IDR: Nucleic Acid-Lipid Films - Programmable Structural Transitions for Drug Delivery and Regulating Gene Expression

IDR：核酸-脂质膜 - 用于药物输送和调节基因表达的可编程结构转变

• 批准号: 1015026
• 负责人: Matthew Tirrell
• 金额: $ 60万
• 依托单位: University of California-Berkeley
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-15 至 2015-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1015026&HistoricalAwards=false
• 关键词: IDR; Nucleic; Acid; Lipid; Films

1015026TirrellIntellectual Merit: This interdisciplinary program aims to bring new developments in self-assembled materials to bear on frontier problems in bioactive nucleic acid (NA) delivery and gene expression. Materials comprising nucleic acids and lipids, assembled based on a balance of electrostatic, hydrophobic and hydration forces, form stable, layered films, alternating nucleic acid and lipid layers. Recently published work from our group has shown that this structure can be manipulated in various ways, for example, by changing the temperature or state of hydration, or by varying the molecular weights of the nucleic acids included. This provides a versatile platform of potentially enabling technology to advance and develop new capabilities in gene delivery and the regulation of gene expression. The first aim of the proposed work is to optimize these constructs for the stated delivery applications. Preliminary work included in the proposal demonstrates that nucleic acid delivery, leading to transfection of stem cells, is possible with these constructs. Maximization of this capability will be explored by varying lipid choices (to have the best possible disassembly characteristics and to minimize any toxicity), by including multiple nucleic acids to have the desired structural features, and more importantly, to be able to have simultaneous, multiple transfection ability. The structures and disassembly profiles of all of these constructs will be thoroughly characterized. A second aim of this work, which will be conducted in parallel with the first, is to use these constructs to examine transfection efficiency for mouse embryonic stem cells in culture, using expression of green fluorescent protein as an indicator of efficiency. Stem cells are notably difficult to transfect; our aim is to exploit the high concentration of nucleic acids in our constructs, and the direct physical contact of the cells with the nucleic acid-lipid layers to increase transfection efficiency. A third aim of the work will be to apply our new material delivery vehicles to the delivery of microRNAs. MicroRNAs (miRNAs) are a novel class of small, regulatory non-coding RNA, serving as potent regulators for gene expression at posttranscriptional level. Contact-mediated delivery will be explored to accomplish this goal. The further goal of the third phase of this work is to integrate plasmid DNA and miRNA delivery to reprogram adult cells into induced, pluripotent stem cells. iPS cells have numerous profound scientific and biomedical implications in personalized therapies and platforms for high-throughput screening of pharmaceuticals. The technical challenge with microRNA delivery for reprogramming is not delivery efficiency, per se, but rather sustained delivery to achieve sustained expression over a span of approximately ten days. Interdisciplinary Nature of the Proposed Research: The team assembled spans several different disciplines from chemical engineering and materials science, to stem cell and tissue engineering, to the molecular and cellular biology of gene regulation. The proposed work will take a discovery in materials science quite far toward new enabling technology in engineering nucleic acid delivery and gene expression. The connection this team embodies, among chemical and materials engineering, biology and biological engineering, is essential to realize the potential of this new delivery system, both to have the insight into how to optimize the materials involved, and to assure that the biological engineering objectives are achieved in a meaningful way. Broader Impacts: An exciting, interdisciplinary development project such as this is an ideal opportunity to bring undergraduate engineering students to the forefront of an important research area. The very nature of this project, spanning three laboratories in chemical, materials and biological engineering, as well as molecular and cellular biology, gives capacity to bring undergraduates with varied interests into participation in this work. The specific plan is to engage two undergraduate students per year, in the summer (six total over the project lifetime), from underrepresented groups as participants in this research. These students will be admitted to the Amgen Scholars Summer Research Program at UC Berkeley. The Amgen Scholars Program is a national program attracting approximately 25 participants each year. Joining this group of 25, the two undergraduate participants will benefit significantly in numerous ways as members of the summer research cohort. They will participate in all program activities including weekly meetings and the poster session and oral presentations at the end of the summer. As a result of these collaborative activities, the undergraduate participants in this project will be fully involved in a broad and comprehensive summer experience.

1015026 Tirrell智力优点：这个跨学科的计划旨在带来自组装材料的新发展，承担在生物活性核酸（NA）交付和基因表达的前沿问题。基于静电力、疏水力和水合力的平衡组装的包含核酸和脂质的材料形成稳定的分层膜，交替的核酸和脂质层。我们小组最近发表的工作表明，这种结构可以以各种方式操纵，例如，通过改变温度或水合状态，或通过改变所包含核酸的分子量。这提供了一个通用的平台，潜在地使技术能够推进和开发基因递送和基因表达调控的新能力。所提出的工作的第一个目的是优化这些结构所述的交付应用程序。该提案中包括的初步工作表明，核酸递送，导致干细胞转染，是可能的与这些构建体。将通过改变脂质选择（以具有最佳可能的分解特征并使任何毒性最小化），通过包括多个核酸以具有所需的结构特征，并且更重要的是，能够具有同时的多重转染能力，来探索这种能力的最大化。所有这些结构的结构和拆卸轮廓将被彻底表征。这项工作的第二个目的，这将与第一个平行进行，是使用这些构建体，以检查转染效率的小鼠胚胎干细胞培养，使用绿色荧光蛋白的表达作为效率的指标。干细胞特别难以转染;我们的目的是利用我们构建体中的高浓度核酸，以及细胞与核酸-脂质层的直接物理接触来提高转染效率。这项工作的第三个目标是将我们的新材料递送载体应用于microRNA的递送。microRNA（miRNAs）是一类新型的小分子非编码RNA，在转录后水平上对基因表达起着重要的调控作用。接触介导的交付将探讨实现这一目标。这项工作的第三阶段的进一步目标是整合质粒DNA和miRNA递送，以将成体细胞重编程为诱导的多能干细胞。iPS细胞在个性化治疗和药物高通量筛选平台方面具有许多深刻的科学和生物医学意义。用于重编程的microRNA递送的技术挑战不是递送效率本身，而是持续递送以在大约十天的跨度内实现持续表达。拟议研究的跨学科性质：该团队聚集了几个不同的学科，从化学工程和材料科学，干细胞和组织工程，到基因调控的分子和细胞生物学。拟议的工作将在材料科学中发现相当远的新技术，在工程核酸交付和基因表达。该团队体现了化学和材料工程，生物学和生物工程之间的联系，对于实现这种新输送系统的潜力至关重要，既要了解如何优化所涉及的材料，又要确保以有意义的方式实现生物工程目标。更广泛的影响：一个令人兴奋的，跨学科的发展项目，如这是一个理想的机会，使本科工程专业的学生到一个重要的研究领域的最前沿。这个项目的性质，跨越化学，材料和生物工程，以及分子和细胞生物学三个实验室，使能力，使不同兴趣的本科生参与这项工作。具体计划是每年夏季从代表性不足的群体中招募两名本科生（整个项目周期共六名）作为本研究的参与者。这些学生将被加州大学伯克利分校的安进学者夏季研究计划录取。安进学者计划是一项国家计划，每年吸引约25名参与者。加入这个25人的小组，两名本科生参与者将在许多方面作为夏季研究队列的成员受益匪浅。他们将参加所有的项目活动，包括每周会议和海报会议，并在夏季结束时进行口头报告。作为这些合作活动的结果，在这个项目的本科生参与者将充分参与一个广泛而全面的夏季经验。