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Rhabdomyosarcomas (RMS) represent a family of aggressive soft tissue sarcomas that present in both children and adults. Pathologic risk stratification for RMS has been based on histologic subtype, with poor outcomes observed in alveolar rhabdomyosarcoma (ARMS) and the adult-type pleomorphic rhabdomyosarcoma (PRMS) compared to embryonal rhabdomyosarcoma (ERMS). Genomic sequencing studies have expanded the spectrum of RMS, with several new molecularly defined entities, including fusion-driven spindle cell/sclerosing rhabdomyosarcoma (SC/SRMS) and MYOD1-mutant SC/SRMS. Comprehensive genomic analysis has previously defined the mutational and copy number spectrum for the more common ERMS and ARMS and revealed corresponding methylation signatures. Comparatively, less is known about epigenetic correlates for the rare SC/SRMS or PRMS histologic subtypes. Herein, we present exome and RNA sequencing, copy number analysis, and methylation profiling of the largest cohort of molecularly characterized RMS samples to date. In addition to ARMS and ERMS, we identify two novel methylation subtypes, one having SC/SRMS histology and defined by MYOD1 p. L122R mutations and the other matching adult-type PRMS. Selected tumors from adolescent patients grouped with the PRMS methylation class, expanding the age range of these rare tumors. Limited follow-up data suggest that pediatric tumors with MYOD1-mutations are associated with an aggressive clinical course.

Rhabdomyosarcoma is a malignant neoplasm of mesenchymal cells, which shows varying degrees of striated muscle cell differentiation. It predominantly occurs in children while rarely found in adults. Involvement of the oral cavity accounts for only 10-12% of all head and neck cases. Herewith, we report a rare case of oral spindle cell / sclerosing rhabdomyosarcoma in a 47-year-old male presented with a small mass involving the gingiva of right upper incisor. The mass was excised with a preoperative diagnosis of gingival epulis and subjected to histopathological and immunohistochemical examination which confirmed it to be spindle cell / sclerosing rhabdomyosarcoma. Data regarding its clinical course, genetic abnormalities and prognosis as a combined subtype is scant. Keywords: Adult spindle cell/sclerosingrhabdomyosarcoma (S­ScRMS) Oral Cavity, Rhabdomyosarcoma, Maxillary Gingiva, Immunohistochemistry

Spindle cell/sclerosing rhabdomyosarcoma (ssRMS) is a rare subtype of rhabdomyosarcoma (RMS) that has fascicular spindle cell and/or sclerosing morphology. SsRMS has a diverse molecular background and is categorized into three groups: congenital/infantile ssRMS with a gene fusion involving the NCOA2 and VGLL2, ssRMS with the MYOD1 mutation, and ssRMS with no recurrent identifiable genetic alterations. Because ssRMS is a newly defined disease concept of RMS, the optimal treatment methods have not been determined. This results in unfavorable prognosis and consequently signals the urgent need for continuous research. Patient-derived cell lines are essential tools in basic and translational research. However, only two ssRMS cell lines with the MYOD1 mutation have been reported to date. Thus, we established a novel ssRMS cell line named NCC-ssRMS2-C1 using a surgically resected tumor tissue from an adult ssRMS patient. NCC-ssRMS2-C1 cells retained the copy number alterations corresponding to the original tumor and are categorized into the group with no recurrent identifiable genetic alterations. NCC-ssRMS2-C1 cells demonstrated constant proliferation, spheroid formation, and capability for invasion in vitro, reflecting the malignant features of the original tumor tissue. In a drug screening test, ssRMS demonstrated remarkable sensitivity to romidepsin, trabectedin, actinomycin D, and bortezomib. Hence, we conclude that the NCC-ssRMS2-C1 cell line is the first ssRMS cell line which belongs to the group with no recurrent identifiable genetic alterations, and it will be a useful resource in both basic and translational studies for ssRMS.

Spindle-cell/sclerosing rhabdomyosarcoma (ssRMS) is a rare subtype of rhabdomyosarcoma, characterized by unique pathological features. Although distinctive molecular backgrounds such as frequent mutations in MyoD1 have been reported, optimized therapy has not been fully developed, and further investigations are required. Patient-derived cancer models are critical tools for basic and pre-clinical studies. However, there is no model for ssRMS. Thus, this study aimed to develop a novel cell line from the tumor tissue of a patient with ssRMS. Using surgically resected tissue, we successfully established this cell line, named NCC-ssRMS1-C1. These cells exhibited spindle-shape morphology, consistent with the pathological observations of the original tumor tissue. Genetic studies demonstrated that NCC-ssRMS1-C1 cells retained original copy number alterations and the typical point mutation in MyoD1. Malignant phenotypes such as proliferation, spheroid formation, and invasion were confirmed in vitro by studying NCC-ssRMS1-C1 cells. Upon screening an anti-cancer agent library, sensitivity to conventional chemotherapeutic agents such as actinomycin D was revealed. We conclude that the NCC-ssRMS1-C1 cell line will be a useful resource for basic and pre-clinical studies.

