Molecular Genetic Analysis of Small Round Cell Sarcomas and its Application for Pathologic Differential Diagnosis

1) EWS/Fli1在Ewing肉瘤组中的DNA断点连接分析我们采用分子生物学技术对13例Ewing肉瘤进行了研究，检测了所有病例的EWS/Fli1融合转录本。在检查的七个案例中，有四个案例，在两个基因的断点附近发现了一个共识序列，5‘-AGAAAARDRR-3’。在大多数情况下，与Alu重复序列和/或真核拓扑异构酶II高度同源的序列也位于断点附近。这些提示EWS/Fli1可能是尤因肉瘤组特异性的。2) c-kit基因在小圆细胞肿瘤中的表达及突变检测c-kit基因编码一种跨膜受体激酶，在大多数胃肠道间质瘤（GIST）中表达，在大多数胃肠道间质瘤中，c-kit基因外显子11发生突变。我们检测了c-kit的近膜结构域突变；采用RT-PCR方法对GIST 26例、横纹肌肉瘤8例、神经母细胞瘤1例、神经源性肿瘤9例、平滑肌肉瘤9例的外显子9 ~ 11进行分析。免疫组化结果显示，所有GIST患者c-kit阳性，24例患者外显子9-11发生突变。在20例中检测到外显子11缺失或插入。尽管非GIST间质肿瘤组均无c-kit蛋白免疫组化表达，但7例横纹肌肉瘤、1例神经母细胞瘤、9例神经源性肿瘤和7例平滑肌肉瘤的9-11外显子突变与GIST相似。11外显子异常5例，1例，5例，3例。GIST组与非GIST间质肿瘤组的相似性提示小圆细胞瘤的癌变机制相同。

1) Analysis on the DNA breakpoint junction of EWS/Fli1 in Ewing sarcoma groupWe studied 13 cases of Ewing sarcoma by a molecular biological technique and detected the EWS/Fli1 fusion transcripts in all cases. In four cases of the examined seven, a consensus sequence, 5'-AGAAAARDRR-3' was found near the breakpoints of both genes. And sequences highly homologous to Alu repeats and/or eukaryotic topoisomerase II cleavage site were also located near the breakpoints in most of cases. These suggest that EWS/Fli1 may be specific to Ewing sarcoma group.2) Detection of expression and mutaion of c-kit gene in small round cell tumors C-kit gene encodes a transmembrane receptor kinase which is expressed in the majority of gastrointestinal stromal tumor (GIST) and in most cases of GIST, mutations in exon 11 of c-kit were reported. We examined a juxtamembrane domain mutation of c-kit ; exon 9 to 11 in 26 cases of GIST, eight of rhabdomyosarcoma, one of neuroblastoma, nine of neurogenic tumor, and 9 of leiomyosarcoma by using RT-PCR method. Immunohistochemically, all of GIST showed positivity for c-kit, and mutaions of exon 9-11 were found in 24. Deletion and/or insertion in exon 11 were detected in 20 cases. Although none of non-GIST mesenchymal-tumor group expressed c-kit protein immunohistochemically, 7 cases of rhabdomyosarcomas, one neuroblastoma, 9 neurogenic tumors, and 7 leiomyosarcomas had mutations in exon 9-11 simillar to those of GIST.Furthermore, abnormalities in exon 11 were found in 5 cases, one, 5, and 3, respectively. These findings of the similarity between GIST group and non-GIST mesenchymal-tumor group is suggested to be the same mechanism of carcinogenesis in small round cell tumors.