Rad9-Based Mouse Model of Prostate Carcinogenesis

描述（由申请人提供）： 当细胞暴露于损害DNA的物理或化学试剂时，可能会产生有害影响，包括突变，癌症或死亡。然而，有机制可以修复损伤，稳定基因组并中和有害影响。我们从酵母、小鼠和人类中分离并鉴定了一个进化上保守的基因Rad 9，它对维持基因组稳定性和促进细胞对DNA损伤的抵抗力至关重要。基于这些以及其他发现，我们研究了Rad 9在致癌作用中的作用，并发现了一个显着的联系，特别是前列腺癌。我们的研究结果包括观察到Rad 9在四种常用的前列腺癌细胞系CWR 22，DU 145，LNCaP和PC-3中显著过表达，相对于正常的人前列腺PrEC细胞和永生化但非致瘤细胞系PWRE-1。我们还发现siRNA降低Rad 9蛋白水平的程度与这些细胞注射到裸鼠后致瘤性的降低程度相关。此外，非致瘤性永生化PWRE-1细胞可以在裸鼠中形成异常生长，如果细胞被工程化以过量产生Rad 9。此外，我们在339例（45.1%）人前列腺癌中的153例中检测到高Rad 9蛋白水平，但在52例正常前列腺组织对照中的2例中检测到低丰度。相对较高水平的Rad 9蛋白通常与更晚期的疾病相关。因此，我们的研究结果表明，Rad 9在前列腺癌发生中起着重要作用。该项目的重点是Rad 9蛋白异常高丰度导致前列腺癌的假设。主要的实验目标是建立和表征一种新的小鼠模型的前列腺癌，特别是在该器官的异常高表达的Rad 9的基础上。为此，我们有初步的数据表明，这样一个系列的小鼠已经建成。本研究的具体目的集中在1）检查亲代以及前列腺中具有高水平Rad 9的转基因小鼠的前列腺组织关于作为年龄的函数的自发性以及辐射诱导的前列腺组织异常和肿瘤发生的形成; 2）确定高水平Rad 9与自发性以及电离辐射诱导的转移性前列腺癌之间的关系;和3）确定高水平的Rad 9介导前列腺疾病的能力与Rad 9的反式作用功能的下游靶基因的表达以及已确定密切参与前列腺癌的其它基因的表达之间的关系，从而开始理解Rad 9介导的前列腺癌发生的机制基础。最后，本项目将提供一个独特的模型，以确定前列腺癌的分子机制，并作为一个体内系统，阐明环境致癌的风险。公共卫生相关性：该提案侧重于Rad 9在介导前列腺癌发生中的作用。我们的目标是设计和表征一种独特的小鼠模型，该模型基于前列腺中高水平的Rad 9蛋白，证明了这种癌症的遗传易感性。将评估前列腺疾病的自发性和辐射诱导性发展。因此，该项目很重要，因为其结果可能会影响人类健康，无论是在前列腺癌的治疗方面，还是在了解遗传学和环境与致癌作用的相互作用方面。

DESCRIPTION (provided by applicant): When cells are exposed to physical or chemical agents that damage DNA, deleterious effects can ensue, including mutation, cancer or death. However, mechanisms are available to repair the damage, stabilize the genome and neutralize harmful effects. We isolated and characterized an evolutionarily conserved gene called Rad9, from yeast, mouse and human, which is critical for maintaining genomic stability and promoting cellular resistance to DNA damage. Based on these as well as other findings, we examined the role of Rad9 in carcinogenesis, and found a significant link in particular to prostate cancer. Our results include the observation that Rad9 is dramatically overexpressed in four commonly used prostate cancer cell lines, CWR22, DU145, LNCaP and PC-3, relative to normal human prostate PrEC cells and the immortalized but nontumorigenic cell line PWRE-1. We also found that the degree to which siRNA reduces Rad9 protein levels correlates with the extent of decrease in tumorigenicity of these cells after injection into nude mice. Moreover, nontumorigenic immortalized PWRE-1 cells can form aberrant growths in nude mice if the cells are engineered to overproduce Rad9. In addition, we detected high Rad9 protein levels in 153 of 339 (45.1%) human prostate adenocarcinomas, but just low abundance in 2 of 52 normal prostate tissue controls. Relatively higher levels of Rad9 protein were in general associated with more advanced disease. Our results thus indicate that Rad9 plays a functional role in prostate carcinogenesis. This project focuses on the hypothesis that aberrantly high abundance of Rad9 protein leads to prostate cancer. The major experimental goal is to create and characterize a novel mouse model of prostate carcinogenesis, based on aberrant high expression of Rad9 specifically in that organ. To this end, we have preliminary data indicating that such a line of mouse has been constructed. Specific aims of the study focus on 1) examining prostate tissue of parental, as well as transgenic mice with high levels of Rad9 in prostate with respect to spontaneous as well as radiation-induced formation of prostate tissue abnormalities and tumorigenesis as a function of age; 2) determine the relationship between high levels of Rad9 and spontaneous as well as ionizing radiation-induced metastatic prostate cancer; and 3) defining the relationship between the ability of high levels of Rad9 to mediate prostate disease and expression of downstream target genes of the transactivity function of Rad9 as well as expression of other genes well-established to be intimately involved in prostate cancer, to begin to understand the mechanistic basis for Rad9 mediated prostate carcinogenesis. In the end, this project will provide a unique model to define molecular mechanisms of prostate carcinogenesis, and serve as an in vivo system for elucidating risk o environmental carcinogenesis. PUBLIC HEALTH RELEVANCE: This proposal focuses on the role of Rad9 in mediating prostate carcinogenesis. The goal is to engineer and characterize a unique mouse model demonstrating genetic predisposition to this kind of cancer, based on high levels of Rad9 protein in prostate. The spontaneous as well as radiation-induced development of prostate disease will be assessed. As such, this project is important since the results could impact on human health, both in terms of treatment for prostate cancer and understanding the interplay of genetics and the environment with respect to carcinogenesis.