Qian Geng-Sun, Kuang Shuang-Yuan, He Xia, Groopman John D, Jackson Peta E
Department of Environmental Health Sciences, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland 21205, USA.
Cancer Epidemiol Biomarkers Prev. 2002 Oct;11(10 Pt 1):1126-9.
Hepatocellular carcinoma (HCC) has several major etiological risk factors, including infection with hepatitis viruses and exposure to aflatoxin B(1). A specific missense mutation resulting from a guanine to thymine transversion at the third position of codon 249 in the p53 tumor suppressor gene has been reported in 10-70% of HCCs from areas of high dietary exposure to aflatoxin B(1.) This mutation has not only been detected in tumor samples but has also been measured in DNA isolated from the blood of patients with HCC in two separate studies by two independent methods: RFLP and short oligonucleotide mass analysis (SOMA), an electrospray ionization mass spectrometry technique. To compare the relative sensitivities of these methodologies, a set of serially diluted samples was analyzed by both techniques. The detection limits of RFLP and SOMA were 6% and 2.4% mutant alleles in the presence of wild-type alleles, respectively. When the DNA samples were predigested with HaeIII before SOMA, the detection limit was improved to 0.4% mutant allele in the presence of wild-type alleles. We have therefore found that SOMA is about 2.5-15-fold more sensitive than RFLP for detection of specific p53 mutations. A set of 26 DNA samples from HCC and normal liver was analyzed by RFLP and SOMA, and 5 samples were positive for the p53 mutation. An additional 4 samples were found to be positive for the mutation when SOMA was repeated after HaeIII predigestion.
肝细胞癌（HCC）有几个主要的病因风险因素，包括感染肝炎病毒和接触黄曲霉毒素B1。在膳食中大量接触黄曲霉毒素B1地区的10%-70%的HCC中，已报道p53肿瘤抑制基因密码子249第三位发生鸟嘌呤到胸腺嘧啶的颠换导致的特定错义突变。这种突变不仅在肿瘤样本中被检测到，而且在两项独立研究中，通过两种独立方法：限制性片段长度多态性（RFLP）和短寡核苷酸质量分析（SOMA，一种电喷雾电离质谱技术），在从HCC患者血液中分离的DNA中也被检测到。为了比较这些方法的相对灵敏度，用这两种技术分析了一组系列稀释的样本。在存在野生型等位基因的情况下，RFLP和SOMA的检测限分别为6%和2.4%的突变等位基因。当DNA样本在SOMA之前用HaeIII预消化时，在存在野生型等位基因的情况下，检测限提高到0.4%的突变等位基因。因此，我们发现SOMA在检测特定p53突变方面比RFLP敏感约2.5至15倍。用RFLP和SOMA分析了一组来自HCC和正常肝脏的26个DNA样本，5个样本p53突变呈阳性。当在HaeIII预消化后重复进行SOMA时，又发现4个样本突变呈阳性。
