcoma with EWSR1::non-ETS fusions. Although this classification underscores the critical role of molecular genetics in the diagnosis of small round cell sarcoma, each entry is recognized as a specific entity not only because they have different genetics but because their phenotypes are distinct and reasonably robust to support the diagnosis. This review focuses on the morphological aspects of Ewing sarcoma and a subset of Ewing-like sarcomas (CIC-rearranged sarcoma, BCOR-associated sarcoma, and EWSR1::NFATC2 sarcoma) for which phenotypic characteristics have been well established. Classic histological findings, uncommon variations, and recurrent diagnostic pitfalls are addressed, along with the utility of recently developed immunohistochemical markers (NKX2.2, PAX7, ETV4, BCOR, CCNB3, and NKX3.1). Phenotypic expertise would significantly expedite the diagnostic process and complement (or sometimes outperform) genetic testing, even in well-resourced settings. Morphological knowledge plays an even more substantial role in facilities that do not have easy access to molecular testing.

[CIC-rearranged sarcoma with rhabdoid features: a clinicopathological analysis].

Objective: To investigate the histopathologic, immunohistochemical, molecular genetic characteristics of CIC-rearranged sarcoma (CRS) with rhabdoid features. Methods: The clinical and pathologic data of two cases of CRS diagnosed between 2019 and 2021 at the Department of Pathology, Jiangsu Province Hospital were analyzed. Immunohistochemical study and fluorescence in situ hybridization (FISH) were performed. The relevant literature was reviewed. Results: Both patients were female, one was 58 years old, with tumor located in left thigh; the other was 43 years old, with tumor located in left pelvic cavity. Microscopically, both tumors were composed of small to medium-sized round, oval cells, arranged in nodules or sheets. The tumor cells showed irregular nuclear outline, coarse chromatin with prominent nucleoli and brisk mitotic activity. Both cases showed rhabdoid phenotype with myxoid stromal changes. Immunohistochemically, both cases were positive for CD99 and c-myc. High WT1 reactivity was seen in classic area, with low reactivity in rhabdoid area. There was no INI1 lost in both cases. Both were negative for NKX2.2 and NKX3.1. By FISH both cases demonstrated convincing break-apart signals of CIC gene. One patient died of disease after 1 month, and the other died of disease after 3 months. Conclusions: CRS is a small round cell undifferentiated sarcoma of the bone and soft tissue defined by molecular genetic characteristics, and may show atypical morphologic and immunophenotypic characteristics such as rhabdoid features. A correct understanding of its rare morphologic and immunophenotypic characteristics, combined with molecular pathologic detection, is conducive to correct diagnosis.

Small round cell sarcomas.

Undifferentiated small round cell sarcomas (SRCSs) of bone and soft tissue comprise a heterogeneous group of highly aggressive tumours associated with a poor prognosis, especially in metastatic disease. SRCS entities mainly occur in the third decade of life and can exhibit striking disparities regarding preferentially affected sex and tumour localization. SRCSs comprise new entities defined by specific genetic abnormalities, namely EWSR1-non-ETS fusions, CIC-rearrangements or BCOR genetic alterations, as well as EWSR1-ETS fusions in the prototypic SRCS Ewing sarcoma. These gene fusions mainly encode aberrant oncogenic transcription factors that massively rewire the transcriptome and epigenome of the as yet unknown cell or cells of origin. Additional mutations or copy number variants are rare at diagnosis and, depending on the tumour entity, may involve TP53, CDKN2A and others. Histologically, these lesions consist of small round cells expressing variable levels of CD99 and specific marker proteins, including cyclin B3, ETV4, WT1, NKX3-1 and aggrecan, depending on the entity. Besides locoregional treatment that should follow standard protocols for sarcoma management, (neo)adjuvant treatment is as yet ill-defined but generally follows that of Ewing sarcoma and is associated with adverse effects that might compromise quality of life. Emerging studies on the molecular mechanisms of SRCSs and the development of genetically engineered animal models hold promise for improvements in early detection, disease monitoring, treatment-related toxicity, overall survival and quality of life.

A case of CIC-rearranged sarcoma with CIC-LEUTX gene fusion in spinal cord.

