Elucidating the Relationship Between Cell Proliferation and Invasion

项目摘要/摘要 在过去的五年中，三种细胞周期蛋白依赖性激酶4/6抑制剂(CDK4/6i)已被批准用于 转移性乳腺癌的治疗。CDK4/6I通过阻断癌细胞的进展来阻止其分裂 通过细胞周期的G1/G0期。然而，体外研究的结果表明，外源和 内源性CDK4/6I增加了某些形式癌症的侵袭性。G1/G0细胞的后果 在体内对癌细胞行为的周期停滞仍不清楚。秀丽线虫锚定细胞(AC)的侵袭 提供了一种出色而简单的体内细胞侵袭行为模型，允许单细胞和亚细胞 细胞通过基底膜侵袭的分析。我们的实验室确定需要特定于G1/G0的逮捕 核激素受体NHR-67/TLX对AC的侵袭起着关键的调节作用 进程。在没有NHR-67的情况下，AC变为非侵袭性的，但过度增殖，表明 当入侵被阻止时，扩散可能会增加。我们的长期目标是理解这两种分子 CDK4/6I单独或联合应用对G1/G0细胞周期停滞的调控机制及体内效应 与哺乳动物雷帕霉素靶标(MTORi)的抑制剂联合使用。CDK4/6i和mTORi共同拥有 已被证明在体外可以诱导癌细胞衰老，这表明将这些药物结合起来 药物在临床环境中可能是有用的。本奖学金申请的总体目标是：(I) 阐明线虫AC中G1/G0是如何维持的，以及(Ii)确定NHR-67缺陷的AC中是否存在ACS 可以通过药物诱导动物从增殖到入侵再到衰老。基于 初步数据和先前的研究，中心假设是冗余机制调节G1/G0 NHR-67基因缺陷动物的野生型AC和增殖性AC的状态可被提示经历 衰老。这一核心假设将通过追求两个具体目标来检验。目标1将确定 调节前侵袭性AC中G1/G0停滞的成分。目标2将评估AC和AC之间的过渡 CDK4/6和/或mTOR缺失时的增殖、侵袭和衰老。为了实现这些目标，这 该项目将使用成熟的基因敲除技术、高分辨率成像和尖端技术 技术，包括一个新的活细胞周期状态传感器和两个条件蛋白质耗尽系统。这个 这些研究的结果可以帮助指导未来侵袭性癌症治疗策略的发展。

PROJECT SUMMARY/ABSTRACT Over the last five years, three cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) have been approved for the treatment of metastatic breast cancer. CDK4/6i prevent cancer cells from dividing by blocking their progression through the G1/G0 phase of the cell cycle. However, results from in vitro studies reveal that both exogenous and endogenous CDK4/6i increase the invasiveness of certain forms of cancer. The consequences of G1/G0 cell cycle arrest on cancer cell behavior in vivo remain unknown. Caenorhabditis elegans anchor cell (AC) invasion provides an excellent yet simple in vivo model of cell invasive behavior that allows single-cell and subcellular analysis of cell invasion through basement membrane. Our lab determined that G1/G0-specific arrest is required for AC invasion to occur and that the nuclear hormone receptor NHR-67/TLX acts as a key regulator of this process. In the absence of NHR-67, the AC becomes non-invasive but over-proliferates, indicating that proliferation can increase as invasion is blocked. The long-term goal is to understand both the molecular mechanisms controlling G1/G0 cell cycle arrest and the in vivo effects of CDK4/6i as individual agents and in combination with inhibitors of the mammalian target of rapamycin (mTORi). Together, CDK4/6i and mTORi have been shown to induce senescence in cancer cells in vitro, suggesting that combining these pharmacological agents might be useful in the clinical setting. The overall objectives of this fellowship application are to (i) elucidate how G1/G0 is maintained in the C. elegans AC and (ii) determine whether ACs in NHR-67-deficient animals can be pharmacologically induced to transition from proliferation-to-invasion-to-senescence. Based on preliminary data and prior research, the central hypothesis is that redundant mechanisms regulate the G1/G0 state of the wild-type AC and that proliferative ACs in NHR-67-deficient animals can be prompted to undergo senescence. This central hypothesis will be tested by pursuing two specifics aims. Aim 1 will identify the components regulating G1/G0 arrest in the pro-invasive AC. Aim 2 will evaluate the transition between AC proliferation, invasion and senescence upon depletion of CDK4/6, mTOR, or both. To address these aims, this project will use established gene knockdown techniques, high-resolution imaging, and cutting-edge technologies, including a novel live cell cycle state sensor and two conditional protein depletion systems. The results of these studies can help guide the development of future treatment strategies for aggressive cancers.