Characterizing the Role of Leucyl Aminoacyl tRNA Synthetase (LARS) in Breast Cancer Metastasis

项目摘要/摘要 乳腺癌转移是美国女性死亡的主要原因。尽管在治疗方面取得了进展 对于早期肿瘤患者，转移性疾病患者的预后仍然很差。之前的工作已经完成 证实转移RNA(TRNAs)作为乳腺癌转移进展的直接驱动因素，通过 不同的丰度导致细胞蛋白质组格局的变化。我假设氨基酰化状态， 例如，tRNA是否带有氨基酸，以及必需的氨基酰基tRNA合成酶(AARS) 负责tRNA充电，也会在乳腺癌转移进展中发挥作用。为了测试这一点 假设，带电的tRNA在不同转移潜力的细胞中被描绘出来，这识别了关键 转移潜能较高的细胞中亮氨酸tRNA电荷的减少。异种和同基因移植的初步研究 小鼠转移研究进一步证实负责充电亮氨酸tRNAs的亮氨酰Aars(LARs)是一种 转移抑制因子。在这项提案中，我将研究并进一步描述LARS在癌症中的作用 转移。在目标1中，我将进一步描述LARS操纵的影响(1)通过击倒和 在异种移植和同基因肿瘤小鼠模型中的过度表达，以及(2)通过基因敲除在小鼠中的表达 乳腺癌的模型。在目标2中，我将研究(1)通过以下途径抑制转移的细胞表型 体外检测和(2)荷电tRNA介导的下游转移抑制机制 RNA测序分析和核糖体图谱。这些拟议的研究将填补文学上的空白： TRNA在癌症进展中发挥作用，与其生物发生相关的分子尚未被确定为 在癌症转移中具有治疗意义的靶点。这些目标的成功实现将为 为未来以新机制治疗转移性乳腺癌的治疗靶点奠定基础。 我是威尔·康奈尔/洛克菲勒/斯隆·凯特林三所学院的医学博士研究生 计划，在洛克菲勒大学的Sohail Tavazoie博士的实验室完成了拟议的目标。塔瓦佐伊医生 与我的委员会成员查尔斯·赖斯博士、平奇博士和基万茨·伯索博士一起，提供了强有力的科学依据 确保项目成功完成的专门知识、指导和资源。共同努力，我们已经取得了 确保我的培训计划也允许发展必要的口头和书面沟通的步骤 技能和指导机会。我的临床导师帕梅拉·查尼博士和池平博士将提供 在我向完成医学培训过渡的时候，关于保持临床敏锐度的关键建议 研究-跟踪内科住院医师计划。成功完成本年度的拟议计划 一流的培训环境将增强我对RNA生物学的基础知识，并教会我 癌症生物学和老鼠模型，让我坚定地走上了成为一名内科科学家和 学术医疗中心的独立调查员。

PROJECT SUMMARY/ABSTRACT Breast cancer metastasis is a leading cause of mortality in US women. Despite advances in treatment for patients with early stage tumors, prognosis remains poor for patients with metastatic disease. Prior work has demonstrated a direct role for transfer RNAs (tRNAs) as drivers of breast cancer metastatic progression, through differential abundance resulting in changes in cell proteomic landscape. I hypothesize that aminoacylation status, as in, whether or not a tRNA is charged with an amino acid, and the requisite aminoacyl tRNA synthetase (aaRS) responsible for tRNA charging, will also play a role in breast cancer metastatic progression. To test this hypothesis, charged tRNAs were profiled across cells of differing metastatic potential, which identified key reductions in leucine tRNA charging in cells of higher metastatic potential. Preliminary xenograft and syngeneic mouse metastasis studies further identified leucyl aaRS (LARS), responsible for charging leucine tRNAs, as a metastasis suppressor. In this proposal, I will examine and further characterize the role of LARS in cancer metastasis. In Aim 1, I will further characterize the effects of LARS manipulation (1) through knockdown and overexpression in xenograft and syngeneic cancer mouse models, and (2) through genetic knockout in mouse models of breast cancer. In Aim 2, I will investigate (1) the cellular phenotype of metastasis suppression through in vitro assays and (2) the mechanism of charged tRNA-mediated metastasis suppression through downstream RNA sequencing analysis and ribosomal profiling. These proposed studies will fill a gap in literature: though tRNAs play a role in cancer progression, molecules related to their biogenesis have yet to be identified as therapeutically meaningful targets in cancer metastasis. Successful completion of these aims will lay the groundwork for future therapeutic targets with novel mechanisms, in treating metastatic breast cancer. I am an MD-PhD student at the Weill Cornell/Rockefeller/Sloan Kettering Tri-Institutional MD-PhD program, completing the proposed aims in the lab of Dr. Sohail Tavazoie at Rockefeller University. Dr. Tavazoie, along with my committee members Drs. Charles Rice, Ping Chi and Kivanc Birsoy, provide strong scientific expertise, guidance and resources to ensure successful completion of the project. Together, we have taken steps to ensure my training plan also allows for development of necessary verbal and written communication skills as well as mentorship opportunities. My clinical mentors, Drs. Pamela Charney and Ping Chi, will offer critical advice on maintaining clinical acumen as I transition towards completion of medical training and select research-track residency programs in Internal Medicine. Successful completion of the proposed plan in this stellar training environment will enhance my foundational knowledge of RNA biology and teach me skills in cancer biology and mouse modeling, setting me solidly on the path towards becoming a physician scientist and independent investigator at an academic medical center.