Clear cell sarcoma of vulva. A case report

Summary

We report the case of a 67-year-old female with clear cell sarcoma (CCS) of the vulva. Grossly, the tumor was a partly exophytical vulvar mass, measuring 20 x 15 cm. At the time of presentation, the patient showed metastases to the lung, inguinal and pelvic lymph nodes. Histologically, the tumor consisted of oval or spindle cells with only mild nuclear pleomorphism and rare mitoses (up to 2/10 HPF). The cytoplasm was pale eosinophilic or clear. The tumor cells were arranged in confluent sheets. There were large areas of necrosis and surface ulceration. Immunohistochemically, the tumor cells showed expression of S-100 protein and focal melan A and HMB45 expression. Fluorescent in situ hybridization analysis revealed rearrangement of the EWSR1 gene. To the best of our knowledge, this is the first report of CCS arising in the vulva.

Keywords:
clear cell sarcoma – EWSR1 gene – melanoma of soft parts – vulva

Clear cell sarcoma (CCS), also known as a melanoma of soft parts, is a malignant neoplasm first described by Enzinger in 1965 (1). This tumor is rare and accounts for 1 % of all soft tissue tumors. CCS typically involves deep soft tissues of the extremities, in close proximity to tendons and aponeurotic structures, especially in young adults (2). However, it can arise in other locations including the head and neck or trunk region and retroperitoneum. We report the case of a 67-year-old female with clear cell sarcoma of the vulva. To the best of our knowledge, our case represents the first report of CCS arising in the vulva.

CASE REPORT

Clinical history

A 67-year-old female presented with a one-year history of a slowly growing vulvar mass, finally measuring 20 x 15 cm at its largest size (Fig. 1). At the time of presentation, the patient showed metastases to the lung, inguinal and pelvic lymph nodes. A biopsy of the vulvar tumor was performed.

MATERIALS AND METHODS

Immunohistochemical analysis. Sections from formalin-fixed, paraffin-embedded tissue blocks were stained with hematoxylin- eosin. Selected sections were analysed immunohistochemically using the avidin-biotin complex method with antibody directed against following antigens: S-100 protein (1:600, Dako, Glostrup, Denmark), Melan A (clone A103, 1:25, Novocastra, Newcastle, UK), HMB45 (1:50, Dako) and Ki-67 (clone MIB-1, 1:50, Dako), cytokeratin AE1/AE3 (1:50, Dako, Glostrup, Denmark), cytokeratin high molecular weight (clone 34betaE12, 1: 200, Dako), estrogen receptors (ER, clone 6F11, 1:50, Novocastra), progesterone receptors (PR, clone 16, dilution 1:200, Novocastra), desmin (clone D33, 1:200, Dako), actin (clone HHF 35, 1:400 Dako), CD34 (clone QBEND 10, 1:50, Dako), CD99 (MIC2, clone 12E7, 1:50, Dako), synaptophysin (clone SY38, 1:25, Dako), chromogranin A (1:50, Dako) and p53 (clone DO-1, 1: 50, Bio, Genex).

Fluorescence in situ hybridization. The rearrangement of the EWSR1 gene (22q12) was analyzed using the Fluorescent in situ hybridization method with probe (Dual Color, Break Apart Rearrangement Probe from Abbott Vysis, Downers Grove, IL, USA). The assay procedure was conducted according to the manufacturer’s instructions.

Isolation of nucleic acid. Sections of formalin-fixed, paraffin-embedded tissue were used for DNA isolation using standard procedures. Approximately seven 10-mm-thick sections from each sample were deparaffinized in xylene. The DNA was then extracted using the QIAamp DNA mini kit (Qiagen, Hamburg, Germany).

Analysis of the BRAF Gene. BRAF 600/601 mutations were detected by polymerase chain reaction (PCR) or reverse-hybridization (StripAssay, ViennaLab, Austria) according to the manufacturer’s instructions.

RESULTS

Five incisional biopsies from the vulva of a 67-year-old female measured 2 mm to 7 x 5 x 3 mm. Histologically, the tumor consisted of oval cells arranged in confluent sheets mostly with only mild nuclear pleomorphism and rare mitoses (up to 2/10 HPF) (Fig. 2). The cytoplasm was pale eosinophilic or clear. There were large areas of necrosis and surface ulceration. Immunohistochemically, the tumor cells showed expression of S-100 protein and focal expression of melan A and HMB45. Ki-67 showed positivity in approximately 10 % of tumor cells (Fig. 3). Other markers examined were negative, including cytokeratin AE1/AE3, cytokeratin high molecular weight, estrogen receptors, progesterone receptors, desmin, actin, CD34, CD99, synaptophysin, chromogranin A and p53. Fluorescent in situ hybridization analysis revealed rearrangement of the EWSR1 gene (22q12). BRAF 600/601 mutations were not detected by polymerase chain reaction (PCR) or reverse- hybridization.

DISCUSSION

CCS is a rare tumor that usually presents as a slowly growing mass. Recurrences occur in 14 - 39 % of patients, whereas metastases develop in approximately 50 % of patients. The prognosis of metastasizing CCS is poor with a mean survival of 18.4 months (3). In non-metastasizing cases, unfavorable prognostic factors include tumor size > 5 cm, histological detection of necrosis, and local recurrence (4). Besides soft tissues, CCS can also arise in the gastrointestinal tract. However, it has been a matter of dispute whether CCS of the gastrointestinal tract represents an entity distinct from soft tissue lesions. Recently, a series of 16 cases was published under the term malignant gastrointestinal neuroectodermal tumor (5). CCS shows some phenotypic and immunohistochemical features overlapping with malignant melanoma (MM), which is the most important differential diagnostic consideration (6). However, in contrast to MM, CCS usually arises in deep soft tissues without involvement of the skin. Moreover, neither nuclear pleomorphism nor high mitotic activity are typical features of CCS. However, pleomorphic cases can occur. Immunohistochemically, CCS and MM show the expression of S-100 protein, HMB-45, Melan A and microphthalmia transcription factor (MiTF). In contrast to MM, CCS is characterized by a rearrangement of the Ewing’s sarcoma (EWS) gene. The most common rearrangement is a balanced translocation (12;22)(q13;q12). This results in the fusion of the EWS gene at 22q12 to the activating transcription factor 1 (ATF1) gene localized at 12q13 (7,8). A translocation (2;22)(q34;q12) resulting in fusion of EWSR1/CREB1 occurs less frequently (9). These changes have not been identified in MM. Moreover, BRAF and NRAS mutations that are commonly found in MM have not been described in CCS (10).

In conclusion, we have described the first reported case of CCS arising in the vulva. The clinician should be aware of the possibility of this unusual primary location of CCS to avoid confusion with other tumors, especially malignant melanoma. In doubtful cases, molecular analysis of the EWSR1 gene will distinguish between these tumors.

ACKNOWLEDGEMENTS

This work was supported by Charles University in Prague (Project PRVOUK-P27/LF1/1, UNCE 204024, Ministry of health, Czech Republic - conceptual development of research organisation 64 165, General University Hospital in Prague, Czech Republic, and by OPPK (Research Laboratory of Tumor Diseases, CZ.2.16/3.1.00/24509).

CONFLICT OF INTEREST

The authors declare that there is no conflict of interest regarding the publication of this paper.

Correspondence address:

Kristýna Němejcová, M.D., Ph.D.

First Faculty of Medicine and General University Hospital,

Charles University in Prague,

Studničkova 2,

Prague 2, 12800,

Czech Republic phone: +420224968632

e-mail: kristyna.nemejcova@vfn.cz