Ultrastructure in resolving a diagnosis of poorly differentiated clear cell sarcoma of soft parts in an adolescent male.

Clear cell sarcoma of soft parts is a rare tumor in children and it requires a high index of suspicion for accurate diagnosis. Early diagnosis leads to radical surgical excision and limits the aggressive behavior of this tumor. We report a case of a 12-year-old boy with a recurrent soft-tissue tumor in the scalp, misdiagnosed on three occasions as epitheloid sarcoma owing to the poorly differentiated appearance of cells. In spite of focal S-100 expression, this tumor was not recognized as a tumor of melanocytic origin till melanosomes were demonstrated on electron microscopy (EM). Detection of melanosomes on electron microscopy helped in clinching the histology diagnosis, reiterating the definite role of EM in diagnosing these tumors. Failure to accurately diagnose this tumor resulted in institution of preoperative chemotherapy, delayed surgical excision, tumor progression and death of patient within a year and half of presentation.

EWS-CREB1: a recurrent variant fusion in clear cell sarcoma--association with gastrointestinal location and absence of melanocytic differentiation.

PURPOSE: Clear cell sarcoma (CCS) usually arises in the lower extremities of young adults and is typically associated with a t(12;22) translocation resulting in the fusion of EWS (EWSR1) with ATF1, a gene encoding a member of the cyclic AMP-responsive element binding protein (CREB) family of transcription factors. CCS arising in the gastrointestinal tract is rare and its pathologic and molecular features are not well defined. EXPERIMENTAL DESIGN: We report a novel variant fusion of EWS to CREB1, a gene at 2q32 encoding another CREB family member highly related to ATF1, detected in three women with gastrointestinal CCS. All three cases contained an identical EWS-CREB1 fusion transcript that was shown by reverse transcription-PCR. In two of the cases tested, EWS gene rearrangement was also confirmed by fluorescence in situ hybridization and the EWS-CREB1 genomic junction fragments were isolated by long-range DNA PCR. RESULTS: Morphologically, all three tumors lacked melanin pigmentation. By immunohistochemistry, there was a strong and diffuse S100 protein reactivity, whereas all melanocytic markers were negative. Ultrastructurally, two of the cases lacked melanosomes. The melanocyte-specific transcript of MITF was absent in two cases, and only weakly expressed in the third case. The Affymetrix gene expression data available in one case showed lower expression of the melanocytic genes MITF, TYR, and TYRP1, compared with four EWS-ATF1-positive CCSs of non-gastrointestinal origin. CONCLUSIONS: EWS-CREB1 may define a novel subset of CCS that occurs preferentially in the gastrointestinal tract and shows little or no melanocytic differentiation. Thus, evidence of melanocytic lineage or differentiation is not a necessary feature of sarcomas with gene fusions involving CREB family members.

Association of the t(12;22)(q13;q12) EWS/ATF1 rearrangement with polyphenotypic round cell sarcoma of bone: a case report.

The t(12;22)(q13;q12) chromosomal rearrangement results in an EWS/ATF1 fusion transcript and is associated with clear cell sarcoma (CCS). CCS is an uncommon tumor arising in tendons and aponeuroses of the extremities and shows evidence of melanocytic differentiation at the light microscopic, immunohistochemical, and/or ultrastructural level. Only 5 cases have been reported to arise in bone, none of which had molecular confirmation of the diagnosis. The current report describes a 7-year-old girl with a primary round cell sarcoma of the left humerus showing polyphenotypic differentiation on immunohistochemical analysis. Antibodies directed at melanocytic antigens were negative, and there was no evidence of melanocytic differentiation by light microscopy or ultrastructural analysis. Cytogenetic analysis revealed rearrangement of the EWS locus within 22q12. RT-PCR and sequence analysis revealed the presence of a fusion transcript bringing together exon 7 of EWS with exon 5 of ATF1, consistent with a type 2 transcript reported in association with CCS. However, given the lack of morphologic features usually present in CCS, a diagnosis of polyphenotypic round cell sarcoma was made. This tumor thus expands the spectrum of neoplasms associated with the t(12;22)(q13;q12) rearrangement.

Clear cell sarcoma of the stomach.

AIM: Clear cell sarcoma is a high-grade sarcoma with morphological features resembling malignant melanoma. This tumour is reported to display a characteristic distribution pattern nearly always involving the extremities. We report the first case of clear cell sarcoma of the stomach. METHODS AND RESULTS: A 30-year-old male developed a huge tumour of the stomach, which at first glance could be considered as a poorly differentiated carcinoma. Immunohistochemical and ultrastructural examinations were consistent with a diagnosis of metastatic melanoma. However, cytogenetic examination revealed a t(12;22) translocation, specific for clear cell sarcoma. This was confirmed by fluorescence in-situ hybridization. CONCLUSION: Making a reliable diagnosis of clear cell sarcoma of the stomach requires cytogenetic or molecular diagnostic investigations, particularly to rule out metastatic melanoma. This diagnosis avoids an unnecessary search for a primary melanoma.

