Using p53 Knock-In Mice to Understand p53's Role in Pancreatic Cancer

描述（由申请人提供）：胰腺癌是癌症相关死亡的第四大原因，几乎总是致命的疾病。这种癌症从侵袭前状态到转移、侵袭状态的进展与p53肿瘤抑制基因的突变有关。虽然p53在应激信号下通过诱导细胞周期阻滞、衰老或凋亡来抑制癌症，但其在体内作用的分子机制尚不清楚。p53表现出作为转录激活因子的活性，但它也具有多种其他生化活性，如调节转录抑制、DNA复制和重组、线粒体膜完整性和中心体复制。为了开始定义p53分子活性对抑制胰腺癌进展的重要性，我们建议通过生成和分析表达不同p53突变体的p53敲入小鼠品系，来解决p53转录激活和抑制功能对肿瘤抑制的重要性。在敲入小鼠中，突变体在天然p53启动子的控制下表达，正常细胞可以在离体和生物体的生理环境中进行研究，这应该为p53在肿瘤抑制中的作用机制提供明确的见解。我们之前产生了表达p53突变体p5325,26的敲入小鼠，该突变体在大多数（但不是全部）p53靶基因的反活化方面受到严重损害，并保留了抑制肺癌发展的能力。这些观察结果表明，p5325,26的肿瘤抑制功能可能是由残留的p53转激活或其他p53活性引起的。我们将首先使用一个由活化的KRas表达驱动的具有良好特征的胰腺癌小鼠模型来检测该突变体抑制胰腺癌进展的能力，从而确定是否需要完全p53反激活潜能来抑制胰腺中的肿瘤。如果p5325,26显示出活性，那么通过检测表达完全缺陷的突变体p5325,26,53,54的敲入小鼠，可以区分有限的转激活与另一种p53功能对肿瘤抑制的重要性。此外，转录抑制对p53功能在阻断胰腺癌进展中的重要性将通过生成表达p53抑制缺陷变体的小鼠来评估。我们建议进一步开发细胞培养模型系统，以更详细地研究p53介导的胰腺肿瘤抑制的机制。总之，这些研究将确定基因是否必须被p53激活和/或抑制，或者其他p53活性是否对其抑制胰腺癌发展的能力至关重要，并将对未来开发更好的癌症治疗这种致命疾病有很大的用处。

DESCRIPTION (provided by applicant): Pancreatic cancer is the 4th leading cause of cancer-associated deaths, and it is virtually always a fatal disease. The progression of this cancer from a preinvasive state to a metastatic, invasive state is associated with mutation of the p53 tumor suppressor gene. While p53 suppresses cancer by inducing cell cycle arrest, senescence or apoptosis in response to stress signals, its molecular mechanism of action in vivo remains unclear. p53 displays activity as a transcriptional activator, but it also has a variety of other biochemical activities, in regulating transcriptional repression, DNA replication and recombination, mitochondrial membrane integrity, and centrosome duplication. To begin to define the p53 molecular activities important for suppressing pancreatic cancer progression, we propose to address the importance of p53 transcriptional activation and repression functions for tumor suppression through the generation and analysis of a panel of p53 knock-in mouse strains expressing different p53 mutants. The use of knock-in mice, in which mutants are expressed under the control of the native p53 promoter, and in which normal cells can be studied both ex vivo and in the physiological context of the organism, should provide definitive insight into the mechanism of p53 action in tumor suppression. We previously generated knock-in mice expressing a p53 mutant, p5325,26, that is severely impaired for the transactivation of most, but not all, p53 target genes and that retains the ability to suppress lung cancer development. These observations suggest that p5325,26 tumor suppressor function is accounted for by either residual p53 transactivation or another p53 activity. We will first examine the ability of this mutant to inhibit pancreatic cancer progression using a well-characterized mouse model of pancreatic cancer driven by activated KRas expression, and we will thus determine if full p53 transactivation potential is required for tumor suppression in the pancreas. If p5325,26 displays activity, the importance of limited transactivation versus another p53 function for tumor suppression will be distinguished by examining knock-in mice expressing a mutant totally defective for transactivation, p5325,26,53,54. In addition, the importance of transcriptional repression for p53 function in blocking pancreatic cancer progression will be assessed by generating mice expressing a repression-defective variant of p53. We propose further to develop a cellular culture model system for more detailed mechanistic studies of p53-mediated tumor suppression in the pancreas. Together, these studies will determine whether genes must be activated and/or repressed by p53, or whether other p53 activities are essential for its ability to suppress pancreatic cancer development, and will have great utility for the future development of better cancer therapies for this deadly disease. PUBLIC HEALTH RELEVANCE: Pancreatic cancer is the 4th leading cause of cancer deaths in the US and is nearly always fatal. It is therefore very important to understand the basis of this disease in order to be able to more effectively treat and cure it. This research aims to elucidate how a gene known as p53 acts to prevent the development of pancreatic cancer, with the ultimate goal of devising better prevention and treatment strategies for cancer.