Rhabdomyosarcomas are malignancies associated with a rhabdomyoblastic phenotype which can be demonstrated morphologically or by immunohistochemistry for MYOD1 and myogenin. Rhabdomyosarcomas are currently subdivided into 4 types in the 2013 WHO classification of tumors of soft tissue and bone, including embryonal rhabdomyosarcoma, alveolar rhabdomyosarcoma, spindle cell/sclerosing rhabdomyosarcoma, and pleomorphic rhabdomyosarcoma. Recent studies have significantly impacted this classification with the emergence of three distinct new subtypes of rhabdomyosarcomas, namely rhabdomyosarcoma with MYOD1 mutations, rhabdomyosarcoma with TFCP2 fusions, and rhabdomyosarcoma with VGLL2/NCOA2 fusions. Although all these tumors share the terminology "rhabdomyosarcoma," their morphology, clinical behavior, and underlying molecular alterations are dramatically different. Finally, the presence of a rhabdomyoblastic phenotype within a tumor is by no means a diagnostic of a rhabdomyosarcoma, as this may be seen in many other mesenchymal malignancies, such as mesenchymal chondrosarcomas, malignant peripheral nerve sheaths tumors, and biphenotypic sinonasal sarcomas. In this review, we present the main clinical, morphological, and molecular features of these tumors and discuss the evolution of the current classification.

The myogenic differentiation 1 gene (MYOD1) p.L122R somatic mutation was first discovered in a subset of clinically aggressive embryonal rhabdomyosarcomas and has since been described in both pediatric and adult spindle cell/sclerosing rhabdomyosarcomas. Relatively little is known about the clinical, molecular, and histopathological features of these tumors in children. In order to further characterize the genomic and clinical features of pediatric MYOD1-mutant sarcomas, we evaluated a cohort of soft-tissue sarcoma patients treated at Texas Children's Hospital. Tumor DNA was subjected to next-generation panel sequencing and/or Sanger sequencing of the MYOD1 hotspot mutation. The MYOD1 p.L122R mutation was identified in six tumors, with a variant allele fraction greater than 0.8 in three cases, suggestive of loss of heterozygosity. One sclerosing rhabdomyosarcoma lacking the MYOD1 hotspot mutation was observed to have a MYOD1 copy number gain, also with evidence of loss of heterozygosity. Cancer gene panel sequencing revealed potentially targetable alterations in six of seven (86%) patients with MYOD1 alterations, including four patients with an alteration in the PI3K-AKT pathway: two hotspot PIK3CA mutations and deletions in PTEN and TSC2. On histopathologic review, MYOD1-altered tumors exhibited spindle and/or round cells and varying degrees of hyaline sclerosis. At last follow-up, six patients had died of disease and the seventh progressed early and was subsequently lost to follow-up. Both pre- and post-therapy patient-derived xenograft models were generated from one patient's tumor. These models were confirmed to harbor the MYOD1 and PIK3CA mutations seen in the primary tumor and were shown to be sensitive to PI3K/mTOR inhibition in vitro and in vivo. In conclusion, this study adds to recent reports describing the clinicopathologic and genomic features of MYOD1-altered soft-tissue sarcomas in children, including dismal prognosis and potential molecular targets for therapy. The novel preclinical models developed will facilitate further biological and preclinical study of this rare and aggressive tumor. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Spindle cell/sclerosing rhabdomyosarcoma is a malignancy classified among skeletal muscle tumors. It is also known as spindle cell rhabdomyosarcoma and sclerosing rhabdomyosarcoma.

Background Ewing sarcoma (ES) is a round cell sarcoma that can be challenging to diagnose on cytologic material given its significant overlap with numerous mesenchymal, epithelial, and lymphoid cytomorphologic mimics. The objective of this study was to assess the utility of a novel marker, NKX2.2, in the diagnosis of ES in cytologic material and its ability to distinguish ES from its mimics. Methods NKX2.2 immunohistochemistry was performed on cell blocks from 107 fine-needle aspirations, and nuclear expression was scored semiquantitatively for extent and intensity. The study cohort included ES (n = 10), well differentiated neuroendocrine tumor (n = 20), melanoma (n = 11), Merkel cell carcinoma (n = 10), small cell carcinoma (n = 10), alveolar rhabdomyosarcoma (n = 2), spindle cell/sclerosing rhabdomyosarcoma (n = 2), synovial sarcoma (n = 12), solitary fibrous tumor (n = 2), chronic lymphocytic leukemia (n = 10), lymphoblastic lymphoma (n = 11), adenoid cystic carcinoma (n = 6), and CIC-rearranged sarcoma (n = 1). Results NKX2.2 had high sensitivity (100%) and moderate specificity (85%) for the diagnosis of ES in cytologic material. NKX2.2 expression also was present in a subset of mesenchymal and epithelial mimics, and staining was most commonly observed in small cell carcinoma (80%) and well differentiated neuroendocrine tumor (45%). Among mesenchymal mimics, 42% exhibited NKX2.2 expression. NKX2.2 staining was absent in melanoma, adenoid cystic carcinoma, and lymphoproliferative neoplasms. Conclusions NKX2.2 is a highly sensitive but only moderately specific marker for ES. Neuroendocrine neoplasms exhibit variable NKX2.2 expression and remain a significant potential diagnostic pitfall. Thus, NKX2.2 expression should be interpreted in the context of an appropriate immunohistochemical panel (and often with confirmatory molecular testing) for the accurate diagnosis of ES.

