Structure-property study for dendronized polymer vectors for CRISPR delivery

非技术总结：该项目开发了一种新的分子递送系统，用于实现基于CRISPR的基因组编辑技术。 基于CRISPR的基因组编辑技术可以在体外或体内精确地添加、删除和“校正”基因;因此，它在治疗应用方面具有巨大的潜力，并且正在积极地用于解决从遗传性疾病、癌症到糖尿病的各种人类疾病。然而，实施这项技术的一个主要障碍是缺乏安全有效的递送系统，可以有效地将所需的分子机器运送到细胞中以执行基因组编辑。为了应对这一挑战，本项目提出了一系列由天然氨基酸和肽制成的递送分子的设计，合成和研究。 将评估这些化合物将两段RNA递送到细胞中进行基于CRISPR的基因组编辑的效率和安全性。 结构-功能研究将揭示有效分子载体的重要设计原则。 本项目的目标是开发一种新的基于RNA的CRISPR合成载体平台，并对这种新的载体系统进行基本的结构-性质研究，为设计新的基因载体提供关键性的见解。技术概要：本项目的目标是开发一种新的基于RNA的CRISPR合成载体平台，并对这种新的载体系统进行结构-性质研究，为设计新的基因载体提供重要的见解。CRISPR/Cas9基因编辑为治疗应用提供了巨大的潜力，并正在积极寻求解决各种人类疾病。然而，实施CRISPR介导的基因组编辑的一个主要障碍是缺乏安全有效的递送载体。PI实验室最近开发了一种生物可还原的树枝化多肽（bdp）的创新设计，其显示出高效率的Cas9 mRNA和gRNA的共递送。对该系统的进一步研究将产生一种新的平台递送系统，该系统可以有效且安全地将CRISPR/Cas9 mRNA和gRNA共递送到各种细胞中。此外，利用BDP的明确定义的分子结构，我们建议进行基本的结构性质的研究，BDP系统的目的是获得关键的见解，设计新的基因传递载体。如果成功，这项研究将为CRISPR技术提供一种新的、通用的、生物相容性的递送系统，并为指导新基因递送系统的设计提供重要见解。研究和广泛的影响活动将提供全面的培训，各级学生，从K-12，本科生，研究生，特别是妇女，代表性不足的少数民族学生。这一奖项反映了NSF的法定使命，并已被认为是值得通过评估使用基金会的智力价值和更广泛的影响审查标准的支持。

Nontechnical Summary:This project develops a new molecular delivery system for enabling the CRISPR-based genome editing technology. CRISPR-based genome editing technology can precisely add, delete, and “correct” genes in vitro or in vivo; therefore, it holds tremendous potential for therapeutic applications and is actively being pursued for addressing a wide range of human diseases ranging from hereditary diseases, cancer, to diabetes. However, one major obstacle to implementing this technology is the lack of safe and efficient delivery systems that can effectively transport the required molecular machinery into cells to execute genome editing. To address this challenge, this project proposes the design, synthesis, and investigation of a series of delivery molecules made from natural amino acids and peptides. These compounds will be evaluated for their efficiency and safety for delivering two pieces of RNAs into cells for performing CRISPR-based genome editing. Structure-function studies will shed light on important design principles for effective molecular carriers. The proposed interdisciplinary research activities also provide excellent trainings to students at various levels, ranging from K-12, undergraduate, to graduate students, especially women, underrepresented minority students, and those from institutes with limited research opportunities.Technical Summary:The goal of this project is to develop a new synthetic delivery platform for RNA-based CRISPR and conduct fundamental structure-property studies on the new delivery system to gain critical insights for the design of new gene delivery vectors. CRISPR/Cas9 gene editing offers a tremendous potential for therapeutic applications and is actively being pursued for addressing a wide range of human diseases. However, one major obstacle to implementing CRISPR-mediated genome editing is the lack of safe and efficient delivery vehicles. The PI lab recently developed an innovative design of bioreducible dendronized polypeptides (BDPs) that show high efficiency for co-delivery of Cas9 mRNA and gRNAs. It is proposed that further investigation of this system will generate a new platform delivery system that can effectively and safely co-deliver CRISPR/Cas9 mRNA and gRNA into various cells. Furthermore, capitalizing on BDP’s well-defined molecular structure, we propose to conduct fundamental structure-property studies on the BDP system with the aim to gain critical insights for the design of new gene delivery vectors. If successful, the proposed study will provide a new, general, and biocompatible delivery system for CRISPR technology and offer critical insights for guiding the design of new gene delivery systems. The research and broad impact activities will provide holistic trainings to students at various levels, ranging from K-12, undergraduate, to graduate students, especially women, underrepresented minority students.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.