Quantifying Biochemical Communication in Hepatocellular Carcinoma

摘要 在美国，肝细胞癌（HCC）的发病率一直在稳步上升。一种解释 因为这是肥胖的惊人趋势。发展与非发展之间存在正相关关系， 酒精性脂肪肝（NAFLD）和肝细胞癌已被确定在临床上，支持之间的联系， 肥胖和HCC。HCC的公共卫生影响因HCC检测的困难而进一步加剧 肿瘤在临床上，导致大多数在临床上确定的肿瘤表现为既存 转移诊断时疾病的侵袭性导致5年生存率为15 百分之这项工作的长期目标是描绘纵向信号扰动， 导致NAFLD和随后进展为转移性HCC。该项目将：1）开发一个信令配置文件 2）描绘与HCC进展为转移表型相关的信号传导变化，和 3)研究转移性HCC中的空间定位信号传导。作为这项工作的核心组成部分， 直接的蛋白激酶活性传感器将用于炎症、生长、存活、细胞运动， 代谢途径这一组新型激酶活性传感器依赖于掺入一种 将磷酸化敏感的氨基酸转化为激酶选择性底物，允许基于直接荧光的 复杂生物样品中激酶酶活性的读出。为配合这一举措，该项目 将利用多学科方法将观察到的激酶信号传导的变化与 RNA和代谢产物。这种多管齐下的方法也将有助于确定监管机构 作为在疾病状态中观察到的改变的激酶活性的效应物。综合而言，拟议项目将 提供NAFLD中信号扰动的定量图像，建立激酶之间的相关性， 活性和侵袭性表型，并能够分析HCC中的空间限制性激酶活性 细胞系从长远来看，这项工作将为临床诊断的发展提供基础，并确定 在HCC的发展和进展中进行治疗干预的潜在新靶点。

ABSTRACT Rates of hepatocellular carcinoma (HCC) have been steadily increasing in the United States. One explanation for this observation is the alarming trend in obesity. A positive correlation between the development of non- alcoholic fatty liver disease (NAFLD) and HCC has been identified in the clinic, supporting a link between obesity and HCC. The public health impact of HCC is further exacerbated by the difficulty in detecting HCC tumors in the clinic, leading to the majority of tumors identified in the clinic presenting with preexisting metastasis. The aggressive nature of the disease at diagnosis results in a dismal five-year survival rate of 15 percent. The long-term goal of the proposed work is to delineate the longitudinal signaling perturbations that lead to NAFLD and subsequent progression to metastatic HCC. This project will: 1) develop a signaling profile of NAFLD, 2) delineate signaling changes associated with progression of HCC to a metastatic phenotype, and 3) investigate spatially localized signaling in metastatic HCC. As a central component of this work, a panel of direct protein kinase activity sensors will be employed for inflammatory, growth, survival, cell motility, and metabolic pathways. This panel of novel kinase activity sensors relies on the incorporation of a phosphorylation-sensitive amino acid into kinase-selective substrates, allowing for a direct fluorescence-based readout of kinase enzymatic activity in complex biological samples. To complement this initiative, this project will leverage a multidisciplinary approach to correlate observed changes in kinase signaling with changes in RNA and metabolite production. This multipronged approach will permit the identification of regulators, as well as effectors, of altered kinase activity observed in the disease state. Taken together, the proposed project will provide a quantitative picture of signaling perturbations in NAFLD, establish correlations between kinase activity and invasive phenotypes in HCC, and enable the analysis of spatially restricted kinase activity in HCC cell lines. In the long-term, this work will provide a basis for the development of clinical diagnostics and identify potential new targets for therapeutic intervention in the development and progression of HCC.