A Microfluidic System for Image-Guided RNA-Seq of Single Cells in Glioblastoma

描述（由申请人提供）：候选人提出了一个为期五年的职业发展和研究计划下教授安德烈卡利法诺的指导。候选人的职业目标是在哥伦比亚大学建立一个独立的研究项目，专注于工程新工具，用于在生物医学中直接应用的单细胞系统范围内的询问。研究部分涉及微流体阵列平台的开发，该平台允许高度并行的光学成像和从单细胞中捕获RNA。重要的是，阵列元件是可单独寻址的，使得可以提取来自感兴趣的单个细胞的RNA文库用于RNA-Seq。该系统将允许候选人和他的合作者解决有关胶质母细胞瘤（一种常见且异常异质性的脑肿瘤）的肿瘤组成和进展的关键问题。虽然候选人有很强的技术导向背景，但他需要大量的培训才能执行提案的生物医学方面。因此，职业发展部分侧重于获得与以下相关的技能：1）组织处理，从组织中分散单细胞悬液，培养组织衍生细胞; 2）标记，FACS和组织衍生细胞的激光捕获; 3）RNA稳定/提取和生物信息学。此外，职业发展计划包括在这些领域的具体课程，以及指导赠款写作和科学交流和假设发展的生物医学研究。哥伦比亚在这些领域有完善的研究计划和大量的共享资源，这些资源将在提案的各个部分得到利用。该奖项将促进候选人作为独立生物医学研究人员的过渡，并形成有价值的合作。 公共卫生相关性：细胞异质性对胶质母细胞瘤的常规批量研究构成了严重挑战，胶质母细胞瘤是一种常见的脑肿瘤，特别难以治疗。通过定量成像实验中表现型特征的确切单个细胞的转录组，该项目将显着提高我们理解这些复杂系统的能力，分析临床样本，并最终设计更有效的治疗方法。

DESCRIPTION (provided by applicant): The Candidate proposes a five-year career development and research program under the mentorship of Prof. Andrea Califano. The Candidate's career goal is to build an independent research program at Columbia University focusing on engineering new tools for system-wide interrogation of single cells with direct applications in biomedicine. The research component involves the development of a microfluidic array platform that allows highly parallel optical imaging and RNA capture from single cells. Importantly, the array elements are individually addressable so that RNA libraries from individual cells-of-interest can be extracted for RNA-Seq. This system will allow the Candidate and his collaborators to address key questions about tumor composition and progression in glioblastoma, a common and unusually heterogeneous type of brain tumor. Although the Candidate has a strong technology-oriented background, he requires significant training in order to carry out the biomedical aspects of the proposal. Hence, the career development component focuses on acquiring skills related to 1) tissue handling, dispersing single cell suspensions from tissue, and culturing tissue-derived cells; 2) labeling, FACS, and laser capture of tissue-derived cells; and 3) RNA stabilization/extraction and bioinformatics. In addition, the career development plan includes specific coursework in these areas as well as mentoring in grant-writing and scientific communication and hypothesis development in biomedical research. Columbia has well-established research programs in these areas and substantial shared resources which will be leveraged in various parts of the proposal. This award will facilitate the candidate's transitio as an independent biomedical researcher and the formation of valuable collaborations. PUBLIC HEALTH RELEVANCE: Cellular heterogeneity poses a serious challenge to conventional bulk studies of glioblastoma, a common form of brain tumor that is particularly difficult to treat. By enabling quantification of transcriptomes from the exact individual cells phenotypically characterized in an imaging experiment, this project will significantly enhance our ability to understand these complex systems, analyze clinical samples, and ultimately design more effective treatments.