Project 2: To determine the consequences of activating Rb function in cancer cells

项目摘要 在人类癌症中，编码Rb肿瘤抑制基因的基因经常被沉默、缺失、突变或缺失。 通过过表达的细胞周期蛋白D-Cdk 4/6激酶复合物磷酸化而功能失活。由于其 由于细胞周期蛋白D-Cdk 4/6-Rb轴在癌症中的重要性，它是FDA批准的Cdk 4/6抑制剂的靶标。的 这种努力的基本假设是Cdk 4/6抑制导致Rb重新激活和肿瘤抑制， 编码Rb的基因未被破坏的癌细胞。虽然大量的研究表明， 研究了Rb功能丧失的后果，很少有研究研究Rb再- 细胞中的激活，即使这种再激活在用Cdk 4/6治疗的患者中在临床上高度相关 抑制剂的 在这里，我们建议解决Rb活性抑制癌症的机制这一核心问题 使用遗传学、细胞生物学、生物化学和结构方法的组合进行进展。一是 使用新的可诱导小鼠等位基因测定Rb失活癌细胞中Rb再活化的体内效应 其中Rb功能可以打开和关闭。使用这个等位基因，我们将重新激活肿瘤细胞中的Rb，以确定 Rb重新引入的分子后果，作为一种更好地了解Rb野生型癌细胞 对Cdk 4/6抑制剂有反应。其次，我们将研究Rb家族可能的肿瘤抑制作用 成员p107和p130对Rb突变的癌细胞中Cdk 4/6抑制剂的应答。目标是 鉴定增强p107和p130对细胞周期抑制的方法。第三，我们将研究翻译后 控制Rb与E2 F相互作用及其在细胞中的肿瘤抑制作用的修饰，包括乙酰化 与Rb的磷酸化功能性相互作用的事件。鉴定这些关键残基， Rb的修饰可能会开辟新的研究途径，以防止其功能失活或增强其肿瘤 抑制活性。 综上所述，这些实验将提供一个深入的分析功能激活的Rb和其 在与癌症相关的背景下，家族成员可能指向新的效应机制下游， 与Rb平行。最终，这些实验将有助于确定增强肿瘤抑制效果的新方法。 FDA批准的Cdk 4/6抑制剂，如palbociclib，ribociclib或abemaciclib。

PROJECT SUMMARY In human cancers, the gene coding for the Rb tumor suppressor is frequently silenced, deleted, mutated, or functionally inactivated by phosphorylation by over-expressed Cyclin D-Cdk4/6 kinase complexes. Due to its importance in cancer, the Cyclin D-Cdk4/6-Rb axis is the target of FDA-approved Cdk4/6 inhibitors. The underlying assumption of this effort is that Cdk4/6 inhibition leads to Rb re-activation and tumor suppression in cancer cells where the gene coding for Rb has not been disabled. While a large number of studies have investigated the consequences of Rb loss of function, few studies have investigated the consequences of Rb re- activation in cells, even though this re-activation is highly relevant in the clinic in patients treated with Cdk4/6 inhibitors. Here, we propose to tackle the central question of the mechanisms by which Rb activity can suppress cancer progression using a combination of genetic, cell biological, biochemical, and structural approaches. First, we will determine the effects of Rb re-activation in Rb-inactive cancer cells in vivo using a novel inducible mouse allele in which Rb function can be turned on and off. Using this allele, we will re-activate Rb in tumor cells to determine the molecular consequences of Rb re-introduction as a way to better understand how Rb wild-type cancer cells respond to Cdk4/6 inhibitors. Second, we will investigate the possible tumor suppressor role of the Rb family members p107 and p130 in response to Cdk4/6 inhibitors in cancer cells that are mutant for Rb. The goal is to identify ways to enhance cell cycle inhibition by p107 and p130. Third, we will investigate post-translational modifications that control Rb interactions with E2F and its tumor suppressive effects in cells, including acetylation events that functionally interact with phosphorylation of Rb. The identification of such key residues that are modified in Rb may open new avenues of research to prevent its functional inactivation or enhance its tumor suppressive activity. Taken together, these experiments will provide an in-depth analysis of the functional activation of Rb and its family members in contexts relevant to cancer and may point to new effector mechanisms downstream of and parallel to Rb. Ultimately, these experiments will help identify new ways to enhance the tumor suppressive effects of FDA-approved Cdk4/6 inhibitors such as palbociclib, ribociclib, or abemaciclib.