Understanding and Controlling p120 Dysfunction in CRC

结直肠癌的肿瘤进展涉及遗传和表观遗传的积累 最终导致恶变的变化。E-钙粘素下调频繁且广泛 被认为是向转移过渡的关键事件。在大多数E-钙粘附素缺陷型癌中， P120异常定位于细胞质，E-钙粘素丢失/胞浆p120密切相关 预后不良。P120本身在CRC的子集中下调，但其意义尚不清楚。我们 已经发现在体外和体内p120消融会破坏E-钙粘附素(和相关的连环蛋白)的稳定性，导致 细胞形态和粘附性严重缺陷。有趣的是，在体外，p120-消融在许多细胞类型中 诱导Rho的结构性激活，在无血清的情况下细胞生长，并失去接触抑制。在……里面 体内，类似的效应导致ROCK/NFKB/COX-2信号级联的细胞自主激活和 慢性炎症与非甾体抗炎药和环氧合酶-2抑制剂疗效的潜在相关性观察 在CRC。我们的数据提示p120和E-钙粘蛋白之间新的相互依赖的相互作用通过 调节Rho以抑制转移和炎症。进度报告描述了这些影响， 以及对Rho、p120和E-钙粘蛋白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结直肠癌的肿瘤进展和临床干预。在目标1中，我们将 评估简化的p120 KO小鼠模型，以确定药物干预是否在 P120(或E-钙粘附素)下调激活的通路可以抑制肿瘤进展，肿瘤生长， 或转移。目标2根据新的观察结果探索了两个不同的假设。首先，要检查 P120丢失、错配修复(MMR)缺陷CRC与TGF0IIR缺陷之间的隐含关系 信号，我们将在注释良好的MMR缺陷组和MMR缺陷组之间进行逐个比较 熟练的肿瘤。其次，我们将跟进在mrna水平上p120下调的新观察结果。 在高级CRC中。我们将确定mRNA水平是否在大肠癌进展的早期阶段下降，是否 这种现象在任何水平上都与肿瘤的类型或结果有关，并且在 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来 询问与p120依赖相关的特征良好但尚不清楚的信号通路 E-钙粘附素失活。与比彻姆实验室(项目2)和范德比尔特研究所合作 化学生物学(VICB)高通量筛选设备，我们开发了高通量小规模 分子筛查(HTS)以确定挽救结直肠癌模型细胞系中E-钙粘蛋白功能的新化合物。 HTS的应用代表了一种描述控制信号通路的创新方法 P1207E-钙粘附素的功能，并可能导致鉴定新的能够抑制肿瘤的化合物 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.