Understanding and Controlling p120 Dysfunction in CRC

结直肠癌（CRC）的肿瘤进展涉及遗传和表观遗传的积累， 最终导致恶性肿瘤的变化。E-钙粘蛋白下调频繁发生，并广泛存在于 被认为是向转移转变的关键事件。在大多数E-钙粘蛋白缺陷型CRC中， p120异常定位于细胞质，并且E-钙粘蛋白丢失/细胞质p120强烈相关 预后不良。p120本身在CRC的一个子集中下调，但其意义尚不清楚。我们 已经发现体外和体内p120消融使E-钙粘蛋白（和相关的连环蛋白）不稳定， 细胞形态和粘附的严重缺陷。有趣的是，在许多细胞类型中的体外p120消融 诱导Rho的组成性激活、在无血清情况下的细胞生长和接触抑制的丧失。在 在体内，类似的作用导致ROCK/NF κ B/考克斯-2信号级联的细胞自主激活， 慢性炎症，与NSAIDS和考克斯-2抑制剂疗效潜在相关性的观察 在《儿童权利公约》。我们的数据表明，p120和E-钙粘蛋白之间的新型相互依赖作用， 调节Rho以抑制转移和炎症。进度报告描述了这些影响， 沿着的是对Rho、p120和E-cadherin之间关系的一种新的分子解释。目标 （下文）寻求将我们的发现应用于CRC的肿瘤进展和临床干预。在目标1中， 评估简化的p120 KO小鼠模型，以确定药物干预是否在 由p120（或E-钙粘蛋白）下调激活的途径可以抑制肿瘤进展，肿瘤生长， 或转移。目的2探讨了基于新观察的两个独立假设。第一，检查 p120缺失、错配修复（MMR）缺陷型CRC和TGF β 1 IR缺陷之间的隐含关系 信号，我们将进行一个头对头之间的比较，以及注释组的MMR缺陷和 熟练的肿瘤。其次，我们将在mRNA水平上追踪p120下调的新观察结果。 晚期CRC我们将确定mRNA水平是否在CRC进展的早期阶段下降， 这种现象在任何水平上与肿瘤类型或结果相关， 分子水平上的因果关系。最后，在目标3中，我们将使用化学遗传学， 询问一个充分表征的但未知的与p120依赖性 E-钙粘蛋白失活。与Beauchamp实验室（项目2）和范德比尔特研究所合作， 化学生物学（VICB）高通量筛选设施，我们开发了一种高通量小 分子筛选（HTS）以鉴定在CRC模型细胞系中拯救E-钙粘蛋白功能的新型化合物。 HTS的应用代表了一种创新的方法来描绘信号通路，控制 p1207 E-钙粘蛋白功能，并可能导致能够抑制肿瘤的新化合物的鉴定 进展或转移。

Tumor progression in colorectal cancer (CRC) involves accumulation of genetic and epigenetic changes that ultimately lead to malignancy. E-cadherin downregulation occurs frequently and is widely believed to be a pivotal event in the transition to metastasis. In the majority of E-cadherin-deficient CRCs, p120 localizes aberrantly to the cytoplasm, and E-cadherin-loss / cytoplasmic p120 is strongly associated poor prognosis. p120 itself is downregulated in a subset of CRC but the significance is not yet clear. We have found that p120 ablation in vitro and in vivo destabilizes E-cadherin (and associated catenins) causing severe defects in cell morphology and adhesion. Interestingly, in vitro p120-ablation in many cell types induces constitutive activation of Rho, cell growth in the absence of serum, and loss of contact inhibition. In vivo, similar effects lead to cell autonomous activation of a ROCK/NFKB/COX-2 signaling cascade and chronic inflammation, an observation of potential relevance to the efficacy of NSAIDS and COX-2 inhibitors in CRC. Our data suggest novel co-dependent interactions between p120 and E-cadherin that act through regulation of Rho to suppress metastasis and inflammation. The progress report describes these effects, along with a novel molecular explanation for the relationship between Rho, p120, and E-cadherin. The aims (below) seek to apply our findings to tumor progression and clinical intervention in CRC. In aim 1, we will evaluate a simplified p120 KO mouse model to ascertain whether pharmaceutical intervention at the level of pathways activated by p120 (or E-cadherin) downregulation can suppress tumor progression, tumor growth, or metastasis. Aim 2 explores two separate hypotheses based on novel observations. First, to examine implied relationships between p120 loss, mismatch repair (MMR) deficient CRC, and defects in TGF0IIR signaling, we will perform a head-to-head comparison between well-annotated groups of MMR-deficient and proficient tumors. Second, we will follow up on novel observations of p120 downregulation at the mRNA level in advanced CRC. We will determine whether mRNA levels fall at early stages of CRC progression, whether this phenomenon is associated at any level with tumor type or outcome, and the significance at the molecular level with respect to cause and effect. Finally, in aim 3 we will use chemical genetics to interrogate a well characterized but as yet unknown signaling pathway associated with p120-dependant inactivation of E-cadherin. In collaboration with the Beauchamp lab (project 2) and the Vanderbilt Institute for Chemical Biology (VICB) high throughput screening facility, we have developed a high throughput small molecule screen (HTS) to identify novel compounds that rescue E-cadherin function in CRC model cell lines. The application of HTS represents an innovative approach to delineating signaling pathways that control p1207E-cadherin function, and could lead to identification of novel compounds capable of suppressing tumor progression or metastasis.