Collaborative Research: An Experimental/Theoretical Program on Reconfigured Polycationic Architectures for Improved Gene Therapy

这项由材料研究部生物材料项目授予马萨诸塞州大学和科罗拉多的合作研究奖项，旨在进行实验和模拟，以了解聚阳离子-DNA复合作用，以改善使用合成聚合物的非病毒基因治疗。该奖项由材料研究部的聚合物项目共同创立，旨在了解DNA与定制的大分子阳离子结构的复合，以及不同复合模式对DNA递送的影响。建议的实验集中在合成聚阳离子，其中阳离子低聚物接枝悬垂到聚合物主链在精确的接枝间距。该架构平台将通过将核定位序列以优选方向插入骨架中来增强。将在多种条件下在细胞培养物中测定多聚物稳定性和DNA递送效率，目的是将高细胞活力与优异的转染效率结合联合收割机。结合实验进行的计算方法将包括：1）原子模拟，以解释系统地改变聚阳离子的结构和化学特征对DNA-聚阳离子结合自由能的影响，并反过来连接到聚合物稳定性的实验趋势;以及2）集中于复合物结构的粗粒度模拟（相对大小、形状和电荷）作为聚阳离子结构和组成的函数，其直接涉及聚合物结构的实验，并且对血清稳定性和转染效率具有关键意义。除了改变对基于聚合物的基因治疗的理解和实践之外，该项目还将提供一个独特的机会，通过SkypeTM会议定期进行组间互动，并为研究人员提供时间和资源，以培训研究生进行综合多学科实验/理论项目。PI将继续致力于本科生参与本研究项目，利用科罗拉多大学和马萨诸塞州大学的本科生研究机会和研究经验。计划中的研讨会活动将连接美国和世界各地的领导者在实验和计算的聚电解质和络合，并将涉及年轻的科学家在他们的职业生涯的开始，通过在碲化物科学研究会议，一个高度互动的设置为研究演示和讨论的会议组织。 设计和实施有效的聚合物为基础的基因传递将推进基因组研究，并开辟新的途径，提供DNA治疗许多疾病，如肌营养不良症。迫切需要改进DNA递送，因为目前使用的基于病毒的递送方法具有其自身的健康和安全风险。在这个合作项目中，计算机模拟将帮助实验人员决定准备哪些聚合物用于复合DNA，以及如何修改这些聚合物以实现最有效的治疗作用。 该项目的成果包括开发新型和更有效的DNA递送剂，以及对带电聚合物及其与关键治疗生物分子（如DNA和RNA）相互作用的更透彻的基本理解。这个实验/理论项目的多学科性质将允许学生在马萨诸塞州和科罗拉多，在高中，本科和研究生水平，在分子模拟，聚合物合成和基因传递领域的全面培训。

This collaborative research award by the Biomaterials program in the Division of Materials Research, made to the University of Massachusetts and University of Colorado, is to carry out experiments and simulations to understand polycation-DNA complexation towards improved non-viral gene therapy using synthetic polymers. This award cofounded by the Polymer program in the Division of Materials Research seeks to understand DNA complexation with tailored macromolecular cationic architectures, and the impact of different modes of complexation on DNA delivery. The proposed experiments focus on synthetic polycations, in which cationic oligomers are grafted pendent to a polymer backbone at precise inter-graft spacing. This architecture platform will be augmented by insertion of nuclear localization sequences into the backbone in preferred orientations. Polyplex stability and DNA delivery efficiency will be determined in cell culture under a variety of conditions, aiming to combine high cell viability with excellent transfection efficiency. The computational approach, carried out in conjunction with experiments, will involve: 1) atomistic simulations to explain the effects of systematically varying architectural and chemical features of the polycation on free energy of DNA-polycation binding, and in turn connect to experimental trends in polyplex stability; and 2) coarse-grained simulations that focus on polyplex structure (relative size, shape and charge) as a function of polycation architecture and composition, which directly relate to experiments on polyplex structure, and carries key implications for serum stability and transfection efficiency. In addition to transforming the understanding and practice of polymer-based gene therapy, the project will open a unique opportunity to train graduate students in an integrated multidisciplinary experimental/theoretical project through regular inter-group interactions via SkypeTM sessions, and by devoting time and resources for the researchers to visit their collaborator's institution. The PIs will maintain a commitment to participation of undergraduates in this research project, utilizing the Undergraduate Research Opportunity and Research Experience for Undergraduates programs at University of Colorado and University of Massachusetts. Planned workshop activities would connect U.S. and worldwide leaders in both experiment and computation of polyelectrolytes and complexation, and would involve young scientists at the outset of their careers, through the organization of meetings at the Telluride Science Research Conference, a highly interactive setting for research presentations and discussions. The design and implementation of effective polymer based gene delivery will advance genomic research and open new avenues to deliver DNA for treating many diseases, such as muscular dystrophy. There is a pressing need for improved DNA delivery, since currently used delivery methods, based on viruses, have their own health and safety risks. In this collaborative project, computer simulations will help experimentalists decide which polymers to prepare for complexing DNA, and how to modify those polymers for most effective therapeutic action. Outcomes of this project include both the development of novel and more efficient DNA delivery agents, and a more thorough fundamental understanding of charged polymers and their interactions with key therapeutic biomolecules such as DNA and RNA. The multidisciplinary nature of this experimental/theoretical project will allow for a well-rounded training of students in Massachusetts and Colorado, at the high school, undergraduate, and graduate levels, in areas of molecular simulation, polymer synthesis, and gene delivery.