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

该合作研究奖由麻省大学和科罗拉多大学材料研究部生物材料项目颁发，旨在开展实验和模拟，以了解聚合阳离子- dna络合作用，从而利用合成聚合物改进非病毒基因治疗。该奖项由材料研究部聚合物项目共同创立，旨在通过量身定制的大分子阳离子结构了解DNA络合，以及不同模式的络合对DNA递送的影响。提出的实验重点是合成阳离子，其中阳离子低聚物以精确的接枝间距接枝到聚合物主链上。该结构平台将通过将核定位序列以首选方向插入到主干中来增强。在多种条件下的细胞培养过程中，我们将测定Polyplex的稳定性和DNA传递效率，旨在将高的细胞活力与优异的转染效率结合起来。计算方法与实验相结合，将涉及：1)原子模拟，以解释系统变化的多聚离子的结构和化学特征对dna -多聚离子结合自由能的影响，并反过来与多聚离子稳定性的实验趋势联系起来；2)粗粒度模拟，重点关注多聚体结构（相对大小、形状和电荷）作为多聚体结构和组成的函数，这直接关系到多聚体结构的实验，并对血清稳定性和转染效率具有关键意义。除了改变基于聚合物的基因治疗的理解和实践外，该项目还将通过SkypeTM会议定期进行小组间互动，并为研究人员投入时间和资源访问合作机构，为培养研究生提供一个综合多学科实验/理论项目的独特机会。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大学本科生项目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在聚电解质和络合的实验和计算方面的领导者，并将通过在特柳赖德科学研究会议（一个高度互动的研究报告和讨论场所）组织会议，让年轻的科学家参与到他们的职业生涯的开始。设计和实施有效的基于聚合物的基因传递将推进基因组研究，并为传递DNA治疗许多疾病（如肌肉萎缩症）开辟新的途径。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本身就存在健康和安全风险。在这个合作项目中，计算机模拟将帮助实验学家决定准备哪种聚合物来络合DNA，以及如何修改这些聚合物以获得最有效的治疗作用。该项目的成果包括开发新的和更有效的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）的相互作用的更彻底的基本理解。这个实验/理论项目的多学科性质将允许马萨诸塞州和科罗拉多州的高中生、本科生和研究生在分子模拟、聚合物合成和基因传递领域进行全面的培训。

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.