Sclerosing/spindle cell rhabdomyosarcoma (SRMS) is a rare type of rhabdomyosarcoma. We immunohistochemically and genetically examined a case of SRMS of the oral floor. The patient was a 19-year-old female, who noticed swelling in the right oral floor. Under a diagnosis of a sublingual tumor, tumor resection was performed. The tumor consisted of atypical round to short spindle-shaped cells that exhibited a hyalinized stroma and a trabecular growth pattern. The tumor cells were diffusely positive for vimentin, MyoD1, and bcl-2, whereas they were partially positive for desmin, cytokeratin (CK)7, CK5/6, and MDM2. They were negative for pan-CK, S-100 protein, and CD99(MIC2). Fluorescence in situ hybridization analysis did not detect any FOXO1A or NCOA2 gene split signals. The cancer cells harbored a point mutation in codon 122 of the MYOD1 gene. This case was finally diagnosed as SRMS. We discuss the differentiation of SRMS from salivary gland tumors. Keywords: Sclerosing rhabdomyosarcoma, Oral cavity, Cytokeratin, FISH, MYOD1 gene

Aims Rhabdomyosarcomas of bone are extremely rare, with fewer than 10 reported cases. A very rare subtype of spindle cell/sclerosing rhabdomyosarcoma harbouring a FUS-TFCP2 fusion and involving both soft tissue and bone locations has been reported very recently. We report only the fourth case of this unusual, clinically aggressive rhabdomyosarcoma. Material and results A previously well 72-year-old male presented with a destructive lesion of the mandible. Morphological and immunohistochemical study of a needle biopsy and the subsequent resection showed a spindle cell rhabdomyosarcoma. RNA-seq, RT-PCR and FISH confirmed the presence of the FUS-TFCP2 fusion. Conclusions Spindle cell rhabdomyosarcomas carrying the FUS-TFCP2 fusion are very rare rhabdomyosarcoma variants with osseous predilection. The classification and differential diagnosis of this unusual molecular variant of spindle cell/sclerosing rhabdomyosarcoma are discussed.

Background Despite extensive immunohistochemical (IHC) and molecular studies combined with morphologic findings, a group of round/ovoid cell tumors histologically similar to Ewing sarcomas (ES) but lacking EWSR1-rearrangements may remain unclassifiable. Design We retrospectively analyzed 41 Ewing-like tumors (formalin-fixed, paraffin-embedded) previously determined as negative or non-informative for EWSR1-rearrangements by FISH and/or RT-PCR. A new histopathology revision and additional IHC and molecular analyses were carried out in order to investigate whether additional IHC and/or molecular testing in combination with the morphological findings may help in reaching a definitive diagnosis. Results Almost all the tumors (n = 40) involved soft tissue and/or bone and half the patients died of disease. In the archival cases all diagnoses were Ewing sarcoma (ES), Ewing-like sarcoma (ELS), myoepithelial tumor and undifferentiated sarcoma (US). In the new review all the tumors were re-classified as, ES (n = 16), Ewing-like tumor with EWSR1 rearrangement and amplification and possible EWSR1-NFATC2 gene fusion (n = 1), CIC-rearranged sarcomas or undifferentiated sarcoma, most consistent with CIC-rearranged sarcoma (n = 7), sarcoma with BCOR-alteration or undifferentiated sarcoma, consistent with BCOR-associated sarcoma (n = 3), neuroblastoma (n = 2), unclassifiable neoplasm with neuroblastic differentiation (n = 1), malignant rhabdoid tumor (n = 2), lymphoblastic lymphoma (n = 1), clear cell sarcoma of the gastrointestinal tract (n = 1), small cell carcinoma (n = 1), sclerosing rhabdomyosarcoma (n = 1), desmoplastic small round cell tumor (n = 1), malignant peripheral sheath nerve tumor (n = 1), poorly-differentiated synovial sarcoma (n = 1), Possible gastrointestinal stromal tumor/GIST with predominant round cells (n = 1) and possible SMARCA4-deficient-sarcoma (n = 1). NKX2.2, ETV4 and BCOR immunoreactivity was observed in all ES, CIC-rearranged sarcomas and sarcomas with BCOR alteration, respectively. CIC-rearrangement by FISH was observed in many of the CIC-rearranged sarcomas. Conclusion Our analysis of 41 Ewing-like tumors confirms that there may be a significant pathological and IHC overlap among Ewing-like tumors, with prognostic and therapeutic impacts. Additional IHC (NKX2.2, ETV4 and BCOR) and molecular studies including FUS, CIC or BCOR analysis may support the final diagnosis when FISH or RT-PCR fail to detect EWSR1-rearrangements. Any molecular findings should always be interpreted in relation to the specific clinical and pathological context.

Background Spindle cell/sclerosing rhabdomyosarcoma is a genomically heterogeneous, uncommon subtype of rhabdomyosarcoma, particularly rare in adults. Its MYOD1-mutant variant is aggressive irrespective of age. Cytogenetic data on spindle cell/sclerosing rhabdomyosarcoma are sparse and disparate. Materials and methods Cytogenetic and molecular analyses were performed on an adult sclerosing rhabdomyosarcoma. Results The karyotype of the sclerosing rhabdomyosarcoma displayed clonal evolution corresponding to two hyperdiploid clones: 48,XY,+i(19)(p10),+22/48,idem,der(9)t(2;9)(q21~22;p21). The changes were gain of chromosome 19 with the overrepresentation of 19p arm, gain of chromosome 22, gain of the 2q arm, and loss of 9p21. Mutation analysis revealed a homozygous c.T365G (p.L122R) mutation of the MYOD1 gene, but none of PIK3CA. Conclusion To our knowledge, this is the first adult MYOD1-mutant sclerosing rhabdomyosarcoma studied cytogenetically. The only other reported sclerosing rhabdomyosarcoma with MYOD1 mutation and abnormal karyotype was pediatric. Since these tumors are highly aggressive, further studies unravelling their cytogenetic and molecular characteristics are warranted.

