Myxoid liposarcoma with or without a round cell component is the most common subtype of liposarcoma. The diagnosis of myxoid liposarcoma could be challenging with histology, as a variety of soft tissue tumors with myxoid change might mimic myxoid liposarcoma, especially on small biopsy tissues. Chromosomal translocations of t(12,16) (q13;p11) and t(12;22) (q13;q12), rendering gene fusions of DDIT3 (previously CHOP) with FUS and EWSR1, have been found to be characteristic of myxoid liposarcoma, and were identifiable in more than 95% cases. These genetic alterations, therefore, are ideal as molecular markers to facilitate the diagnosis of this type of tumor. DDIT3 (12q13) dual-color break-apart rearrangement probe for fluorescence in situ hybridization has been commercially available. However, its consistency with DDIT3-associated gene fusion and its clinical use, including sensitivity and specificity, have not been adequately evaluated. In this study, we assessed the locus specificity of the probe on metaphase, and then tested it on 8 cases of myxoid liposarcoma, 12 cases of other sarcomas, and 18 cases of tumors with myxoid differentiation. All 8 myxoid liposarcomas showed DDIT3 gene break-apart, whereas all 12 other sarcomas were negative. All the cases with DDIT3 break-apart also showed FUS-DDIT3 fusion by reverse transcription-polymerase chain reaction, with 100% consistency. In addition, the FISH assay has been clinically applied on 18 myxoid tumors with promising outcome. In conclusion, FISH with DDIT3 break-apart probe is a highly sensitive and specific assay for detection of DDIT3-associated gene fusions, and therefore is a valuable adjunct in diagnosis or differential diagnosis of myxoid liposarcoma.

Expression of the transducin-like enhancer of split 1 (TLE1) by immunohistochemistry (IHC) has been widely used as a biomarker for the diagnosis of synovial sarcoma. Although TLE1 expression can be identified in more than 90% of synovial sarcomas, positive staining has been reported in up to one third of nonsynovial sarcomas, including peripheral nerve sheath tumors and neoplasms of fibrous and adipose tissues. The low specificity of this test in soft tissue tumors raises concern on its clinical application as a diagnostic biomarker. As synovial sarcoma is frequent among the differential diagnosis of unclassified high-grade sarcomas, and considering that the specificity of TLE1 antibody in this tumor group remains unclear, we evaluated TLE1 expression by IHC in 42 unclassified high-grade sarcomas. SS18 (SYT) gene break-apart analyses by fluorescence in situ hybridization were simultaneously performed as a gold standard biomarker for synovial sarcoma. Five cases that were positive for the SS18 break-apart by fluorescence in situ hybridization were also positive for TLE1 by IHC, whereas the remaining 37 cases negative for SS18 break-apart were all negative for TLE1. The results showed no evidence of nonspecific TLE1 expression in the nonsynovial high-grade sarcomas. We concluded that TLE1 is a highly specific biomarker for synovial sarcoma in the setting of differential diagnosis of unclassified high-grade sarcomas.

TMPRSS2 gene fusions with ETS transcription factor family members ERG, ETV1, or ETV4 have been recently discovered as a common molecular event in prostate cancer. Much attention has been focused on exploring their clinical application as a genetic tumor marker for the diagnosis, prognosis, and prediction of response to therapy. Although several studies have been done, the clinical utility of TMPRSS2 genetic alterations as biomarkers for prostate carcinoma remains indeterminate. In this study, we examined adenocarcinomas, prostatic intraepithelial neoplasia (PIN), and normal epithelium of the prostate retrieved from radical prostatectomy specimens to determine the frequency, specificity, tissue heterogeneity, and prognostic value of TMPRSS2 genetic alterations using a direct-labeled TMPRSS2 dual-color break-apart fluorescence in situ hybridization (FISH) probe cocktail designed to detect all known TMPRSS2-associated deletions or translocations. Seventy-one patients (161 samples) with normal prostate tissue, 60 patients (153 samples) with PIN, and 61 patients (142 samples) with carcinoma in formalin-fixed paraffin-embedded tissue microarrays were tested. None of the 161 normal prostate samples showed TMPRSS2 translocation or deletion. Sixty-two percent patients of prostate carcinomas demonstrated TMPRSS2 gene alterations, including 39% with translocation, 16% with deletion, and 7% with a mixed pattern. Tissue heterogeneity for TMPRSS2 gene alterations was identified in 28% of prostate carcinomas. No difference in the frequency of TMPRSS2 gene alterations was found between Gleason 6 and 7 tumors. Seventeen percent of PIN had TMPRSS2 gene alterations and showed the same FISH patterns as in the carcinomas from respective prostatectomy specimens. The TMPRSS2 dual-color break-apart FISH probe cocktail provides a simple and reliable method for the detection of TMPRSS2-related genetic alterations in formalin-fixed paraffin-embedded tissue. TMPRSS2 genetic alterations detectable by this method are strictly restricted in prostate neoplasia, and can be identified in the majority of prostate carcinomas. Tissue heterogeneity for TMPRSS2 alterations is common, and it should be considered when sampling and evaluating biopsy specimens.

