ANALYSIS OF GENETIC ALTERATIONS DURING HEPATOCARCINOGENESIS

肝癌发生过程中的基因改变分析

• 批准号: 3838444
• 负责人: P J WIRTH
• 金额: --
• 依托单位: DIVISION OF CANCER EPIDEMIOLOGY AND GENETICS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3838444
• 关键词: China; Macaca fascicularis; aflatoxins; chemical carcinogen; chemical carcinogenesis; chemical related neoplasm /cancer; chromosomes; colorectal neoplasms; complementary DNA; laboratory rat; messenger RNA; mutagens; neoplasm /cancer genetics; nutrition related neoplasm /cancer; nutrition related tag; point mutation; polymerase chain reaction; quinoline; restriction fragment length polymorphism; tumor suppressor genes

Mutations in the putative p53 tumor suppressor gene were analyzed in a series of 9 aflatoxin B1 (AFLB1) induced monkey tumors, 6 AFLB1 transformed rat liver cell lines, as well as a group of 19 human hepatocellular carcinomas (HCC) obtained from Qidong, China, and 17 HCCs induced by the food mutagen, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) in cynomolgus monkeys. Among the AFLB1 induced monkey tumors only a single point mutation at codon 175 (G to T transversion) was identified in only one HCC by sequencing analysis of the four conserved domains (II to V) in the p53 gene. In 3/17 of the IQ induced monkey tumors the same G to T transversion was detected at codons 159, 175, and 248. Sequence analysis of amplified cDNA by RT-PCR of p53 mRNA from the six AFLB1 transformed rat liver cell lines showed the same point mutation, a transition from G to A in codon 173 (CGC to CAC) which results in a change of the encoded amino acid from arginine to histidine. No wild- type alleles of the p53 gene were observed in any of the AFLB1 cell lines by genomic DNA analysis. The half life of the mutant p53 protein in the AFLB1 transformants was greater than 3 hours versus 30 minutes for normal p53 protein. Analysis of 19 HCCs from Qidong using PCR-SSCP as well as RFLP analysis revealed that 10/19 HCCs (53%) showed p53 mutation. In two tumors single mutations occurred in exons 5 and 6, while in 8 HCCs a G to T mutation at the third position of codon 249 in exon 7 was observed. Loss of heterozygosity (LOH) of the p53 and retinoblastoma (RB) genes was detected in 7/9 (78%) and 3/6 (50%) of informative cases, respectively. No LOH of adenomatis polyposis coli locus was observed in 10 informative cases. LOH of chromosomes 16q and 16p was detected in 10/18 (56%) and 6/13 (46%) informative cases, respectively. These data suggest that the selective mutation and frequent LOH of the p53 play an important role in the development of HCCs from Qidong and at least three additional tumor suppressor genes might be involved in the human hepatocarcinogenesis.

假定的 p53 肿瘤抑制基因的突变在 9 种黄曲霉毒素 B1 (AFLB1) 诱导猴肿瘤系列，6 种 AFLB1 转化的大鼠肝细胞系，以及一组 19 个人类肝细胞系 取自中国启东的肝细胞癌 (HCC) 和 17 个 HCC 由食物诱变剂 2-氨基-3-甲基咪唑[4,5-f]喹啉 (IQ) 诱导 在食蟹猴中。 在 AFLB1 诱导的猴肿瘤中，只有 鉴定出密码子 175 处的单点突变（G 至 T 颠换） 通过对四个保守结构域的测序分析，仅在一个 HCC 中发现了这一点（II 至V)在p53基因中。 3/17的智商诱发猴肿瘤相同 在密码子 159、175 和 248 处检测到 G 到 T 的颠换。 序列 通过 RT-PCR 分析来自 6 个 AFLB1 的 p53 mRNA 的扩增 cDNA 转化的大鼠肝细胞系显示出相同的点突变， 密码子 173 中从 G 到 A 的转变（CGC 到 CAC），导致 编码的氨基酸从精氨酸变为组氨酸。 没有野生- 在任何 AFLB1 细胞系中均观察到 p53 基因的等位基因型 通过基因组 DNA 分析。 突变型 p53 蛋白的半衰期 AFLB1 转化体的时间超过 3 小时，而正常则为 30 分钟 p53 蛋白。 使用 PCR-SSCP 和 PCR-SSCP 分析启东市 19 例 HCC RFLP 分析显示 10/19 HCC (53%) 显示 p53 突变。 分成两份 肿瘤单一突变发生在外显子 5 和 6，而在 8 个 HCC 中，G 到 在外显子 7 的密码子 249 的第三个位置观察到 T 突变。 p53 和视网膜母细胞瘤 (RB) 基因杂合性丢失 (LOH) 为 分别在 7/9 (78%) 和 3/6 (50%) 的信息病例中检测到。 10个资料中未观察到腺瘤性息肉病大肠杆菌位点的LOH 案例。 10/18 (56%) 中检测到染色体 16q 和 16p 的 LOH 分别为 6/13 (46%) 个信息丰富的案例。 这些数据表明 p53 的选择性突变和频繁的 LOH 在 来自启东的 HCC 和至少另外三种肿瘤的发展 抑制基因可能参与人类肝癌的发生。