A 16-year-old male was admitted to the hospital for weakness of both lower extremities. Magnetic resonance imaging revealed an intraspinal extramedullary subdural mass at the thoracic 9 level. Microscopically, the tumor cells were small to medium sized and round to ovoid in shape. They were distributed in diffuse sheets or showed nodular appearance. The nucleus of the tumor had mild-to-moderate atypia, with vesicular chromatin and prominent nucleoli. A smaller proportion of tumor cells demonstrated rhabdoid morphology. Focal myxoid stromal change was present, in which tumor cells exhibited spindle shapes. Approximately two mitoses were counted per 10 high-power fields. No necrosis was observed. The tumor cells were focal positive for CD99; multifocal positive for WT1; diffuse positive for nestin, synaptophysin, and D2-40; partial positive for GFAP; focal positive for desmin and SSTR2; and scattered positive for S-100 protein. The Ki-67 labeling index was approximately 20%. Genetic testing revealed CIC-LEUTX gene fusion. Considering the patient's history, clinical data, pathological findings and genetic findings, we rendered a rare tumor named CIC-rearranged sarcoma with CIC-LEUTX gene fusion.

Novel ATXN1/ATXN1L::NUTM2A fusions identified in aggressive infant sarcomas with gene expression and methylation patterns similar to CIC-rearranged sarcoma.

CIC-rearranged sarcomas are newly defined undifferentiated soft tissue tumors with CIC-associated fusions, and dismal prognosis. CIC fusions activate PEA3 family genes, ETV1/4/5, leading to tumorigenesis and progression. We report two high-grade CNS sarcomas of unclear histological diagnosis and one disseminated tumor of unknown origin with novel fusions and similar gene-expression/methylation patterns without CIC rearrangement. All three patients were infants with aggressive diseases, and two experienced rapid disease deterioration and death. Whole-transcriptome sequencing identified an ATXN1-NUTM2A fusion in the two CNS tumors and an ATXN1L-NUTM2A fusion in case 3. ETV1/4/5 and WT1 overexpression were observed in all three cases. Methylation analyses predicted CIC-rearranged sarcoma for all cases. Retrospective IHC staining on case 2 demonstrated ETV4 and WT1 overexpression. ATXN1 and ATXN1L interact with CIC forming a transcription repressor complex. We propose that ATXN1/ATXN1L-associated fusions disrupt their interaction with CIC and decrease the transcription repressor complex, leading to downstream PEA3 family gene overexpression. These three cases with novel ATXN1/ATXN1L-associated fusions and features of CIC-rearranged sarcomas may further expand the scope of "CIC-rearranged" sarcomas to include non-CIC rearrangements. Additional cases are needed to demonstrate if ATXN1/ATXN1L-NUTM2A fusions are associated with younger age and more aggressive diseases.

CIC-DUX4 sarcomas.

PURPOSE OF REVIEW: CIC-DUX4 sarcoma (CDS) is a high-grade undifferentiated round cells sarcoma that belongs to the undifferentiated round cell sarcomas family. It represents less than one percent of sarcomas, defining a rarest among rare malignancies. It affects young adults, displaying soft tissue mass. Considered very aggressive, a high proportion of cases display an advanced disease with lung metastasis at diagnosis. Here we discuss recent progress in molecular characterization of CDS, the main tracks of CDS biology and the current and future prospects of therapeutic approaches. RECENT FINDINGS: CDS is characterized by a specific oncogenic translocation CIC::DUX4 that induce ETV4 overexpression. Patients with CDS show an aggressive clinical course and have a significantly unfavorable outcome compared to Ewing sarcoma. As of today, there is a lack of consensus on whether they should be treated with an Ewing-like approach, as currently done by most sites, or regarded as high-grade soft tissue sarcoma (STS). Anyway, when feasible, combination regimens including anthracycline and alkylating agents should be favored and patients should not benefit from a therapeutic de-escalation. Overall, registration within clinical trials and prospective registries is recommended. SUMMARY: Overall, CDS showed a poor prognosis regardless of the patterns of treatment that warrant biological studies to better understand the disease.

Systemic treatments and outcomes in CIC-rearranged Sarcoma: A national multi-centre clinicopathological series and literature review.