A man in his 30s presented with back pain and was found to have a tumour in the posterior chest wall. Imaging showed an expansile mass measuring up to 6.7 cm, centred on the left posterior 11th rib with diffuse cortical destruction. The tumour displaced the left hemidiaphragm, and was in contact with the spleen, with no infiltration seen. There was no involvement of the overlying skin. PET-CT showed the lesion to be inhomogeneously hypermetabolic, with a mainly peripheral distribution of FDG. Staging imaging showed no other abnormality. The patient was otherwise well with no medical history and no significant family history. A CT-guided biopsy was performed. Review the high-quality, interactive digital Aperio slide at http://virtualacp.com/JCPCases/jclinpath-2019-206338/ and consider your diagnosis. 1. Metastatic carcinoma. 2. Alveolar rhabdomyosarcoma (ARMS). 3. Sclerosing epithelioid fibrosarcoma (SEF). 4. Spindle cell/sclerosing rhabdomyosarcoma. 5. Osteosarcoma. 6. Ossifying fibromyxoid tumour (OFMT). This is a tumour composed of relatively uniform, minimally to mildly pleomorphic, medium-sized cells with round to ovoid nuclei and clear to lightly eosinophilic cytoplasm. The cells are present in groups, nests and occasionally individually, within densely fibrous stroma, with areas suspicious of osteoid formation. In this case the tumour is diffusely and strongly positive for MUC4 and CD99, and focally for CD57. This particular case is negative for AE1/AE3, Cam5.2, CK7, TTF1, desmin, smooth muscle actin (SMA), myogenin, FLI1, neuron-specific enolase, CD56, synaptophysin, chromogranin, glial fibrillary acidic protein, TLE1, CD45 and CD138. There is a normal staining pattern for p53, and INI expression is retained in nuclei. …

Spindle cell and sclerosing rhabdomyosarcoma (ssRMS) is a rare variant of rhabdomyosarcoma, which includes three distinct subtypes. In infants, these tumors are commonly associated with recurring fusions involving VGLL2 or NCOA2 and have a favorable prognosis. We present four cases of ssRMS and 16 additional cases from the literature, which show that these patients present with localized disease and have an excellent prognosis regardless of surgical margin or lack of radiation therapy. Molecularly defined spindle cell rhabdomyosarcoma in infants is likely a biologically distinct entity which may not require the aggressive multimodal treatment used for other subtypes of rhabdomyosarcoma.

Background Primary breast sarcoma (PBS) is rare, comprising approximately 1% of breast malignancies. Rhabdomyosarcoma (RMS) accounts for an extremely small proportion of PBSs, often leading to delayed histologic confirmation. Methods Upon reviewing Asan Medical Center's pathology database between 2000 and 2018, 41 PBS cases were retrieved, including three cases of primary RMS of the breast. Their clinicopathological features were analyzed, and the literature related to PBS and primary RMS of the breast was reviewed. Results We identified three primary breast RMS cases from our institution database, comprising 7.3% of PBS: one case each of spindle cell/sclerosing RMS (ssRMS), alveolar RMS (aRMS), and embryonal RMS (eRMS). All cases involved adolescents or young adults (14, 16, and 25 years, respectively) who underwent mastectomy or radiotherapy and were confirmed using immunohistochemical testing for myogenin, desmin, and myogenic differentiation. The ssRMS patient experienced recurrence at the operation site 4 months post-surgery despite undergoing concurrent chemoradiotherapy. The aRMS patient had multiple metastases at diagnosis and showed FAX3-FOXO1 fusion transcripts; she died 22 months after the diagnosis. The eRMS patient had enlarged axillary lymph nodes; post-radiotherapy, the lesion recurred as multiple metastases to the bone and lung. She died 18 months post-diagnosis. Conclusions Our experience on RMS cases suggests that spindle cell or small round cell malignancy in breasts of young female should raise suspicion for the possibility of primary or secondary RMS. To our knowledge, this is the second report of primary breast ssRMS and it may help clinicians who encounter this rare disease in the future.

Based on their distinctive histologic and genetic features, the latest WHO classification of soft tissue tumors includes four pathologic variants of rhabdomyosarcoma (RMS): embryonal (ERMS), alveolar (ARMS), spindle cell-sclerosing (SRMS-ScRMS) and pleomorphic RMS. The aim of this study focused on a detailed clinicopathologic and survival analysis of head and neck RMS (HNRMS) using the latest pathologic and molecular criteria reflecting this new subclassification in a large cohort.Patients managed for HNRMS in our institution (1996-2015) were analyzed. The presence of a FOXO1 fusion was required for the classification of ARMS. MYOD1 mutations in SRMS-ScRMS were tested when material available. Univariate and multivariate analyses were performed to evaluate variables related to overall survival (OS).Ninety-nine HNRMS patients (52 males and 47 females, mean of 16years) were included in the study after pathologic re-review. The most common location was parameningeal (PM) (n=64), followed by non-orbital/non-PM (n=25) and orbital (n=10). There were 53 ERMS, 33 fusion-positive ARMS and 13 SRMS-ScRMS [SRMS (8); ScRMS (5)]. The 5-year OS rate for ERMS patients was significantly higher (82%) compared to ARMS (53%) and SRMS-ScRMS (50%) [SRMS (75%); ScRMS (30%)]. Univariate analysis showed that survival was dependent on histology (P=0.012), tumor size5cm (P0.001), regional lymph node involvement (P=0.002), metastasis at initial presentation (P0.001), stage (P0.001), and recurrence (P=0.002). Multivariate analysis confirmed histologic subtype to be significant (P=0.043).Our findings reinforce that HNRMS is a heterogenous disease with ARMS and SRMS-ScRMS having an equally unfavorable outcome.