HER2 gene amplification by fluorescence in situ hybridization and protein expression by immunohistochemistry (IHC) have been used for prognosis and guiding treatment of invasive ductal carcinoma of the breast with trastuzumab. Accurate evaluation of HER2 status is important in the management of patients with candidacy for the HER2-targeting therapy. Despite previous studies, effects of polysomy of chromosome 17, at which HER2 is located, on HER2 protein expression remains controversial. In this study, we calculated the average copy numbers of HER2 and chromosome 17 (CEP17) per nucleus in 109 cases of invasive breast carcinoma and analyzed their correlations with the HER2/CEP17 ratio and protein expression. As expected, there were close correlations between HER2 protein expression and the HER2/CEP17 ratio (CC: 0.49, P<0.001), along with the HER2 copy number per nucleus (CC: 0.48, P<0.001) and between the CEP17 copy number per nucleus and the HER2 copy number per nucleus (CC: 0.45, P<0.001). Correlation between the CEP17 copy number per nucleus and the HER2/CEP17 ratio was not significant (CC: 0.2, P>0.05). There was a weak, but statistically insignificant, correlation between the CEP17 copy number per nucleus and HER2 protein expression by IHC (CC: 0.26, P>0.05). The cases were then grouped on the basis of the amplification of the HER2 gene by fluorescence in situ hybridization. In the cases that showed no amplification, there was a significantly higher CEP17 copy number per nucleus in cases with strong HER2 protein expression (2.99) when compared with cases with weak (2.39) or absent (1.86) expression. In conclusion, a high HER2 gene copy number-associated polysomy 17 is a significant contributing factor in HER2 protein overexpression in unamplified invasive breast carcinomas. Cases without HER2 amplification, but carrying polysomy 17, should be further evaluated for HER2 protein overexpression by IHC. Those with polysomy 17-associated HER2 protein overexpression, like any other IHC+ ones, should be eligible candidates for trastuzumab therapy.

The characteristic immunoprofile for the diagnosis of synovial sarcoma, a neoplasm of unclear tissue origin, is expression of transducer-like enhancer of split 1 (TLE-1), CD99, partial expression of cytokeratin, and epithelial membrane antigen by immunohistochemistry (IHC). Diagnostic dilemma or misdiagnosis can occur due to overlap in IHC and morphology with carcinomas, and particularly poorly differentiated and metastatic tumors. The frequency of TLE-1 and CD99 expression in carcinomas by IHC has not been previously assessed. We evaluated TLE-1 and CD99 expression in various carcinomas and evaluated the expression of the SS18 (SYT) gene rearrangement (a characteristic biomarker for synovial sarcoma) in tumors with TLE-1 and/or CD99 expression. Immunostains of TLE-1 and CD99 were performed in 100 various carcinomas. Seven of the 98 cases (7%) of carcinomas showed TLE-1 expression, including 1 each of prostate adenocarcinoma (ADCA), esophageal ADCA, basal cell carcinoma, adrenocortical carcinoma, endometrial ADCA, ovarian serous carcinoma, and small cell carcinoma. Twenty-one of the 100 cases (21%) of carcinomas demonstrated CD99 expression, including 6 prostate ADCA, 3 esophageal ADCA, 5 squamous cell carcinomas, 2 hepatocellular carcinomas, 1 each for endometrial ADCA, renal cell carcinoma, urothelial cell carcinoma, neuroendocrine carcinoma, and mucoepidermoid carcinoma. An esophageal ADCA was positive for both TLE-1 and CD99. None of the carcinomas with positive TLE-1 (n=7) or CD99 (n=21) by IHC showed SS18 gene rearrangement by fluorescent in situ hybridization. TLE-1 and CD99 expression were identified in 7% and 21% of carcinomas, respectively. This is a potential pitfall in the IHC interpretation for diagnosis of synovial sarcoma. SS18 gene rearrangement by fluorescent in situ hybridization is helpful for the diagnostically challenging cases, either for confirmation or exclusion of synovial sarcoma.