Requirements for Normal Rb Function In Vivo

视网膜母细胞瘤肿瘤抑制基因(Rb1)突变导致儿童癌症视网膜母细胞瘤，并与大多数类型的人类癌症有关。尽管它在肿瘤发生中起着核心作用，但针对这一途径的治疗方法开发得很慢。对Rb1介导的肿瘤抑制的分子机制的详细了解将有助于此类治疗方法的发展。这个持续项目的长期目标是识别和描述这些分子机制。Rb1蛋白(PRB)是细胞周期的负性调节因子，这种细胞周期调节活性有助于抑制肿瘤生长。多种机制可能参与了pRB介导的细胞周期控制，包括pRB与E2F转录因子结合，导致细胞周期所需基因的转录抑制。另一种可能的机制包括pRb靶向降解Skp2，从而稳定p27Kip1。PRb调节E2F转录因子的能力与其降解Skp2的能力在基因上是可以分离的。我们推测pRb介导的Skp2的降解和p27Kip1的稳定有助于体内前列腺癌的抑制。我们的工作模型是，p27Kip1(P27)的翻译后稳定通常会对致癌压力产生类似衰老的反应，从而抑制肿瘤的发生和发展。在没有pRb的情况下，这种肿瘤抑制反应是沉默的。该应用程序的目标是使用体外和体内实验模型系统来验证该假说。我们已经创造了一个含有突变的Rb1等位基因(R654W)的小鼠品系，其编码的蛋白缺乏E2F结合和调节，但可能能够靶向Skp2进行降解。我们建议使用这个突变的等位基因来评估pRb/E2F和pRb/p27机制在体内对前列腺癌抑制的相对贡献。提出了两个特定的目标：1)检测R654W pRb是否能抑制小鼠前列腺癌的形成；2)表征pRb/Skp2/p27机制在 R654W pRb抑瘤活性。这些特定目标的成功完成将确定有助于抑制Rb1肿瘤的新机制，并提出治疗前列腺癌的新治疗策略。

Mutation of the retinoblastoma tumor suppressor gene (Rb1) causes the pediatric cancer retinoblastoma and contributes to most types of human cancer. Despite its central role in tumorigenesis, therapies designed to target this pathway have been slow to develop. A detailed understanding of the molecular mechanisms underlying Rb1 mediated tumor suppression will facilitate the development of such therapies. The long-term goal of this continuing project is to identify and characterize these molecular mechanisms. Rb1 protein (pRb) is a negative regulator of the cell cycle, and this cell cycle regulatory activity contributes to tumor suppression. Multiple mechanisms likely contribute to pRb mediated cell cycle control, including the well characterized mechanism involving binding of pRb to E2F transcription factors resulting in transcriptional repression of genes required for cell cycle progression. Another possible mechanism involves targeted degradation of Skp2 by pRb and consequent stabilization of p27Kip1. The ability of pRb to regulate E2F transcription factors is genetically separable from its ability to degrade Skp2. We hypothesize that pRb mediated degradation of Skp2 and stabilization of p27Kip1 contributes to prostate tumor suppression in vivo. Our working model is that post translational stabilization of p27Kip1 (p27) normally enforces a senescence like response to oncogenic stress, thereby curtailing tumor initiation and progression. In the absence of pRb, this tumor suppressive response is muted. The goal of this application is to test the hypothesis using in vitro and in vivo experimental model systems. We have created a mouse strain containing a mutant Rb1 allele (R654W) whose encoded protein is deficient for E2F binding and regulation, but may be competent to target Skp2 for degradation. We propose to use this mutant allele to assess the relative contribution of the pRb/E2F and pRb/p27 mechanisms to prostate tumor suppression in vivo. Two specific aims are proposed: 1) Test whether R654W pRb can suppress prostate tumorigenesis in the mouse; 2) Characterize the contribution of the pRb/Skp2/p27 mechanism to R654W pRb tumor suppressor activity. Successful completion of these specific aims will identify a new mechanism contributing to Rb1 tumor suppression and suggest new therapeutic strategies to treat prostate cancer.