Recurrent mutations in the myogenic transcription factor MYOD1 and PIK3CA were initially described in a subset of embryonal rhabdomyosarcomas. Recently, two independent studies demonstrated presence of MYODI (L122R) mutations as the basis to re-classify a spindle cell rhabdomyosarcoma, along with a sclerosing rhabdomyosarcoma, distinct from an embryonal rhabdomyosarcoma. We analyzed a much larger cohort of 49 primary rhabdomyosarcoma tumor samples of various subtypes, collected over a period of 9 years, for the presence of MYOD1 (L122R), PIK3CA (H1047), and PIK3CA (E542/E545) mutations, along with immunohistochemical analysis of desmin, myogenin, and MYOD1. Although activating PIK3CA mutations were absent across the sample set analyzed, we report 20% MYOD1 (L122R) mutation in rhabdomyosarcomas, found exclusively in 10 of 21 spindle cell and sclerosing rhabdomyosarcomas, occurring mostly in the head and neck region along with extremity sites (64%), than the paratesticular and intra-abdominal sites. Furthermore, while all 10 MYOD1 mutant spindle cell and sclerosing rhabdomyosarcoma samples showed diffuse and strong MYOD1 immunoexpression, 7 of 31 samples of rhabdomyosarcoma with wild-type MYOD1 were negative for MYOD1 expression. Clinically, a striking correlation was found between MYOD1 mutation and the clinical outcomes available for 15 of 21 cases: 5 of 7 patients with spindle cell and sclerosing rhabdomyosarcomas, harboring MYOD1 mutation, were alive-with-disease and 2 of 8 patients with spindle cell and sclerosing rhabdomyosarcomas, with mutant MYOD1, were free-of-disease. Taken together, we present the first report of MYOD1 (L122R) mutation in the largest cohort of 49 rhabdomyosarcomas reported so far, that are associated with a relatively aggressive clinical course. Moreover, consistent with the earlier two studies, this study further reinforces a relationship between spindle cell and the sclerosing rhabdomyosarcoma—now recognized as a single subtype, distinct from an embryonal rhabdomyosarcoma.

Sclerosing and spindle cell rhabdomyosarcoma is a rare histologic subtype, designated in the latest WHO classification as a stand-alone pathologic entity. Three genomic groups have been defined: an infantile subset of spindle cell rhabdomyosarcoma harboring VGLL2-related gene fusions, a MYOD1-mutant subset commonly associated with sclerosing morphology, and a subset lacking recurrent genetic abnormalities. In this study, we focus on MYOD1-mutant rhabdomyosarcoma to further define their clinicopathologic characteristics and behavior in a larger patient cohort. We investigated 30 cases of MYOD1-mutant rhabdomyosarcoma (12 previously reported and 18 newly diagnosed) with an age range of 2-94 years, including 15 children. All cases showed morphology within the spectrum of spindle cell/sclerosing rhabdomyosarcoma (8 cases showing pure sclerosing morphology, 8 cases showing pure spindle cell morphology and 14 cases showing a hybrid phenotype of spindle, sclerosing and primitive undifferentiated areas). All tumors harbored either homozygous or heterozygous MYOD1 (p.L122R) exon 1 mutations. In 10 (33%) cases, a co-existent PIK3CA mutation was identified. Hot-spot mutations in NRAS and HRAS were each identified in a single case, respectively. Follow-up was available on 22 (73%) patients with a median duration of 28 months. Local recurrence was seen in 12 (55%) and distant recurrence in 12 (55%) cases, despite multimodality chemoradiation therapy. At last follow-up, 15 (68%) patients died of the disease, one patient was alive with disease and five had no evidence of disease. The prognosis was equally poor in pediatric and adult patients. In conclusion, MYOD1 mutation defines an aggressive rhabdomyosarcoma subset, with poor outcome and response to therapy, irrespective of age. Given that this distinct molecular subtype is characterized by an aggressive biologic behavior compared to other genetic subtypes of spindle and sclerosing rhabdomyosarcoma, the MYOD1 genotype should be used as a molecular marker in both subclassification and prognostication of rhabdomyosarcoma.

Sclerosing rhabdomyosarcoma (RMS) is a rare subtype of RMS with unique prominent stromal hyalinization and a pseudovascular architecture. It overlaps morphologically with spindle cell RMS and poses both diagnostic and therapeutic challenges because of its rarity and aggressive clinical course. In this article, we report a case of sclerosing RMS arising from a prior craniotomy site, which demonstrated both sclerosing and spindle cell components. A literature review of RMS with sclerosing morphology identified 122 cases. Our review documents the following: sclerosing RMS occurs in both childhood and adult populations, has a predilection for the head and neck areas, and has a worse prognosis in adults. Sclerosing RMS harbors a high frequency of MYOD1 mutations, conferring a poor clinical outcome. Sclerosing RMS and spindle RMS likely represent a morphologic spectrum of one entity.

