Original Article
A Combined Cytogenetic and Molecular Approach to Diagnosis in a Case of Desmoplastic Small Round Cell Tumor with a Complex Translocation (11;22;21)

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Abstract

Desmoplastic small round cell tumor (DSRCT) has recently been described as a discrete tumor entity. It is distinguished from other small round cell tumors by its prominent desmoplastic quality, its preponderance in adolescent males, its almost exclusive intraabdominal location, a multi-immunophenotypic profile, and its aggressive nature. Diagnosis on histology alone is not always unequivocal. A recurrent t(11;22)(p13;q12) translocation has recently been described in this tumor, and a chimeric RNA fusion product formed from the WT1 and EWS genes is detectable by reverse transcriptase-polymerase chain reaction (RT-PCR). We describe the use of a multi-faceted approach using conventional G-banding, fluorescence in situ hybridization (FISH) and RT-PCR to assist the diagnosis of a case of DSRCT with a complex variant t(11;22;21)(p13;q12;q22.1) translocation and demonstrate the value of a combined approach to genetic investigation of solid tumors.

Introduction

Cytogenetic and molecular investigation of small round cell tumors has enhanced our knowledge of tumor pathogenesis and facilitated diagnosis, particularly in cases where histological results are equivocal. Desmoplastic small round cell tumor (DSRCT) is a recently recognized discrete tumor entity [1] presenting with some diagnostic difficulty, where genetic investigation can be of importance. It is found predominantly in adolescent males, usually in an intraabdominal location. Histologically, the tumor shows a “divergent” multi-immunophenotypic profile, with abundant desmoplastic stroma. The clinical behavior is aggressive and its response to treatment is poor. Current treatment protocols follow those for other histologically similar soft tissue tumors. Understanding the pathogenesis of this disease has improved since the observation of a recurrent t(11;22)(p13;q12) translocation was described 2, 3, 4, 5. Further, reverse transcriptase polymerase chain reaction (RT-PCR) investigation has shown that the rearrangement produces a chimeric RNA product formed from the rearrangement of the genes WT1 at 11p13 and EWS at 22q12 [6]. We have investigated a case of DSRCT by conventional G-banding, fluorescence in situ hybridization (FISH) and RT-PCR. Our case showed a complex variant t(11;22;21) translocation indicating movement of 11p to 22q. Fluorescence in situ hybridization using a WT1 probe confirmed movement of WT1 to 22q. Reverse transcriptase polymerase chain reaction using primers located in the region of WT1 and EWS confirmed the presence of a WT1-EWS chimeric fusion transcript. We therefore advocate the use of one or more of these genetic techniques to assist in the diagnosis of this poorly understood tumor entity.

Section snippets

Case report and histology

A 14-year-old male presented with 2–3 months of abdominal pain and shortness of breath, and subsequently developed dysuria. An ultrasound scan revealed an intraabdominal mass. A computerized tomography (CT) scan confirmed the presence of multiple abdominal and pelvic masses, the largest of which was inseparable from the bladder. The cerebro-spinal fluid (CSF) was clear, and bone marrow was found to contain 1–2% malignant infiltration. Surgical incision revealed right iliac adenopathy, omental

Cytogenetics

Cell suspensions for cytogenetic analysis were obtained from bone marrow and tumor tissue. Bone marrow was cultured in RPMI 1640 medium (GIBCO) supplemented with 20% fetal calf serum (LabTech), glutamine (GIBCO), and antibiotics (GIBCO), and incubated at 37° C in a closed system in 0.2 μg/mL colcemid (Sigma) for 17 hours. Harvesting was completed following incubation for 15 minutes in 0.075M KCl and three washes in Carnoy’s fixative.

Tumor tissue was minced by mechanical disaggregation using

Cytogenetics

The short-term suspension and 8-day-flask cultures from the tumor produced an abnormal karyotype containing a complex three-way translocation between chromosomes 11, 22, and 21 Figure 3, Figure 4, and several other abnormalities: 48,X,−Y,+1,der(1;14)(q10;q10),+5,t(11;22;21)(p13; q12;q22.1),+15,+18. Bone marrow cultures produced cells with a normal male (46,XY) karyotype.

Fish

Following G-banded analysis, very little fixed suspension was available for FISH studies. The majority of the small number of

Discussion

Differential diagnosis of small round cell tumors (including primitive neuroectodermal tumor, Ewing sarcoma, rhabdomyosarcoma, mesenchymal chondrosarcoma, small cell osteosarcoma, hemangiopericytoma, and neuroblastoma) can be extremely difficult.

Genetic investigation of solid tumors is increasingly being used to assist with diagnosis and assessment of prognosis. G-banded analysis is useful in providing a broad overview of the genetic picture of the disease; however, occasional problems of

Acknowledgements

The RT-PCR work was supported by the Candlelighter’s Trust. We are grateful to Dr. K. Pritchard-Jones from the Department of Pediatric Oncology, Royal Marsden NHS Trust, Sutton, Surrey, UK for donation of the E1241 probe.

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