Clear-cell sarcoma diagnosis by fine-needle aspiration: cytologic, histologic, and ultrastructural features; potential pitfalls; and literature review.

A definitive diagnosis of clear-cell sarcoma of soft parts (CCSSP) is possible by fine-needle aspiration (FNA) biopsy alone. The aspirates are markedly cellular, consisting predominantly of discohesive cells but also of cohesive cells. The cytoplasm is eosinophilic and eccentric. The nuclei are round and contain macronucleoli. CCSSP should be considered when FNA of a soft-tissue tumor shows uncharacteristically high cellularity and relatively uniform cells with macronucleoli. Cohesion of some tumor cells does not rule out CCSSP. Melanin pigment and cytoplasmic clearing are infrequent and not necessary for the diagnosis. Sufficient material should always be procured for immunohistochemical studies on the cell block. Seven other cases are found in the literature, all correctly diagnosed by FNA. Although it is rare, CCSSP is a highly malignant tumor that can be diagnosed readily by FNA without resorting to incisional biopsy.

Molecular diagnosis of clear cell sarcoma: detection of EWS-ATF1 and MITF-M transcripts and histopathological and ultrastructural analysis of 12 cases.

Clear cell sarcoma (CCS), also known as melanoma of soft parts, is an uncommon deep soft tissue tumor presenting typically in the lower extremities of young adults. Previous cytogenetic studies have established the specificity of the recurrent t(12;22)(q13;q12), resulting in a EWS-ATF1 fusion, for CCS. The prevalence of the EWS-ATF1 fusion in CCS remains unclear, since most genetically confirmed CCS have been reported as isolated cytogenetic or molecular diagnostic case reports. We therefore studied histologically confirmed CCS from 12 patients for the presence of EWS-ATF1 by reverse-transcriptase polymerase chain reaction (RT-PCR), using RNA extracted from either frozen (four cases) or formalin-fixed paraffin-embedded (eight cases) material. All primary tumors were located in the deep soft tissues of the extremities. Histologically, 10 cases had a typical epithelioid nested appearance. Most or all cases showed immunostaining for HMB45 (12 of 12), S-100 protein (10 of 12), and MITF (12 of 12). Ultrastructural analysis showed melanosomes in six of seven cases. The presence of an EWS-ATF1 fusion transcript was identified by RT-PCR in 11 of 12 cases (91%), all of which showed the same fusion transcript structure, namely the previously described in-frame fusion of EWS exon 8 to ATF1 codon 65. RT-PCR analysis for the melanocyte-specific splice form of the MITF transcript was positive in all cases tested (4 of 4). These data confirm that EWS-ATF1 detection can be used as a highly sensitive diagnostic test for CCS and that CCS expresses the melanocyte-specific form of the MITF transcript, further supporting its genuine melanocytic differentiation.

Clear cell sarcoma of kidney in an adolescent and in young adults: a report of four cases with ultrastructural, immunohistochemical, and DNA flow cytometric analysis.

Clear cell sarcoma of the kidney is a distinct, highly malignant pediatric neoplasm. Its occurrence in adults is extremely rare and the subject of isolated case reports. We present a series of four cases (three males and one female) identified in an adolescent and in young adults (16, 18, 20, and 25 years) with flank mass (three cases), hematuria (two cases), flank pain (two cases), and hypertension (one case). Three patients had stage III disease and one had stage I disease (National Wilms' Tumor Study staging system). All tumors had predominantly or exclusively the classic histology of a monotonous proliferation of uniform small round cells with evenly distributed fine chromatin, although focal microcyst formation (two cases) and spindled architecture (one case) (variant patterns) were also noted. Therapy in all cases consisted of surgery and chemotherapy with or without radiation. Follow-up data (29-202 months) showed distant metastases in all four cases, including the lung (four cases), bone (two cases), and the liver (two cases). Three patients died of disease at 29, 59, and 63 months (mean, 50.3 months), and one patient is alive with no evidence of disease at 202 months. Ultrastructural features included scattered primitive junctions, short and irregular cytoplasmic extensions, and scant to a moderate amount of mitochondria. Immunohistochemical study (three cases) showed immunoreactivity with vimentin (two cases) and no reaction with cytokeratin, epithelial membrane antigen, S-100 protein, or desmin. Flow cytometric analysis showed diploid DNA content in three primary tumors and tetraploidy in one metastatic tumor. The proliferative activity (S-phase fraction) was low to intermediate (mean, 9.8%). Our data suggest that clear cell sarcoma of the kidney in the young adult age group resembles its pediatric counterpart in ultrastructural and immunohistochemical characteristics, proclivity for skeletal and visceral metastasis, DNA diploid status with relatively low S-phase, and aggressive clinical course. Clear cell sarcoma of the kidney in adult patients, although rare, must be differentiated from sarcomatoid carcinoma, sarcomas, and round cell tumors because of its unique characteristics in comparison to other renal neoplasms.