CIC-rearranged sarcoma is a recently established, ultra-rare, molecularly defined sarcoma subtype. We aimed to further characterise clinical features of CIC-rearranged sarcomas and explore clinical management including systemic treatments and outcomes. METHODS: A multi-centre retrospective cohort study of patients diagnosed between 2014-2019. RESULTS: Eighteen patients were identified. The median age was 27 years (range 13-56), 10 patients were male (56%), 11 patients (61%) had localised disease and 7 patients had advanced (metastatic or unresectable) disease at diagnosis. Of 11 patients with localised disease at diagnosis, median overall survival (OS) was 40.6 months and the 1-, 2- and 5-year OS estimates were 82%, 64% and 34% respectively. Nine patients (82%) underwent surgery (all had R0 resections), 8 (73%) patients received radiotherapy to the primary site (median dose 57Gy in 28 fractions), and 8 (73%) patients received chemotherapy (predominantly Ewing-based regimens). Metastases developed in 55% with a median time to recurrence of 10.5 months. In patients with advanced disease at diagnosis, median OS was 12.6 months (95% CI 5.1-20.1), 1-year OS was 57%. Median progression-free survival was 5.8 months (95% CI 4.5-7.2). Durable systemic therapy responses occurred infrequently with a median duration of systemic treatment response of 2.1 months. One durable complete response of metastatic disease to VDC/IE chemotherapy was seen. Responses to pazopanib (n = 1) and pembrolizumab (n = 1) were not seen. CONCLUSION: In this series, CIC-rearranged sarcomas affected young adults and had a high incidence of presenting with, or developing, metastatic disease. The prognosis overall was poor. In advanced disease, durable systemic therapy responses were infrequent.

Primary pancreatic Ewing sarcoma: a cytomorphologic and histopathologic study of 13 cases.

INTRODUCTION: Ewing sarcoma (ES) is a small, round cell sarcoma that rarely occurs in solid organs, including the pancreas. A diagnostic overlap exists with other primary pancreatic neoplasms, especially for specimens from small biopsies and fine needle aspiration (FNA). To improve the diagnosis of this rare pancreatic tumor, we have reported a series of 13 cases of primary pancreatic ES and reviewed the cytopathologic, surgical pathology, clinical, and radiologic features of these neoplasms. MATERIALS AND METHODS: We performed a retrospective case review of 13 patients with a diagnosis of pancreatic ES from 2 tertiary academic medical centers. A combination of cytology and histopathologic slides were reviewed, and the patient demographics, clinical information, somatic genetics, and radiologic findings were obtained from the electronic medical records. RESULTS: Five FNA specimens from 5 patients and 8 surgical biopsy or resection specimens were identified and reviewed. The patients included 9 males and 4 females, with a median age of 27 years (range, 15-78 years). The cytology smears were highly cellular and showed a combination of complex tissue fragments and singly dispersed small round blue cells. The final diagnosis was ES for all 5 FNA specimens in accordance with the characteristic cytomorphology, diffuse and/or strong membranous immunolabeling for CD99, membranous ß-catenin, and molecular confirmation of EWSR1 using fluorescence in situ hybridization or reverse transcriptase polymerase chain reaction. CONCLUSIONS: The cytologic diagnosis of ES is challenging, especially in unusual locations such as the pancreas. However, the correct cytologic diagnosis is important because these patients will require neoadjuvant therapy before surgery. Confirmatory molecular studies should be required to render the diagnosis of pancreatic ES.

Ultrastructure of CIC-DUX4 sarcoma: the first pathological report.

CIC-DUX4 sarcoma (CDS) is a recently identified subtype of small round cell sarcoma. Morphologically, CDS partially resembles Ewing sarcoma (ES) and has been classified as "ES-like sarcoma"; however, detailed clinicopathologic and molecular genetic analyses have indicated that CDS is a new independent disease. Many studies have provided light microscopic, immunohistochemical, and genetic information about CDS. However, ultrastructural findings associated with this sarcoma are lacking. The aim of this study was to investigate the ultrastructure of CDS tumors and to compare their features with those of ES. We examined two cytogenetically confirmed CDS cases. We found that, compared to typical ES, CDS presented heterogeneity: in cell density, from tightly packed to loosely unconnected areas; in cell shape, from polygonal to pleomorphic with small processes; and in nuclear shape including round, oval, polygonal, elongated, invaginated, or wrinkled formations. However, abundant glycogen in the cytoplasm and rare cell adhesion apparatus between cells are major similarities between CDS and typical ES. Neuroendocrine granules, which are seen in rare ES cases, could not be identified in these two CDS cases. Although cytogenetic differences can validate a definite diagnosis, ultrastructural features could also provide important information about the differences between CDS and ES.