CAREER: Biomolecular Engineering Approach to Controlling Cell Processes

职业：控制细胞过程的生物分子工程方法

• 批准号: 1453098
• 负责人: Josephine Allen
• 金额: $ 50.17万
• 依托单位: University of Florida
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-02-01 至 2021-09-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1453098&HistoricalAwards=false
• 关键词: CAREER; Biomolecular; Engineering; Approach; Controlling

PI: Allen, JosephineProposal Number: 1453098Developing strategies to control cellular functions is critical in the field of tissue engineering. This is often achieved through the activation of receptor proteins on the surface of the cells, which triggers a cascade of events leading to a desired cellular response. Of interest are the cellular responses that contribute to the formation of a blood vessels network. Controlling blood vessel formation is paramount to the success of many engineered tissue substitutes, and continues to be a challenge and a barrier to progress within the field. Current strategies include the use of receptor specific growth factors, which are difficult to control, can be unstable, require high doses, and may bind to unintentional targets. There is a need to promote blood vessel formation using alternative strategies. In this proposal, the PI seeks to develop a new strategy to control cellular processes through the use of highly tunable small molecules termed, aptamers. If successful, the proposed studies will advance our understanding of the fundamental science involved in controlling cellular behavior, and have a significant impact on the field of tissue engineering. Education and outreach activities have been integrated with the proposed research and involve providing hands-on research experience to students in local middle and high school, the creation of online educator resources which will be available to K-12 teachers nationwide, as well as outreach to underrepresented minority undergraduate students.Growth factors are used extensively in the field of tissue engineering to bind and activate cell surface receptors, with the ultimate goal to control receptor mediated process. This is particularly true for cell processes involved in angiogenesis, with the most successful approach involving the use of vascular endothelial growth factor. However, this strategy is limited due to instability of growth factors, the high doses required for efficacy, off target effects, and the lack of control of vessel development at distant sites. To address these shortcomings the investigator proposes the use of a highly tunable aptamer based platform as a strategy to engineer tissue constructs that can promote and support the formation of a vascular network. The challenge is to engineer aptamer structures to create a supramolecular aptamer assembly (SAA) that can be used to not only bind but activate target receptors and thereby control receptor mediated processes. The objective is to reveal the structural constraints that govern SAA-receptor binding and agonist behavior. Towards this goal the scientific aims set forth by this proposal are 1) characterization of supramolecular aptamer assembly (SAA) structures and target receptor interactions; and 2) correlate supramolecular aptamer assembly (SAA) structure with overall function to induce and promote angiogenesis. To successfully complete these aims, techniques established in the PI's lab will be employed for aptamer synthesis, receptor screening, and molecular evaluations of receptor interactions, including protein and gene expression changes. Structural biology and biophysical techniques will be employed to assess the structural constraints involved in aptamer-receptor interactions. Lastly, a three-dimensional model of angiogenesis will be used to quantitatively assess SAA induced angiogenesis. Completion of the tasks set forth will lay the foundation for a targeted and robust aptamer mediated neovascularization approach.

主要研究者：艾伦，约瑟芬提案号：1453098开发控制细胞功能的策略在组织工程领域至关重要。这通常通过激活细胞表面上的受体蛋白来实现，这触发了导致所需细胞应答的级联事件。感兴趣的是有助于血管网络形成的细胞反应。控制血管形成对于许多工程组织替代物的成功至关重要，并且仍然是该领域内进展的挑战和障碍。目前的策略包括使用受体特异性生长因子，其难以控制，可能不稳定，需要高剂量，并且可能与无意的靶结合。需要使用替代策略来促进血管形成。在这项提案中，PI试图开发一种新的策略，通过使用高度可调的小分子（称为适体）来控制细胞过程。如果成功，这些研究将促进我们对控制细胞行为的基础科学的理解，并对组织工程领域产生重大影响。教育和推广活动已与拟议的研究相结合，并涉及为当地初中和高中的学生提供实践研究经验，创建在线教育资源，供全国K-12教师使用，以及推广到代表性不足的少数民族本科生。生长因子广泛用于组织工程领域，以结合和激活细胞表面受体，最终目的是控制受体介导的过程。对于参与血管生成的细胞过程尤其如此，最成功的方法涉及使用血管内皮生长因子。然而，由于生长因子的不稳定性、有效性所需的高剂量、脱靶效应以及缺乏对远端部位血管发育的控制，该策略受到限制。为了解决这些缺点，研究者提出使用高度可调的基于适体的平台作为一种策略来工程化组织构建体，其可以促进和支持血管网络的形成。挑战在于设计适体结构以产生超分子适体组装体（SAA），其不仅可用于结合而且可用于激活靶受体，从而控制受体介导的过程。目的是揭示SAA受体结合和激动剂行为的结构约束。为了实现这一目标，本提案提出的科学目的是1）表征超分子适体组装（SAA）结构和靶受体相互作用;以及2）将超分子适体组装（SAA）结构与诱导和促进血管生成的总体功能相关联。为了成功完成这些目标，PI实验室建立的技术将用于适体合成，受体筛选和受体相互作用的分子评价，包括蛋白质和基因表达变化。将采用结构生物学和生物物理技术来评估适体-受体相互作用中涉及的结构约束。最后，血管生成的三维模型将用于定量评估SAA诱导的血管生成。所提出的任务的完成将为靶向和稳健的适体介导的新血管形成方法奠定基础。