Clear cell tumors of the somatic soft tissues.

The prototypic soft tissue tumor showing "clear" cytoplasmic features is the so-called clear cell sarcoma or malignant melanoma of soft parts, a tumor characterized by immunophenotypic and ultrastructural melanogenic differentiation and specific cytogenetic and molecular abnormalities, ie, t(12;22)(q13;q12) translocation and Ewing's sarcoma oncogene/activating transcription factor 1 rearrangement. A number of other malignant soft tissue tumors occasionally show cytoplasmic clarity, including epithelioid leiomyosarcomas, malignant peripheral nerve sheath tumors, and sclerosing fibrosarcomas, as well as metastatic carcinomas and malignant melanomas. These tumors are discussed in relation to their differential diagnosis with clear cell sarcomas.

Clear cell rhabdomyosarcoma.

A clear cell rhabdomyosarcoma was studied by light microscopy, histochemistry, immunohistochemistry, and electron microscopy. It was a large, painful left parapharyngeal mass in a 10-year-old boy with intracranial extension and cervical metastatic enlarged lymph nodes. Tumor tissue was macroscopically grayish. At microscopic examination, the architecture was diffuse and focally alveolar. Tumor cells were of three types. Most cells were large, round or polygonal, with abundant clear vacuolated cytoplasm. Fibrils were sometimes found to be present around the nucleus. Nuclei often had irregular outlines and multiple nucleoli. Mitotic activity was high. Some round or elongated cells had eosinophilic fibrillar cytoplasm and were found to have a few double striations. A few cells were round and medium sized with a high nucleocytoplasmic ratio. Periodic acid-Schiff stain demonstrated huge amounts of intracytoplasmic glycogen in clear cells. Tumor cells showed positive immunostaining for muscle markers (desmin, muscle actins, dystrophin). Electron microscopy showed large lakes of glycogen, lipid droplets, and striated muscle features.

A patient with rhabdomyosarcoma and clear cell sarcoma of the skin.

We describe a patient with rhabdomyosarcoma of the posterior cervical region and clear cell sarcoma on the occipital scalp. These two tumors later metastasized to distant skin. We differentiated these tumors by histopathologic, histochemical, immunohistochemical, and ultrastructural findings. Cells of the posterior cervical tumor showed differentiation toward striated muscle, whereas those of the occipital tumor showed findings compatible with melanocytic differentiation.

[Benign nasal melanosis transformed into malignant mucosal melanoma: a documented case]. / Melanosis benigna nasal hacia melanoma maligno mucoso: una transformación documentada.

We report a case of nasal mucosal melanosis transformed into malignant mucosal melanoma. We review the feature of these disorders, their association, and preventive treatment.

Establishment and characterization of a clear-cell sarcoma (malignant melanoma of soft parts) cell line.

A clear cell sarcoma (CCS) cell line, designated as NCS-1, was established in monolayer culture from a xenograft line originating from a metastatic CCS. Marked karyotypic aberrations and tumorigenicity in nude mice revealed the malignant derivation of the NCS-1 cell line. These cells contained abundant glycogen and were amelanotic by light microscopy. By electron microscopy, however, melanosomes in various developmental stages were seen, and some of them were partially melanized. The electron microscopic dopa reaction revealed the presence of tyrosinase activity. Enzyme-linked immunoadsorbent assay revealed that NCS-1 cells expressed a 75-kDa glycoprotein which was identified as a marker of highly differentiated melanoma cells. From these results, NCS-1 cells were found to retain both cytochemical and morphological properties of CCS. Application of NCS-1 cells to a panel of monoclonal antibodies recognizing melanocytic differentiation antigens showed that they corresponded approximately to highly differentiated melanoma cells. In conclusion, the present study strongly supports the close relationship between CCS and malignant melanoma.

[Clear-cell sarcoma of the kidney: the clinico-morphological, immunohistochemical and electron microscopic study of 22 cases]. / Svetlokletochnaia sarkoma pochki: kliniko-morfologicheskoe, immunogistokhimicheskoe i élektronno-mikroskopicheskoe issledovanie 22 nabliudenii.

22 cases of clear cell sarcoma of the kidney (CCSK) were studied, 15 of them ultrastructurally and 7--immunohistochemically. CCSK is a rare malignant tumour of children, predominantly males, with an involvement of one kidney and frequent multiple metastases to the bones. Typical, epithelioid-trabecular, hyalinizing, cystic and myxoid variants are distinguished at the light microscopic level. As vimentin was found in tumour cells in all cases, antibodies against vimentin and cytokeratin can be recommended for differential diagnosis between CCSK and Wilms tumour and rhabdoid tumour. Peculiar vascular pattern can be revealed by means of antibodies against factor VIII. Immunohistochemical properties (presence of vimentin) and electron microscopy allow suggesting histogenic relation of CCSK with pericytes.