TUMOR SUPPRESSOR P53 GENE IN VIRAL HEPATOCARCINOMA

编码一种生长调节蛋白的p53基因经常是 在人类癌症中失活，被认为是肿瘤抑制基因。在……里面 探索肝细胞癌发生的潜在分子机制， 我们在原代人肝细胞中发现P53基因失活 癌(PHC)。这些肿瘤中这种突变的频率仍然存在。 有待确定，但在我们的初步研究中，我们发现了p53 在9个PHC来源的细胞系中，有8个出现异常。我们的结果很强劲 提示P53基因失活是一种重要的遗传损伤 在肝细胞癌(HCC)中。我们正在开始研究以确定 P53基因失活与其他可疑协作性疾病的关系 肝细胞癌发生过程中的因素，特别是 乙肝病毒(乙肝病毒)。我们推测乙肝病毒可能在肝细胞癌中起作用。 通过直接改变P53基因的结构或表达。 另一种可能是P53功能失活和慢性乙肝病毒感染 可能在肝细胞癌的多步骤过程中起累积作用 转型。我们将通过确定频率开始我们的研究 PHCS中P53基因失活。肿瘤，以及未受影响的肝脏样本 将在蛋白质水平上进行分析，使用 免疫化学方法；对p53基因的研究将包括限制 基因组DNA的酶分析和保守区的测序 聚合酶链式反应扩增。P53基因阳性的PHCS 异常将用于制备基因组文库，从基因组文库 P53基因以及乙肝病毒整合位点将被克隆和测序。 为进一步探讨P53、HBV3和P53之间的关系。 肝细胞癌的发生，我们将进行转基因研究 三个PHC来源的细胞系：“最小偏差”的Hep G2细胞， 似乎表达‘野生型’P53、Hep 3B细胞，这些细胞没有内源性 P53和HuH-7细胞，表达突变的P53蛋白。这些细胞系 将用表达‘野生型’或突变型P53的载体 和/或乙肝病毒基因组。将对转基因细胞进行后续评估 表型变化，如生长能力和致瘤性的改变。 通过我们证明p53基因在肝细胞癌中失活，我们有 为我们的肝细胞研究开辟了新的视角 致癌。我们的目标是确定乙肝病毒是否与病毒直接相互作用 P53的抑癌功能独立地参与了 转型过程。我们相信，这些研究将是有益的。 在阐明肝细胞癌的发病机制方面。

The p53 gene, which encodes a growth regulatory protein, is frequently inactivated in human cancer and is considered a tumor suppressor gene. In exploring potential molecular mechanisms of hepatocellular carcinogenesis, we discovered p53 gene inactivations in primary human hepatocellular carcinoma (PHC). The frequency of such mutations in these tumors remains to be determined, but in our preliminary studies, we found p53 abnormalities in 8 out of 9 PHC-derived cell lines. Our results strongly suggest that the inactivation of p53 gene is a significant genetic lesion in hepatocellular carcinoma (HCC). We are beginning studies to determine the relationship between p53 gene inactivation and other suspected co- factors in the process of hepatocellular carcinogenesis, particularly Hepatitis B Virus (HBV). We hypothesize that HBV could play a role in HCC by directly altering the structure or expression of p53 gene. Alternatively, inactivation of p53 function and chronic infection with HBV may have cumulative effects in the multi-step process of hepatocellular transformation. We will begin our studies by determining the frequency of p53 gene inactivation in PHCs. Tumors, as well as uninvolved liver samples from the same patients, will be analyzed at the protein level using immunochemical methods; studies of the p53 gene will include restriction enzyme analysis of genomic DNA and sequencing of conserved regions after polymerase chain reaction amplification. PHCs with demonstrated p53 gene abnormalities will be used for preparation of genomic libraries, from which p53 genes as well as HBV integration sites will be cloned and sequenced. To further investigate the relationship between p53, HBV, and hepatocellular carcinogenesis, we will conduct transfection studies on three PHC-derived cell lines: 'minimally deviant' Hep G2 cells, which appear to express 'wide-type' p53, Hep 3B cells, which have no endogenous p53, and Huh 7 cells, which express a mutant p53 protein. These cell lines will be transfected with plasmids expressing 'wild-type' or mutant p53 and/or the HBV genome. Transfected cells will be assessed for consequent phenotypic changes, such as altered growth capability and tumorigenicity. Through our demonstration that the p53 gene is inactivated in HCC, we have developed a new perspective in our studies of hepatocellular carcinogenesis. We aim to determine whether HBV interacts directly with the tumor suppressor function of p53 or contributes independently to the transformation process. We believe that these studies will be instrumental in clarifying the pathogenesis of HCC.