Regulation of non-proteolytic ubiquitination in cancer chemoresistance and progression

项目摘要 三阴性乳腺癌(TNBC)是一组发病率较高的侵袭性疾病。 复发、转移和患者死亡率与其他乳腺癌亚型的比较。遗传图谱研究 已确定至少有六个TNBC亚群显示出不同的分子特征。这些亚组- 相关的分子特征，或“生物标记物”，最终可能被用来预测治疗反应； 然而，并不是所有的生物标志物都会导致肿瘤进展和/或治疗耐药。因此，至关重要的是 更好地理解构成TNBC亚群的分子驱动因素。 TNBC表达丰富的Twist，是TNBC进展的基本和多功能调节因子。尽管 作为一个长期的药物靶点，目前还没有专门针对Twist活动的方法。泛素是一种 重要的监管信号。K48相关泛素化导致蛋白质降解和下调 底物，而新表征的K63连接泛素化激活蛋白质介导的信号转导 小路。E3连接酶是给予泛素信号特异性的底物识别蛋白。许多 E3连接酶在人类癌症中上调。几种使致癌E3连接酶失活的抑制剂是 目前正在进行临床试验。通过确定K63连接泛素化的原因是哪些E3连接酶 在Twist的基础上，我们可能会发现专门阻断Twist治疗TNBC的新药靶点。 该应用程序的目标是表征K63连接的泛素化网络，该网络协调 Twist的激活和蛋白表达及与Twist K63连锁的E3连接酶的鉴定 泛素化。我们将进行一系列的生化和生物学研究，以揭示这些机制 其中与K63相关的泛素化在翻译后调节Twist的活动。我们将建立病理性的 Twist K63连接泛素化及其控制的E3连接酶在动物研究中的作用。我们将验证 E3连接酶作为TNBC肿瘤功能生物标记物的生理学相关性及抑制作用 这些E3连接酶具有治疗潜力。与公共卫生的相关性：我们对E3连接酶特性的研究 作为TNBC的分子驱动因素，将为开发有效治疗TNBC的新战略打开大门。 通过无偏E3连接酶筛选，我们确定了两个主要的扭曲控制E3连接酶，它们似乎 以一种相互排斥的方式存在于大多数TNBC组织中，很可能是肿瘤的结果 异质性。如果这两个E3连接酶单独或组合确实影响TNBC的调控，那么它们可能 可作为个性化医疗中的功能性预后标记物。除了TNBC，获得的知识 这项工作可能适用于其他与Twist激活有关的肿瘤类型，如 前列腺癌、肺癌、头颈癌。

Project Summary Triple-negative breast cancer (TNBC) is a group of aggressive diseases with a greater incidence of relapse, metastasis and patient mortality compared to other breast cancer subtypes. Genetic profiling studies have identified at least six subgroups of TNBC exhibiting distinct molecular features. These subgroup- associated molecular features, or ‘biomarkers’, may eventually be used to predict response to treatment; however, not all biomarkers cause tumor progression and/or treatment resistance. It is therefore critical to better understand the molecular drivers that underlie TNBC subgroups. TNBC expresses abundant Twist, a fundamental and versatile regulator of TNBC progression. Despite being a long-standing drug target, there is no approach to specifically target Twist activity. Ubiquitin is an important regulatory signal. K48-linked ubiquitination causes degradation and downregulation of protein substrates, whereas the newly characterized K63-linked ubiquitination activates protein-mediated signaling pathways. E3 ligases are the substrate recognition protein that impart specificity to the ubiquitin signal. Many E3 ligases are upregulated in human cancers. Several inhibitors that inactivate the oncogenic E3 ligases are currently being tested in clinical trials. By identifying which E3 ligases account for the K63-linked ubiquitination of Twist, we could potentially discover new drug targets that specifically block Twist to treat TNBC. The goal of this application is to characterize the K63-linked ubiquitination network that orchestrates Twist activation and protein expression and to identify the E3 ligases responsible for Twist K63-linked ubiquitination. We will conduct a series of biochemical and biological studies to uncover the mechanisms by which K63-linked ubiquitination regulates Twist activity post-translationally. We will establish the pathological roles of Twist K63-linked ubiquitination and its controlling E3 ligases in animal studies. We will validate the physiological relevance of E3 ligases as functional biomarkers in TNBC tumors and assess whether inhibiting these E3 ligases has therapeutic potential. Relevance to Public Health: Our study characterizing E3 ligases as molecular drivers of TNBC will open the door to developing new strategies for effectively treating TNBC. Through a non-biased E3 ligase screen, we identified two primary Twist-controlling E3 ligases that appear to be present in most TNBC tissues in a mutually exclusive fashion, most likely as the result of tumor heterogeneity. If the two E3 ligases, individually or in combination, do affect TNBC regulation, then they could be used as functional prognosis markers in personalized medicine. Aside from TNBC, the knowledge gained through this work is potentially applicable to other tumor types where Twist activation is implicated, such as prostrate, lung, head and neck cancers.