Kibel Adam S, Faith Dennis A, Bova G Steven, Isaacs William B
Division of Urologic Surgery, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Prostate. 2002 Sep 1;52(4):305-10. doi: 10.1002/pros.10112.
In an effort to better understand the molecular events responsible for progression of prostate carcinoma to metastatic disease, we have recently identified a homozygous deletion at 12p12-13 involving ETV6 (tel). Although mutational analysis of ETV6 has not been examined previously in prostate carcinoma, it is an attractive candidate prostate cancer tumor suppressor gene since as it previously has been implicated in malignancy. Therefore, we decided to analyze 43 prostate cell lines, xenografts, and metastatic foci for inactivating mutations.
DNA was isolated from 7 cell lines, 18 xenografts, and 18 metastatic deposits. Single-strand conformational polymorphism (SSCP) analysis of ETV6, was performed by polymerase chain reaction (PCR) amplification of each exon by using intron specific primers. PCR products were then resolved by gel electrophoresis, and aberrantly migrating PCR products were then sequenced.
Two previously described polymorphisms and four novel sequence changes were identified. Polymorphisms at nucleotide 258 (G --> A, Thr --> Thr) and 602 (T --> C, Leu --> Pro) were identified in eight and one specimen(s), respectively. Analysis of noncancerous DNA confirmed the presence of the polymorphisms in the germ-line. Four possible mutations were identified at nucleotides 24 (T --> G, Cys --> Trp), 380 (G --> A, Arg --> Glu), 776 (G --> T, Arg --> Leu), and 876 (C --> T, Leu --> Leu). Three were in xenografts or cell lines. Because normal DNA was not available, these could represent rare polymorphisms. The sole mutation in a clinical specimen, at nucleotide 876, did not result in an amino acid change.
Our data suggest that mutational inactivation ETV6 may occur in prostate carcinoma. The functional significance of these potential inactivating mutations remains to be determined.
为了更好地理解导致前列腺癌进展为转移性疾病的分子事件，我们最近在12p12 - 13区域发现了一个涉及ETV6（tel）的纯合缺失。尽管之前尚未对前列腺癌中的ETV6进行突变分析，但它是一个有吸引力的前列腺癌肿瘤抑制基因候选者，因为它之前已与恶性肿瘤相关联。因此，我们决定分析43个前列腺细胞系、异种移植物和转移灶，以寻找失活突变。
从7个细胞系、18个异种移植物和18个转移灶中分离DNA。通过使用内含子特异性引物对ETV6的每个外显子进行聚合酶链反应（PCR）扩增，进行单链构象多态性（SSCP）分析。然后通过凝胶电泳分离PCR产物，并对迁移异常的PCR产物进行测序。
鉴定出两个先前描述的多态性和四个新的序列变化。分别在8个和1个样本中鉴定出核苷酸258（G→A，苏氨酸→苏氨酸）和602（T→C，亮氨酸→脯氨酸）处的多态性。对非癌DNA的分析证实了种系中存在这些多态性。在核苷酸24（T→G，半胱氨酸→色氨酸）、380（G→A，精氨酸→谷氨酸）、776（G→T，精氨酸→亮氨酸）和876（C→T，亮氨酸→亮氨酸）处鉴定出四个可能的突变。其中三个在异种移植物或细胞系中。由于没有正常DNA，这些可能代表罕见的多态性。临床样本中唯一的突变位于核苷酸876，未导致氨基酸变化。
我们的数据表明，ETV6的突变失活可能发生在前列腺癌中。这些潜在失活突变的功能意义仍有待确定。
