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Test Code DSRCT Desmoplastic Small Round-Cell Tumor by Reverse Transcriptase PCR (RT-PCR)

Useful For

Supporting the diagnosis of desmoplastic small round-cell tumor

Additional Tests

Test ID Reporting Name Available Separately Always Performed
SLIRV Slide Review in MG No, (Bill Only) Yes

Reflex Tests

Test ID Reporting Name Available Separately Always Performed
EWSF EWSR1 (22q12), FISH, Ts Yes No

Testing Algorithm

This test is performed in conjunction with slide review. Additional testing may be performed after review by pathologist. Upon approval from the requesting clinician, PATHC / Pathology Consultation may be added, if determined to be appropriate.

Special Instructions

Method Name

Reverse Transcriptase Polymerase Chain Reaction (RT-PCR)/Gel Electrophoresis

Reporting Name

Desmo Small Round-Cell Tumor,RT-PCR

Specimen Type

Varies


Specimen Required


A pathology/diagnostic report including a brief history is required.

 

Specimen Type:

Container/Tube: Surgical Pathology Packaging Kit (T554) requested, but not required

Preferred: Formalin-fixed, paraffin-embedded (FFPE) tissue with a minimum of 5% tumor cell population

Acceptable: Unstained slides with a minimum of 5% tumor cell population; slides may be stained and/or scraped

Collection Instructions:

1. Process all specimens into FFPE blocks prior to submission.

2. If submitting slides, a minimum of ten, 4- to 5-micron thick, unstained slides are required.

Additional Information:

1. A quality specimen is essential for evaluation. Submit only tissue containing tumor cells; minimal tissue is required for evaluation.

2. Special stains performed outside Mayo Medical Laboratories and included with the case may be repeated and charged at the reviewing pathologist's discretion. Testing requested by the referring physician may not be performed if deemed unnecessary by Mayo Clinic pathologist.


Specimen Stability Information

Specimen Type Temperature Time
Varies Ambient (preferred)
  Frozen 
  Refrigerated 

Reject Due To

Note: No specimen should be rejected. If specimen not received at appropriate temperature or in wrong anticoagulant, include note to laboratory. If questions, contact laboratory

Clinical Information

Desmoplastic small round-cell tumor (DSRCT) is a member of the small round-cell tumor group that also includes rhabdomyosarcoma, synovial sarcoma, lymphoma, Wilms tumor, and Ewing sarcoma. DSRCT is a type of sarcoma that affects mainly children and adolescent males, usually in the form of widespread intra-abdominal growth not related to any specific organ system. The tumor is composed of angulated nests of small round cells with an abundant desmoplastic stroma. The tumor cells show multiphenotypic differentiation and are usually positive for cytokeratin and desmin.(1-4) These tumors can express renal, epithelial, muscle, and endocrine markers.

 

While treatment and prognosis depend on establishing the correct diagnosis, the diagnosis of sarcomas that form the small round-cell tumor group can be very difficult by light microscopic examination alone, especially true when only small-needle biopsy specimens are available for examination. The use of histochemical and immunohistochemical stains (eg, desmin, cytokeratin, and WT1) can assist in establishing the correct diagnosis, they cannot distinguish between DSRCT and other small round-cell tumors. Expertise in soft tissue and bone pathology are often needed.

 

Studies have shown that some sarcomas have specific recurrent chromosomal translocations. These translocations produce highly specific gene fusions that help define and characterize subtypes of sarcomas and are useful in the diagnosis of these lesions.(1-4)

 

DSRCT is associated with a unique chromosomal translocation t(11:22)(p13;q12) that involves the EWSR1 and the WT1 genes. EWSR1 is the breakpoint site of translocations associated with Ewing sarcoma and WT1 is a gene altered in some Wilms tumors. The translocation results in a fusion of the 2 genes with expression of a chimeric EWSR1-WT1 product. The most common breakpoints involve the intron between EWSR1 exon 7 and 8 and the intron between WT1 exons 7 and 8. Analyses of these transcripts have shown an in-frame fusion of RNA encoding the amino-terminal domain of EWSR1 to the zinc finger of the DNA-binding domain of WT1.

Reference Values

An interpretative report will be provided.

Interpretation

A positive EWSR1-WT1 result is consistent with a diagnosis of desmoplastic small round-cell tumor (DSRCT).

 

Sarcomas other than DSRCT, and carcinomas, melanomas, and lymphomas are negative for the fusion products.

 

A negative result does not rule out a diagnosis of DSRCT.

Cautions

Reliable results are dependent on adequate specimen collection and processing. This test has been validated on formalin-fixed, paraffin-embedded tissues; other types of fixatives are discouraged. Improper treatment of tissues, such as decalcification, may cause PCR failure. False-negative results may occur in tumor specimens when tumor cells comprise <5% of the cell population. Tumor cells may be enriched by macrodissection to avoid false-negative results

 

Clinical diagnosis or therapy should not be based solely on this assay. The results should be considered in conjunction with clinical information, histologic evaluation, and additional diagnostic tests.

