Kohler M F, Marks J R, Wiseman R W, Jacobs I J, Davidoff A M, Clarke-Pearson D L, Soper J T, Bast R C, Berchuck A
Department of Obstetrics and Gynecology/Division of Gynecologic Oncology, Duke University Medical Center, Durham, N.C. 27710.
J Natl Cancer Inst. 1993 Sep 15;85(18):1513-9. doi: 10.1093/jnci/85.18.1513.
The p53 gene encodes a nuclear phosphoprotein present in low levels in normal human cells. The wild-type form of this protein functions to restrain inappropriate cellular proliferation. Approximately one half of human epithelial ovarian cancers have mutations in the p53 gene and overexpress the mutant protein product. Deletion of one allele of the p53 gene also frequently occurs in these cancers.
We sought to define the spectrum of mutations in the p53 gene in epithelial ovarian cancer with respect to both the specific codons involved and the type of mutations observed. We also examined the frequency of allelic deletion of the p53 gene in cancers containing p53 gene mutations.
Tissue samples from the epithelial ovarian cancers of 62 patients were obtained during initial laparotomy. Histologic examination was done to ensure that the experimental samples used in this study contained more than 75% cancer cells. Total RNA was extracted from these samples and separately from matched control noncancerous regions of the surgical specimen or white blood cells. The purified RNAs were reverse transcribed to generate cDNA copies of exons 4-10 of the p53 gene. Two rounds of polymerase chain reaction (PCR) were conducted to produce enough template for DNA sequence analysis of the regions of interest within the p53 gene. Dideoxy sequencing of at least two independent productions of each amplified DNA template was done to confirm the validity of the mutations found. Allelic deletions were identified by PCR and gel electrophoretic techniques to examine three polymorphisms within the p53 gene in cancer-normal DNA pairs.
We identified 45 mutations in exons 5-8 of the p53 gene, where mutations frequently have been found in other cancer types. An additional mutation was identified in exon 4. Overall, 72% of the mutations were transitions, 24% transversions, and 4% microdeletions. Allelic deletion of the other p53 allele was seen in 67% of ovarian cancers in which a p53 mutation was present. Germ-line p53 mutations were not found in any patients whose cancers had p53 mutations.
Like p53 mutations in other types of human cancers, those in epithelial ovarian cancers are diverse and occur frequently in exons 5-8. The predominance of transition mutations suggests that p53 mutations in ovarian cancer arise because of spontaneous errors in DNA synthesis and repair rather than the direct interaction of carcinogens with DNA. These molecular data are consistent with data from epidemiologic studies that have failed to demonstrate a convincing relationship between exposure to environmental carcinogens and the development of ovarian cancer.
p53基因编码一种在正常人类细胞中低水平存在的核磷蛋白。该蛋白的野生型形式发挥作用抑制不适当的细胞增殖。大约一半的人类上皮性卵巢癌存在p53基因突变并过表达突变蛋白产物。p53基因一个等位基因的缺失在这些癌症中也经常发生。
我们试图确定上皮性卵巢癌中p53基因的突变谱，包括所涉及的具体密码子以及观察到的突变类型。我们还研究了含有p53基因突变的癌症中p53基因等位基因缺失的频率。
在初次剖腹手术期间获取62例患者上皮性卵巢癌的组织样本。进行组织学检查以确保本研究中使用的实验样本含有超过75%的癌细胞。从这些样本以及手术标本的匹配对照非癌区域或白细胞中分别提取总RNA。将纯化的RNA逆转录以生成p53基因外显子4 - 10的cDNA拷贝。进行两轮聚合酶链反应（PCR）以产生足够的模板用于p53基因内感兴趣区域的DNA序列分析。对每个扩增的DNA模板至少两个独立产物进行双脱氧测序以确认所发现突变的有效性。通过PCR和凝胶电泳技术鉴定等位基因缺失，以检查癌症 - 正常DNA对中p53基因内的三种多态性。
我们在p53基因的外显子5 - 8中鉴定出45个突变，这些区域在其他癌症类型中也经常发现突变。在外显子4中鉴定出另外一个突变。总体而言，72%的突变是转换，24%是颠换，4%是微缺失。在存在p53突变的卵巢癌中，67%可见另一个p53等位基因的等位基因缺失。在癌症存在p53突变的任何患者中均未发现种系p53突变。
与其他类型人类癌症中的p53突变一样，上皮性卵巢癌中的p53突变多种多样，且在外显子5 - 8中频繁发生。转换突变占优势表明卵巢癌中的p53突变是由于DNA合成和修复中的自发错误而非致癌物与DNA的直接相互作用引起的。这些分子数据与流行病学研究的数据一致，流行病学研究未能证明接触环境致癌物与卵巢癌发生之间存在令人信服的关系。