The diagnosis of mesenchymal chondrosarcoma, a distinctive biphasic malignant neoplasm harboring the HEY1-NCOA2 gene fusion and consisting of primitive round to spindled cells admixed with foci of relatively mature hyaline cartilage, is usually straightforward by morphologic evaluation alone. However, in the setting of a limited biopsy, specimens lacking cartilage generate a broad differential diagnosis, encompassing a variety of other primitive sarcomas, including spindle cell/sclerosing rhabdomyosarcoma. Although a small number of cases of mesenchymal chondrosarcoma with aberrant skeletal muscle marker expression have been reported, pathologists are largely unaware of this potential diagnostic pitfall. We report 6 additional cases of mesenchymal chondrosarcoma showing expression of multiple skeletal muscle markers, including one case initially misdiagnosed as "spindle cell/sclerosing rhabdomyosarcoma" on needle biopsy. Awareness of this phenomenon and judicious application of molecular diagnostic testing for the HEY1-NCOA2 fusion are critical to avoid misclassification of mesenchymal chondrosarcoma as rhabdomyosarcoma, with potentially adverse patient impact.

Neural, sclerosing, and myofibroblastic lesions of the ear and temporal bone present diagnostic challenges for both clinicians and pathologists due to significant overlap in their clinical presentations, histologic appearances, and immunohistochemical profiles. While some of these lesions, such as schwannomas, are relatively common, others are rendered even more difficult because they are encountered very rarely in routine surgical pathology practice. This review is intended to provide an update on the pathology of some of the most commonly encountered primary diagnostic entities for the ear and temporal bone, and includes the following neural lesions: schwannoma, meningioma, and encephalocele/meningocele. Sclerosing lesions that will be discussed include spindle cell and sclerosing rhabdomyosarcoma, sclerosing epithelioid fibrosarcoma, and sclerosing paraganglioma. Finally, myofibroblastic lesions that will be reviewed are nodular fasciitis, IgG4-related disease, and solitary fibrous tumor. For each of these lesions, the differential diagnosis and useful ancillary tests will be discussed in the context of a broad range of additional primary and secondary lesions.

Spindle cell/sclerosing rhabdomyosarcoma (ssRMS) is a new subtype of rhabdomyosarcoma included in the World Health Organization soft tissue and bone tumor classification in 2013. Despite the increasing number of reported cases of ssRMS, the imaging characteristics of ssRMS are not established. Herein, we present the case of an elderly Japanese woman with ssRMS of the masticator space with intracranial invasion without destruction of the adjacent bone. Attention should be paid to the presence of intracranial infiltration that may indicate a worse prognosis. Tumor growth without bone destruction could be a key finding to differentiate ssRMSs from conventional subtypes of rhabdomyosarcoma.

Spindle cell/sclerosing rhabdomyosarcoma (S-ScRMS) was recently recognized in 2013 by the World Health Organization (WHO) as a stand-alone entity (Parham et al., WHO classification of tumours of soft tissue and bone, IARC Press, Lyon, 2013). Historically, the spindle cell and sclerosing variants were subcategorized under embryonal type rhabdomyosarcoma. Current data supports that certain S-ScRMS cases have a more aggressive clinical course with reduction of long-term survival, and those found in the head and neck region often exhibit extensive local recurrence. Furthermore, due to variable histopathologic appearances and immunohistochemical findings, misdiagnosis is common. We aim to report the variability in histopathologic patterns, immunohistochemical findings, radiographic features, and clinical data on three new cases of S-ScRMS presenting in the oral cavity of young adult males (ages 22, 24, and 39 years). The English-language literature on S-ScRMS is briefly reviewed as well.

Background Clinical characteristics and optimal treatment strategies for spindle cell/sclerosing rhabdomyosarcoma (ssRMS) have not been well established because of its rarity. Procedure Retrospective re-evaluation of sarcoma specimens (1997–2014) identified 16 ssRMSs (median age 20 years, range 7–39 years). Clinicopathological features, clinical course, and outcome were analyzed. Results Primary disease sites were the head and neck (10 cases) and other regions (6 cases). Nine cases were at Intergroup Rhabdomyosarcoma Study preoperative stage 3. The primary tumors were >5 cm in 13 cases. Two patients had lymph node metastases, but none had distant metastases at presentation. At follow-up (median period 39 months, range 4.6–201), seven patients were alive without disease. Among nine patients treated with the vincristine, actinomycin, and cyclophosphamide (VAC) regimen, five responded well, with four surviving free of disease. Among ten patients with recurrent or progressive disease, three experienced local recurrence, four had distant metastases, and three had both. None exhibited bone marrow invasion. Eight of the ten patients died in median time from relapse to death of 18 months (range 11–56). Conclusions Although most ssRMSs present as a bulky tumor, nodal or distant metastases are rare at presentation. ssRMSs initially show good response to VAC, but >50% of tumors recur or progress; these data suggest a worse prognosis of ssRMS compared to the pediatric embryonal variant. As relapse typically occurs as local or distant solitary lesion without bone marrow invasion, localized treatment combined with chemotherapy would contribute to improve the prognosis of recurrent ssRMS. Pediatr Blood Cancer 2015;62:1011–1016. © 2014 Wiley Periodicals, Inc.