Supportive Data

A total of 36 cases that were diagnosed as desmoplastic small round-cell tumor by soft tissue pathology experts and by immunohistochemical staining were analyzed. These consisted of 4 frozen tissue specimens and 32 paraffin sections. Three of the cases from frozen and paraffin tissue sections were the same cases for direct comparison. All specimens were also analyzed by Southern hybridization using a sequence internal to the primers as a probe. A few specimens were also analyzed by sequencing the PCR products.

 

The EWSR1-WT1 product was detected in all 4 (100%) fresh tissues and in 30/32 (94.7%) of the paraffin sections. Southern hybridization detected 1 additional case in the paraffin section that was not detected on the ethidium bromide gel. Sequencing of the fresh tissues and paraffin sections confirmed the sequence of the breakpoint of the fusion transcript of EWSR1 exon 7 to WT1 exon 8. Comparable RT-PCR results were obtained for the 3 cases that were analyzed by frozen and paraffin tissues. Analysis of other small round-cell tumors (alveolar rhabdomyosarcoma, synovial sarcoma, and Ewing sarcoma) (n=12) were all negative for the EWSR1-WT1 fusion transcript.

Clinical Reference

1. Barr FG, Chatten J, D'Cruz CM, et al: Molecular assays for chromosomal translocations in the diagnosis of pediatric soft tissue sarcomas. JAMA 1995;273:553-557

2. Gerald WL, Miller HK, Battifora H, et al: Intra-abdominal desmoplastic small round-cell tumor: Report of 19 cases of a distinctive type of high-grade polyphenotypic malignancy affecting young individuals. Am J Surg Pathol 1991;15:499-513

3. Gerald WL, Rosai J, Ladanyi M: Characterization of the genomic breakpoint and chimeric transcripts in the EWS-WT1 gene fusion of demoplastic, small round-cell tumor. Proc Natl Acad Sci USA 1995;92:1028-1032

4. Jin L, Majerus J, Oliveira A, et al: Detection of fusion gene transcripts in fresh-frozen and formalin-fixed paraffin-embedded tissue sections of soft tissue sarcomas after laser capture microdissection and RT-PCR. Diagn Mol Pathol 2003;12:224-230

Method Description

The paraffin-embedded tissue is deparaffinized, lysed, and digested. RNA is extracted using either the TRIzol kit (Invitrogen) or High Pure FFPE RNA Micro kit (Roche). DNase digestion is performed. RNA is converted to cDNA via reverse transcription (RT); the cDNA is amplified via PCR. Primers specific for the desmoplastic small round-cell tumor EWSR1-WT1 transcript are used. Controls are run with each specimen to assess possible contamination issues and overall test performance. The PCR products are separated by gel electrophoresis and stained with ethidium bromide. The agarose gel is viewed under ultraviolet light and photographed to document the results. In some cases, the specimen is further analyzed by FISH and sequencing.(Jin L, Majerus J, Oliveira A, et al: Detection of fusion gene transcripts in fresh-frozen and formalin-fixed paraffin-embedded tissue sections of soft tissue sarcomas after laser capture microdissection and RT-PCR. Diagn Mol Pathol 2003;12:224-230)

Day(s) and Time(s) Performed

Monday through Friday; Varies

Analytic Time

8 days

Specimen Retention Time

Unused portions of blocks will be returned. Unused slides are stored indefinitely.

Performing Laboratory

Mayo Medical Laboratories in Rochester

Test Classification

This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. This test has not been cleared or approved by the U.S. Food and Drug Administration.

CPT Code Information

81401-EWSR1/WT1 (t(11;22)) (eg, Ewing sarcoma/peripheral neuroectodermal tumor), translocation analysis, qualitative, and quantitative, if performed

88381-Microdissection, manual

LOINC Code Information

Test ID Test Order Name Order LOINC Value
DSRCT Desmo Small Round-Cell Tumor,RT-PCR In Process

 

Result ID Test Result Name Result LOINC Value
54754 Result Summary 50397-9
54755 Result In Process
54756 Interpretation 69047-9
54757 Additional Information 48767-8
54758 Reason for Referral 42349-1
54759 Specimen 31208-2
54760 Source 31208-2
54761 Tissue ID 81178-6
54762 Released By 18771-6

Forms

1. Pathology/Cytology Information (T707) in Special Instructions

2. If not ordering electronically, complete, print, and send 1 of the following forms with the specimen:

Pathology Test Request Form (T246) (http://www.mayomedicallaboratories.com/it-mmfiles/pathology-request-form.pdf)

Oncology Test Request Form (T729) (http://www.mayomedicallaboratories.com/it-mmfiles/oncology-request-form.pdf)