Systems microscopy analysis of tumor cell motility in microenvironment context

微环境背景下肿瘤细胞运动的系统显微镜分析

• 批准号: 8424468
• 负责人: Bojana Gligorijevic
• 金额: $ 14.04万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-07 至 2015-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8424468
• 关键词: Address; Advisory Committees; Affect; Algorithms; Anatomy; Animals; Area; Award; Back; Behavior; Biological; Biological Models; Biomechanics; Biophotonics; Blood Vessels; Cell Culture Techniques; Cell physiology; Cells; Cellular biology; Chemotactic Factors; Classification; Collaborations; Collagen; Computational Biology; Computer Simulation; Consult; Critiques; Custom; Cytoskeleton; Data; Detection; Diagnosis; Diagnostic; Early treatment; Educational process of instructing; Endothelial Cells; Environment; Extracellular Matrix; Faculty; Fellowship; Fibroblasts; Frequencies; Funding; Goals; Growth Factor; Image; Image Analysis; In Vitro; Individual; Institution; Intervention; Investigation; Knowledge; Laboratories; Lead; Legal patent; Life; Link; Literature; Locomotion; Machine Learning; Malignant Neoplasms; Mammary Neoplasms; Measurement; Measures; Mentors; Metalloproteases; Microscopy; Modeling; Modification; Molecular; Mus; Neoplasm Metastasis; Neoplasms in Vascular Tissue; Optics; Outcome; Pathway interactions; Peer Review; Phase; Play; Population; Postdoctoral Fellow; Primary Neoplasm; Publications; Reporter; Reporting; Research; Resources; Role; Serbia; Shapes; Signal Pathway; Signal Transduction; Staging; System; Systems Biology; Techniques; Technology; Testing; Theoretical Biology; Therapeutic Intervention; Time; Training; Tumor Biology; Universities; Work; Writing; abstracting; base; career development; cell behavior; cell motility; design; flexibility; human disease; improved; in vivo; inhibitor/antagonist; innovation; instructor; interest; intravital imaging; macrophage; malignant breast neoplasm; mathematical model; meetings; member; mouse model; multidisciplinary; neoplastic cell; novel; novel diagnostics; novel therapeutics; public health relevance; research study; statistics; time use; tumor; tumor microenvironment; tumor progression

DESCRIPTION (provided by applicant): Candidate: Majoring as analytical chemist during the time in Belgrade, Serbia, Dr. Gligorijevic has become an expert in optics and live cell microscopy over the course of her graduate training at Georgetown University and developed several novel microscopy-based technologies. During the postdoctoral period, Dr. Gligorijevic has focused on studying tumor cell intravasation in breast tumors, in living animals. To do so, she introduced several advancements into in vivo multiphoton microscopy. Dr. Gligorijevic is funded by DOD and Charles Revson Fellowship and has received several honors for her research. Her work to date resulted in one patent, ten peer-reviewed and ten other publications. Recent research has shown that different components of tumor microenvironment have an essential role in tumor progression. In order to decipher the network of interactions which shape the tumor environment and determine tumor cell behavior, it is ideal to apply systems biology approaches combined with in vivo microscopy, as cell cultures are limited in their complexity, mouse models are time-consuming and expensive and in silico models need optimization based on experiments. During the mentored phase of the Award, Dr. Gligorijevic will incorporate the relevant principles of advanced statistics and mathematical modeling of dynamical systems into her background of tumor biology and microscopy. She will take coursework offered at Einstein and meet regularly with her sponsor and co-sponsor. This training will make it possible to build a laboratory which investigates mechanisms of metastasis using the integrative, systems microscopy approach and utilizing information from molecular to population levels. Environment: Sponsor of the PI, Dr. John Condeelis is a Co-Chair of Anatomy Department and Biophotonic Center at Einstein. His lab and the Center create a multidisciplinary environment focused on answering mechanisms of human diseases, such as cancer, through use of microscopy. The Center is well known for its shared imaging resources and Innovation Laboratory where new microscopy systems are custom-built to accommodate specific needs of different projects. Consulting member of Advisory Committee, Dr. Robert Singer, is an expert in combining experimental and theoretical biology and a renowned mentor. The co-sponsor, Dr. Aviv Bergman, is the Founding Chair of Systems Department at Einstein and teaches coursework which will be a part of the career development. Einstein is an institution which highly values collaboration and insists on career development of postdoctoral fellows, instructors and junior faculty. Research: While most research of tumor microenvironments focuses on isolating and understanding single parameters, an integrative, systems-level network of interactions among relevant biological players is missing. In primary tumors, there are numerous biomechanical signals able to direct tumor cell movement towards and into the blood vessels. Growth factors, secreted by host macrophages, fibroblasts and endothelial cells in the tumor microenvironment are the main chemoattractants but recent studies show that the extracellular matrix also plays an important role. In loose extracellular matrix, tumor cells can migrate by reorganizing their cytoskeleton, generating a protrusive force. In regions with stiff extracellular matrix, simple locomotion is not possible. Here, tumor cells become invasive and degrade extracellular matrix, mainly by matrix metalloproteases (MMPs). Tumor cell migration was well characterized in vivo, but the mechanism of the switch from locomotory to invasive state and assembly of invasive protrusions in vivo are unknown. To address the link between integrated microenvironment signaling and the tumor cell behavior, it is necessary to combine cell biology, advanced microscopy and systems biology. In preliminary experiments, tumor cells were recorded using time-resolved 3D multiphoton imaging in living mice and two different protrusion types were observed: a) locomotory protrusions which quickly lead to migration of the tumor cell and b) invasive protrusions, which are persistent and MMP-dependent. The goal of Aim 1 is to investigate the role of invasive protrusions in the tumor cell intravasation and metastasis. A combination of fluorescent reporters will be used for imaging of invasive protrusion assembly and investigation of consequent tumor cell fate. Aim 2 explores signals which drive tumor cells to form either locomotory or invasive protrusions or to switch between behaviors. In areas which contain either locomotory or invasive protrusions, tumor microenvironment parameters are recorded simultaneously (number of macrophages, collagen stiffness, blood vessel size etc.). Imaging analysis results in a data matrix, which is analyzed by a Support Vector Machine classification. Classification shows that tumor cells switch from locomotory to invasive states under very specific conditions. Proposed experiments test the hypothesis that tumor cell behavior can be changed by slightly modifying microenvironment parameters. Relevant microenvironment parameters are incorporated into an integrative mathematical model of the tumor cell switch from locomotory to invasive state, in Aim 3. Using the predictive power of the model, experiments were designed to inhibit the invasive state and subsequent metastasis. Experimental outcomes will be used to optimize and complexify the model. Results of this study will lead to better understanding of the interplay among microenvironment components during tumor progression and the results will be used to improve diagnosis and treatment of early metastasis.

