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计划颁发给马萨诸塞州大学和科罗拉多大学的合作研究奖，是为了进行实验和模拟，以了解聚阳离子-DNA络合作用，从而使用合成聚合物改进非病毒基因治疗。该奖项由材料研究部的聚合物项目共同设立，旨在了解DNA与定制的大分子阳离子结构的络合作用，以及不同络合模式对DNA传递的影响。拟议的实验集中在合成聚阳离子上，其中阳离子低聚物以精确的接枝间距接枝到聚合物主干上。这一架构平台将通过在首选方向上将核定位序列插入主干来增强。在细胞培养中，将在各种条件下测定复合体的稳定性和DNA的输送效率，旨在结合高细胞存活率和优异的转染率。结合实验开展的计算方法将涉及：1)原子模拟，解释系统地改变聚阳离子的结构和化学特征对DNA-多聚阳离子结合自由能的影响，进而与多链稳定性的实验趋势相联系；2)粗粒度模拟，侧重于多链结构(相对大小、形状和电荷)作为多阳离子结构和组成的函数的多链结构的模拟，这与多链结构的实验直接相关，并对血清稳定性和转染率具有关键影响。除了改变对基于聚合物的基因治疗的理解和实践之外，该项目还将通过SkypeTM会议的定期小组间互动，以及通过为研究人员访问他们的合作者的机构提供时间和资源，为研究生提供一个独特的机会，在一个综合的多学科实验/理论项目中培训研究生。PIS将利用科罗拉多大学和马萨诸塞大学为本科生提供的本科生研究机会和研究经验，继续致力于本科生参与这一研究项目。计划中的研讨会活动将把美国和世界各地的聚合电解质和络合实验和计算领域的领先者联系起来，并将通过在Telluride科学研究会议上组织会议，让年轻科学家在职业生涯之初参与进来，这是一个高度互动的研究报告和讨论场所。基于聚合物的有效基因递送的设计和实现将促进基因组研究，并开辟新的途径来递送DNA用于治疗许多疾病，如肌肉营养不良。目前迫切需要改进DNA传递，因为目前使用的基于病毒的传递方法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者决定准备哪些聚合物来络合DNA，以及如何修饰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高效的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)的相互作用有更彻底的基本了解。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学生在分子模拟、聚合物合成和基因传递领域对高中、本科生和研究生进行全面的培训。

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.