Sclerosing and spindle cell rhabdomyosarcoma (RMS) are rare types of RMS recently reclassified as a stand-alone pathologic entity, separate from embryonal RMS (ERMS). Although sclerosing and spindle cell RMS share clinical and morphologic features, a pathogenetic link based on shared molecular alterations has not been established. Spindle cell RMS in children have been associated with a less aggressive clinical course compared to adults. Recently, recurrent MYOD1 mutations were described in 44% of adult spindle cell RMS, but no pediatric tumors or sclerosing RMS were studied for comparison. Thus, we investigated 16 RMS (5 sclerosing and 11 spindle cell) in children and adults for the presence of MYOD1 mutations by targeted Polymerase Chain Reaction (PCR). Remarkably, all 5 sclerosing RMS and 4 of 11 spindle cell RMS showed the MYOD1 p.L122R hot-spot mutation. Of the five pediatric tumors, 2/2 sclerosing RMS and 2/3 spindle cell RMS showed MYOD1 mutations. Three of nine MYOD1-mutant RMS showed coexistent PIK3CA mutations, while no MDM2 amplifications were identified. All four pediatric MYOD1-mutated RMS patients died of the disease at 12-35 months following diagnosis. In conclusion, spindle cell and sclerosing RMS show recurrent MYOD1 mutations, in keeping with a single pathologic entity, regardless of age at presentation. This group however, is distinct from the infantile RMS associated with NCOA2 fusions. Although our study suggests that pediatric MYOD1-mutant RMS follow an aggressive behavior with high mortality, further studies are required to confirm this finding.

Rhabdomyosarcoma, a cancer of skeletal muscle lineage, is the most common soft-tissue sarcoma in children. Major subtypes of rhabdomyosarcoma include alveolar (ARMS) and embryonal (ERMS) tumors. Whereas ARMS tumors typically contain translocations generating PAX3-FOXO1 or PAX7-FOXO1 fusions that block terminal myogenic differentiation, no functionally comparable genetic event has been found in ERMS tumors. Here we report the discovery, through whole-exome sequencing, of a recurrent somatic mutation encoding p.Leu122Arg in the myogenic transcription factor MYOD1 in a distinct subset of ERMS tumors with poor outcomes that also often contain mutations altering PI3K-AKT pathway components. Previous mutagenesis studies had shown that MYOD1 with a p.Leu122Arg substitution can block wild-type MYOD1 function and bind to MYC consensus sequences, suggesting a possible switch from differentiation to proliferation. Our functional data now confirm this prediction. Thus, MYOD1 p.Leu122Arg defines a subset of rhabdomyosarcomas eligible for high-risk protocols and the development of targeted therapeutics.

Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma in children and adolescents, being characterized by expression of genes and morphological and ultrastructural features of sarcomeric differentiation. The spindle cell variant of rhabdomyosarcoma (spindle cell RMS) in adults has been defined as an entity, separated from embryonal rhabdomyosarcoma (ERMS), with unfavourable clinical outcome. So far, no recurrent genetic alteration has been identified in the adult form of spindle cell RMS. We studied a case of adult spindle cell RMS using next-generation sequencing (NGS) after exome capture. Using this approach, we identified 31 tumour-specific somatic alterations and selected four genes with predicted functional relevance to muscle differentiation and growth. MYOD1, KIF18A, NOTCH1, and EML5 were further tested for mutations using Sanger sequencing on DNA from FFPE samples from 16 additional, adult spindle cell RMS samples. The highly conserved sequence homology of MYOD1 with other myogenic transcription factors prompted us to screen the basic DNA-binding domains of MYF5, MYF6 and MYOG for mutations. From the investigated 17 samples, seven (41%) showed homozygous mutation of MYOD1, indicating a critical role in this rare subtype of adult spindle cell RMS, while no mutations were found in any of the other genes involved in myogenic differentiation. The p.L122R mutation occurs in the conserved DNA binding domain in MYOD1 and leads to transactivation and MYC-like functions. MYOD1 homozygous mutations are frequent, recurrent and pathognomonic events in adult-type spindle cell RMS.

Sclerosing rhabdomyosarcoma (SRMS) is a newly recognized and rare variant of rhabdomyosarcoma. This soft tissue tumor has not yet been reported as a thoracic lesion. We report a case of a 26-year-old woman who presented with a large chest wall tumor. The tumor originated from the right anterior chest wall and protruded into the intra- and extrapleural cavity. A transcutaneous needle biopsy revealed spindle cells in an abundant hyalinized and fibrous stroma. Although the tumor was considered as a malignant soft-tissue neoplasm, a definitive diagnosis could not be established. A wide excision of the chest wall including the second, third and fourth rib and a part of sternum was performed. Histologically, cytoplasmic cross-striations were found in a portion of the tumor cells. The tumor cells were positive for muscle markers, and the tumor was diagnosed as rhabdomyosarcoma consistent with a sclerosing type of rhabdomyosarcoma. Eighteen months after the complete resection, the patient has pleural disseminations but is alive and undergoing chemotherapy. This case highlights the histologic features of a rare form of rhabdomyosarcoma, and emphasizes the importance of awareness of its existence and the utility of skeletal muscle markers in distinguishing sclerosing rhabdomyosarcoma from its mimics.

Previously unrecognized but clinicopathologically (and often molecularly) distinct types of soft tissue tumor continue to be characterized, allowing wider recognition, more consistent application of diagnostic criteria, more reliable prediction of tumor behavior and enhancement of existing classification schemes. Examples of such 'entities' that have become much better understood over the past decade or so include deep 'benign' fibrous histiocytoma, hemosiderotic fibrolipomatous tumor, PEComa, spindle cell liposarcoma, myoepithelial tumors of soft tissue and spindle cell/sclerosing rhabdomyosarcoma. These tumor types, as well as the insights which they have engendered, are briefly reviewed here.

