Anti-cancer therapeutic approaches for targeting the ALT telomere maintenance mechanism

项目摘要/摘要 在肿瘤发生过程中，癌细胞需要获得端粒维持机制才能无限增殖。 间叶性肿瘤(如肉瘤、内分泌瘤、胶质母细胞瘤和一些儿童早期肿瘤 癌症)，经常获得一种端粒酶不依赖的端粒维持机制，这一直是 称为端粒交替延长(ALT)。此外，已有研究表明，端粒酶抑制在 端粒酶阳性的人类癌细胞可以参与ALT途径。尽管ALT癌症可以被诊断出来 通过肿瘤切片的端粒荧光原位杂交(FISH)和C-环实验(phi29 肿瘤DNA的聚合酶反应，目前还没有治疗ALT肿瘤的靶点或临床试验 治疗。因此，确定ALT肿瘤的潜在治疗靶点可能会显著改善 通过识别和利用独特的ALT癌症脆弱性，为ALT癌症患者提供选择。最终， 这将导致基于ALT状态的个性化癌症治疗。从我最近的研究中，我发现 ALT的分子机制是由PML小体中端粒的过度和持续聚集启动的。我 我一直在进一步研究这些过程是由一系列翻译后修饰所调节的 和分离：通过相扑靶向泛素化E3连接酶(RNF4)泛素化，以及通过泛素- 选择性伴侣/AAA-ATPase(p97；也称为VCP)。在目标1中，我将进一步剖析 RNF4和p97在ALT途径中的表达，并验证抑制RNF4-p97轴的治疗机会窗 作为一种有效的抗ALT癌症策略。ALT癌症的特征之一是大量聚集 早幼粒细胞白血病(PML)小体中的端粒。我已经开发出一种生物物理系统，它可以重组 PML小体由最少的成分组成，并产生端粒聚集，从而模仿ALT相关的PML 活体(APB)。在目标2中，我将进一步设计模拟ALT癌症表型的生物物理系统 并将这种ALT模型应用于蛋白质组学和候选基因筛选，以确定新的治疗靶点 另一种癌症。我的主要职业目标是在R1大学或研究部获得一个助理教授的职位 在美国的研究所。我的长期职业目标是成为一名独立调查员和领导者 癌细胞生物学领域。K22奖项下的培训将作为我未来方向的基础 研究，以及由此产生的出版物将支持我向独立的过渡，并为我准备 提交额外的拨款申请，包括R01。我从目标1和目标2获得的结果将使我 进一步研究ALT途径的分子机制，寻找新的分子靶点和新的分子靶点 目前还没有针对ALT癌症的治疗方法。

Project Summary/Abstract During tumorigenesis, cancer cells need to acquire a telomere maintenance mechanism to proliferate indefinitely. Cancers of mesenchymal origin (such as sarcomas, endocrine tumors, glioblastoma, and some early childhood cancers), frequently acquire a telomerase-independent telomere maintenance mechanism, which has been termed Alternative Lengthening of Telomeres (ALT). Moreover, it has been shown that telomerase inhibition in telomerase positive human cancer cells can engage the ALT pathway. Although ALT cancers can be diagnosed by telomere fluorescence in situ hybridization (FISH) of tumor sections, and by using the C-circle assay (phi29 polymerase reaction) of tumor DNA, currently there are no therapeutic targets or clinical trials for ALT tumor treatment. Therefore, identification of potential therapeutic targets for ALT tumors may significantly improve the options for patients with ALT cancers by identifying and exploitering unique ALT cancer vulnerabilities. Ultimately, this should lead to personalized cancer treatment based on ALT status. From my recent studies, I found that the molecular mechanism of ALT is initiated by excessive and persistent clustering of telomeres in PML bodies. I have been further investigating that these processes are mediated by a series of post-translational modifications and segregation: ubiquitination by SUMO-targeted ubiquitination E3 ligase (RNF4), and segregation by ubiquitin- selective chaperone/AAA ATPase (p97; also known as VCP). In aim 1, I will further dissect the mechanism of RNF4 and p97 in ALT pathway and validate the therapeutic window of opportunity to inhibit the RNF4-p97 axis as an effective anti-ALT cancer strategy. One of the hallmarks of ALT cancers is the clustering of large amounts of telomeres in promyelocytic leukemia (PML) bodies. I have developed a biophysical system that reconstitutes PML bodies from minimal components and generates telomere clustering, thus mimicking ALT-associated PML bodies (APBs) in vivo. In aim 2, I will further engineer the biophysical system mimicking ALT cancer phenotype and apply this ALT model to proteomics and candidate gene screenings to identify novel therapeutic targets for ALT cancer. My primary career goal is to obtain an assistant professor position at a R1 university or research institute in the United States. My long-term career goal is to become an independent investigator and leader in the field of cancer cell biology. Training under a K22 award will serve as the basis for the future direction of my research, and the resulting publications will support my transition toward independence and prepare me to submit additional grant applications including R01s. The results that I obtain from Aim 1 and Aim 2 will allow me to further investigate the molecular mechanisms of the ALT pathway to identify new molecular targets and novel therapies for ALT cancers that are not currently available.