Synovial Sarcoma by Reverse Transcriptase PCR (RT-PCR), Paraffin
Supporting a diagnosis of synovial sarcoma
Clinical Information Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test
Synovial sarcomas account for 9% to 10% of soft tissue tumors. These tumors occur in 2 major forms: biphasic and monophasic. Monophasic tumors are composed entirely of spindle cells, while biphasic tumors have epithelial cells arranged in glandular structures and mixed with spindle cells. The tumors are usually positive for keratin and epithelial membrane antigen as well as vimentin by immunostaining.
Synovial sarcoma is a member of the small round-cell tumor group that includes rhabdomyosarcoma, lymphoma, Wilms tumor, Ewing sarcoma, and desmoplastic small round-cell tumor. While treatment and prognosis depend on establishing the correct diagnosis, the diagnosis of sarcomas that form the small round-cell tumor group by light microscopic examination alone can be very difficult, especially true when only small-needle biopsy specimens are available for examination. The use of immunohistochemical stains (eg, keratin and epithelial membrane antigen [EMA]) can assist in establishing the correct diagnosis, but these markers are not entirely specific for synovial sarcoma. 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)
Cytogenetic studies have shown a distinctive chromosomal translocation, t(X;18)(p11;q11), in more than 90% of synovial sarcomas. Cloning of the translocation breakpoint showed that t(X;18) results in the fusion of 2 genes designated as SS18 (at 18q11) and SSX (at Xp11). Two closely related genes, SSX1 and SSX2, have 81% homology in proteins. SS18-SSX1 is present in 55% of cases, while SS18-SSX2 is present in 35% of cases. Patients with SS18-SSX2 translocation usually have greater metastasis-free survival than those with SS18-SSX1.
These fusion transcripts can be detected by reverse transcriptase PCR (RT-PCR), by FISH, chromogenic in situ hybridization (CISH), or by classical cytogenetic analyses. The RT-PCR and FISH procedures are the most sensitive methods to detect these fusion transcripts.(3)
Reference Values Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.
A positive SS18-SSX1 or SS18-SSX2 result is consistent with a diagnosis of synovial sarcoma.
Sarcomas other than synovial sarcoma, and carcinomas, melanomas, and lymphomas are negative for the fusion products.
A negative result does not rule out a diagnosis of synovial sarcoma.
Cautions Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances
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 <10% of the cell population. Tumor cells may be enriched by macrodissection to avoid false-negative results.
Clinical diagnosis and therapy should not be based solely on the results of this assay. The results should be considered in conjunction with clinical information, histologic evaluation, and additional diagnostic tests.
A total of 45 cases diagnosed as synovial sarcomas by soft tissue pathology experts and by immunohistochemical staining were analyzed. These consisted of 15 frozen tissue specimens and 30 paraffin sections. Three of the cases from frozen tissues and paraffin tissue sections were the same cases for direct comparison.
The primers used to detect common SS18-SSX1 and SS18-SSX2 resulted in a 130-bp fragment, while the SS18-SSX1 and SS18-SSX2 were 151 bp and 109 bp, respectively. For the frozen tissues, there were 12 (80%) SS18-SSX1-positive cases and 3 (20%) SS18-SSX2-positive cases. For paraffin sections, 27/30 (90%) of the cases were reactive. Paraffin sections had 19 of 27 (70.4%) SS18-SSX1 cases and 8 of 27 (29.6%) SS18-SSX2 cases. Southern hybridization and sequencing supported the ethidium bromide gel finding. Comparable RT-PCR results were obtained for the 3 cases that were analyzed by frozen and paraffin sections.
Analysis of other small round-cell tumors (alveolar rhabdomyosarcoma, desmoplastic round-cell tumor, and Ewing sarcoma) (n=12) were all negative for the SS18-SSX1/SSX2 transcripts. One of 8 malignant nerve-sheath tumors had a weak band for SS18-SSX1.
Clinical Reference Provides recommendations for further in-depth reading of a clinical nature
1. Crew AJ, Clark J, Fisher C, et al: Fusion of SYT to two genes SSX1 and SSX2 encoding proteins with homology to the Kruppel-associated box in human synovial sarcoma. EMBO J 1995;14:2333-2340
2. Clark J, Rocques PJ, Crew AJ, et al: Identification of novel genes, SYT and SSX, involved in the t(X;18)(p11.2;q11.2) translocation found in human synovial sarcoma. Nat Genet 1994;7:502-508
3. Kawaii A, Wodruff J, Healey JH, et al: SYT-SSX gene fusion as a determinant of morphology and prognosis in synovial sarcoma. N Engl J Med 1998;338:153-160
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