Miao Jiangyong, Kusafuka Takeshi, Fukuzawa Masahiro
The Department of Pediatric Surgery, Osaka University Graduate School of Medicine, Suita, Osaka, Japan.
Oncol Rep. 2004 Dec;12(6):1269-72.
BRAF (v-raf murine sarcoma viral oncogene homolog B1) activating mutations in a high proportion of melanomas and in a small fraction of other cancers have been recently reported. All the presented mutations of BRAF are located in exons 11 and 15, and the hotspot mutation at codon 599 accounts for 87% of BRAF mutations. Because the mutational status is unclear in pediatric solid neoplasms, we screened BRAF mutations comprehensively in our tumor series presented in childhood. Two pairs of primers were designed to amplify exons 11 and exon 15, respectively, and 181 tumor samples (65 neuroblastomas, 23 Wilms tumors, 19 hepatoblastomas, 16 teratomas, 17 rhabdomyosarcomas, 13 ganglioneuromas, etc.) were investigated by PCR-SSCP method. On agarose gel electrophoresis, amplified PCR fragments showed no size-altered changes in exons 11 and 15, and SSCP analysis revealed uniform band patterns in both exons. Subsequent direct sequencing of selected 10 samples confirmed only normal sequences without any nucleotide substitutions. The current study represents the first genetic analysis of the BRAF gene in pediatric solid tumors. Our data suggest that mutations of BRAF gene as a mechanism of tumorigenesis is unlikely to be associated with most childhood neoplasms.
最近有报道称，BRAF（v-raf鼠肉瘤病毒癌基因同源物B1）激活突变在高比例的黑色素瘤以及一小部分其他癌症中存在。所有已报道的BRAF突变均位于第11和15外显子，密码子599处的热点突变占BRAF突变的87%。由于儿童实体瘤的突变状态尚不清楚，我们对我们所收集的儿童期肿瘤样本进行了BRAF突变的全面筛查。设计了两对引物分别扩增第11外显子和第15外显子，并通过PCR-SSCP方法对181个肿瘤样本（65个神经母细胞瘤、23个肾母细胞瘤、19个肝母细胞瘤、16个畸胎瘤、17个横纹肌肉瘤、13个神经节瘤等）进行了研究。在琼脂糖凝胶电泳上，扩增的PCR片段在第11和15外显子中未显示大小改变，SSCP分析显示两个外显子中的条带模式均一致。随后对所选的10个样本进行直接测序，仅证实为正常序列，无任何核苷酸替代。本研究是对儿童实体瘤中BRAF基因的首次基因分析。我们的数据表明，BRAF基因突变作为一种肿瘤发生机制不太可能与大多数儿童肿瘤相关。