描述（由申请人提供）： 候选人：Gligorijevic 博士在塞尔维亚贝尔格莱德主修分析化学家，在乔治城大学研究生培训期间已成为光学和活细胞显微镜方面的专家，并开发了几种基于显微镜的新型技术。在博士后期间，Gligorijevic 博士专注于研究活体动物乳腺肿瘤中的肿瘤细胞内渗。为此，她引入了体内多光子显微镜的几项进步。 Gligorijevic 博士由国防部和查尔斯·雷夫森奖学金资助，并因其研究获得了多项荣誉。迄今为止，她的工作成果包括一项专利、十项同行评审以及十篇其他出版物。最近的研究表明，肿瘤微环境的不同组成部分在肿瘤进展中发挥着重要作用。为了破译塑造肿瘤环境并决定肿瘤细胞行为的相互作用网络，最好将系统生物学方法与体内显微镜相结合，因为细胞培养物的复杂性有限，小鼠模型既耗时又昂贵计算机模型需要基于实验进行优化。在该奖项的指导阶段，Gligorijevic 博士将把动态系统的高级统计和数学建模的相关原理融入到她的肿瘤生物学和显微镜学背景中。她将参加爱因斯坦提供的课程，并定期与她的赞助商和共同赞助商会面。该培训将使建立一个实验室使用综合系统显微镜方法并利用从分子到群体水平的信息来研究转移机制成为可能。 环境： PI 的发起人 John Condeelis 博士是爱因斯坦解剖学系和生物光子中心的联合主席。他的实验室和该中心创建了一个多学科环境，专注于通过使用显微镜来解答癌症等人类疾病的机制。该中心以其共享的成像资源和创新实验室而闻名，其中新的显微镜系统是定制的，以满足不同项目的特定需求。顾问委员会顾问罗伯特·辛格博士是实验与理论生物学相结合的专家，著名导师。共同发起人阿维夫·伯格曼 (Aviv Bergman) 博士是爱因斯坦系统系的创始主席，教授的课程将成为职业发展的一部分。爱因斯坦是一所高度重视合作并坚持博士后、导师和初级教师职业发展的机构。 研究：虽然大多数肿瘤微环境研究侧重于分离和理解单个参数，但缺少相关生物参与者之间的综合、系统级相互作用网络。在原发性肿瘤中，有许多生物力学信号能够引导肿瘤细胞向血管运动并进入血管。肿瘤微环境中宿主巨噬细胞、成纤维细胞和内皮细胞分泌的生长因子是主要的化学引诱剂，但最近的研究表明细胞外基质也发挥着重要作用。在疏松的细胞外基质中，肿瘤细胞可以通过重组其细胞骨架来迁移，产生突出力。在细胞外基质坚硬的区域，简单的运动是不可能的。在这里，肿瘤细胞变得具有侵袭性并主要通过基质金属蛋白酶（MMP）降解细胞外基质。肿瘤细胞迁移在体内得到了很好的表征，但从运动状态转变为侵袭状态以及体内侵袭性突起组装的机制尚不清楚。为了解决整合微环境信号传导与肿瘤细胞行为之间的联系，有必要将细胞生物学、先进显微镜和系统生物学结合起来。在初步实验中，使用时间分辨 3D 多光子成像在活体小鼠中记录肿瘤细胞，并观察到两种不同的突起类型：a）快速导致肿瘤细胞迁移的运动突起和 b）持久性和 MMP 的侵入性突起-依赖。目标 1 的目标是研究侵袭性突起在肿瘤细胞内浸润和转移中的作用。荧光报告基因的组合将用于侵入性突起组装的成像和随后的肿瘤细胞命运的研究。目标 2 探索驱动肿瘤细胞形成运动或侵入性突起或在行为之间切换的信号。在含有运动或侵入性突起的区域，同时记录肿瘤微环境参数（巨噬细胞数量、胶原硬度、血管尺寸等）。成像分析产生数据矩阵，并通过支持向量机分类进行分析。分类表明肿瘤细胞在非常特定的条件下从运动状态转变为侵袭状态。拟议的实验验证了肿瘤细胞行为可以通过稍微修改微环境参数来改变的假设。在目标 3 中，相关微环境参数被纳入肿瘤细胞从运动状态转变为侵袭状态的综合数学模型中。利用模型的预测能力，设计实验来抑制侵袭状态和随后的转移。实验结果将用于优化和复杂化模型。这项研究的结果将有助于更好地了解肿瘤进展过程中微环境成分之间的相互作用，并将用于改善早期转移的诊断和治疗。