Spindle cell rhabdomyosarcoma (RMS) is a rare form of RMS with different clinical characteristics and behavior between children and adult patients. Its genetic hallmark remains unknown and it remains debatable if there is pathogenetic relationship between the spindle cell and the so-called sclerosing RMS. We studied two pediatric and one adult spindle cell RMS by next generation RNA sequencing and used FusionSeq for data analysis to detect novel fusions. An SRF-NCOA2 gene fusion was detected in a spindle cell RMS from the posterior neck in a 7 month-old child. The fusion matched the tumor karyotype and was further confirmed by fluorescence in situ hybridization (FISH) and by RT-PCR, which showed fusion of SRF exon 6 to NCOA2 exon 12. Additional 14 spindle cell (from 8 children and 6 adults) and 4 sclerosing (from 2 children and 2 adults) RMS were tested by FISH for the presence of abnormalities in NCOA2, SRF, as well as for PAX3 and NCOA1, identifying NCOA2 rearrangements in two additional spindle cell RMS from a 3 month-old and a 4 week-old child, both arising in the chest wall. In the latter tumor, TEAD1 was identified by rapid amplification of cDNA ends (RACE) to be the NCOA2 gene fusion partner. None of the adult tumors were positive for NCOA2 rearrangement. Despite similar histomorphology in adults and young children, these results suggest that spindle cell RMS is a heterogeneous disease genetically as well as clinically. Our findings also support a relationship between NCOA2-rearranged spindle cell RMS occurring in young childhood and the so-called congenital RMS, which often displays rearrangements at 8q13 locus (NCOA2).

Since the first description of sclerosing rhabdomyosarcoma in 2000, 19 pediatric cases have been reported in the literature. However, it is debated whether sclerosing rhabdomyosarcoma represents a specific rhabdomyosarcoma entity or a variant of embryonal or alveolar rhabdomyosarcoma. To date, 6 sclerosing rhabdomyosarcoma karyotypes and 1 sclerosing rhabdomyosarcoma comparative genomic hybridization profile have been reported. We present the first whole-genome tumoral genotyping of a sclerosing rhabdomyosarcoma by high-density single nucleotide polymorphism array. The single nucleotide polymorphism genotyping revealed a complex pattern including gains and losses of whole chromosomes and an amplification of the 12q13-15 region. Amplification of the 12q13-q15 region containing SAS, GLI, CDK4, and MDM2 has been observed in rhabdomyosarcoma. In the present case, the 2 amplified target genes were MDM2 and HMGA2, excluding CDK4. The identification of a specific MDM2-HGMA2 amplicon excluding CDK4 has only been described so far in well-differentiated and dedifferentiated liposarcoma. Further studies are needed to assess if this anomaly is a specific marker of sclerosing rhabdomyosarcoma.

Sclerosing rhabdomyosarcoma (ScRMS) and spindle cell rhabdomyosarcoma (SRMS) have been recently reclassified as a stand-alone pathologic entity, separate from embryonal RMS. Genetically, a subset of the congenital cases display NCOA2 gene rearrangements, whereas tumors occurring in older children or adults harbor MYOD1 gene mutations with or without coexisting PIK3CA mutations. Despite these recent advances, a significant number of tumors lack known genetic alterations. In this study we sought to investigate a large group of pediatric SRMS/ScRMS, spanning a diverse clinical and pathologic spectrum, by using a combined fluorescence in situ hybridization, targeted DNA, and whole-transcriptome sequencing methodology for a more definitive molecular classification. A total of 26 SRMS and ScRMS cases were selected from the 2 participating institutions for the molecular analysis. Ten of the 11 congenital/infantile SRMS showed recurrent fusion genes: with novel VGLL2 rearrangements seen in 7 (63%), including VGLL2-CITED2 fusion in 4 and VGLL2-NCOA2 in 2 cases. Three (27%) cases harbored the previously described NCOA2 gene fusions, including TEAD1-NCOA2 in 2 and SRF-NCOA2 in 1. All fusion-positive congenital/infantile SRMS patients with available long-term follow-up were alive and well, none developing distant metastases. Among the remaining 15 SRMS patients older than 1 year, 10 (67%) showed MYOD1 L122R mutations, most of them following a fatal outcome despite an aggressive multimodality treatment. All 4 cases harboring coexisting MYOD1/PIK3CA mutations shared sclerosing morphology. All 5 fusion/mutation-negative SRMS cases presented as intra-abdominal or paratesticular lesions.

Sclerosing (pseudovascular) rhabdomyosarcoma in adults has been described as a rare variant of rhabdomyosarcoma characterized by extensive hyaline fibrosis and pseudovascular growth patterns. We describe another case of this rhabdomyosarcoma subtype including ultrastructural and genetic findings-the lesion presented in a 62-year-old male patient in the left lower leg. The tumor was located within the deep soft tissue with maximum diameter of 11.8 cm and skin ulceration. Ultrastructural analysis revealed irregularly distributed disorganized filaments without clear evidence of Z-bands and a richly collagenized matrix. Using comparative genomic hybridization, a sharply delineated loss of chromosomal region 10q22, loss of chromosome Y, and a gain of chromosome 18 (trisomy) were detected. Reciprocal translocations t(1;13) and t(2;13)(q35;q14) which are characteristic of alveolar rhabdomyosarcoma could be excluded. These findings, while showing a relation to other rhabdomyosarcoma subtypes, represent a relatively circumscribed genetic defect pattern in sclerosing (pseudovascular) rhabdomyosarcoma that is somewhat different from patterns described in most other rhabdomyosarcoma subtypes. Six months after tumor resection, the patient presented with metastatic disease. Further studies should concentrate on the identification of genes especially on chromosomal region 10q22 to elucidate more aspects in the pathogenesis of this rhabdomyosarcoma subtype.