CAREER: Acid-Transforming Polypeptides as Stimuli-responsive, Efficient, Biocompatible, and Tunable Nonviral Gene Carriers

ID：MPS/DMR/BMAT(7623)0956091 PI：KWON，Young Jik ORG：加州大学欧文分校题目：CALEAR：酸转化多肽作为刺激响应、高效、生物相容和可调的非病毒基因载体INTELLECTUAL优点：合成基因载体(或非病毒载体)有效地将基因输送到预期的生物靶点，具有高度的生物相容性和通用性，是基础和翻译基因治疗研究中的一项高需求技术。与病毒载体相比，非病毒载体在细胞外和细胞内的性能仍然相对较差，病毒载体虽然被证明是非常有效的，但通常是免疫原性的，很难通过修饰来改变其结构和功能。拟议的职业研究将合成新型的刺激转化多肽，这些多肽在中性pH下将核酸与阳离子臂复合，但在细胞室中水解后，会转化为中性的自然产生的多肽，从而破坏复合体的稳定并释放核酸。具体地说，该项目将研究含有酮化丝氨酸(KSer)残基的新型生物材料，KSer残基类似于赖氨酸(Lys)，但在酸解后转化为丝氨酸(Ser)。KSer残基将与赖氨酸残基、聚乙二醇(PEG)和其他功能多肽一起使用，以创建多功能聚合物非病毒载体。初步的体外研究表明，聚(kSer-Lys)/DNA复合体和聚乙二醇聚(KSer)/DNA胶束通过几种改进的细胞内途径显著提高了基因传递效率。PI建议通过更好地了解关键的细胞外和细胞内基因传递途径来开发这些基于酸转化多肽(ATPP)的新型基因载体。他将(1)研究ATPP与融合和核定位信号肽的杂交，以提高靶向性；(2)探索ATPP与可被此类刺激切断的功能肽和/或聚乙二醇组分之间的刺激敏感连接的效果，以提高疗效；(3)优化含ATPP的聚合物，以改善DNA络合、细胞内化、细胞内靶向、DNA释放和转基因。以生物兼容和多样化的方式对体细胞进行重新编程以诱导干细胞)。根据这项研究开发的新型生物材料将向本科生和高中生提供实践研究体验和暑期研究计划。建议的项目将激励本科生创造性和独立思考，并给他们一个机会用PI来检验他们的假设。该推广计划旨在激发参与计划的高中生对科学和工程职业的兴趣，特别是那些来自附近城市地区代表不足的群体的学生。建议的研究将通过本科生直接参与职业研究和外展研究计划，转化到课堂和其他领域。一门新的本科课程将介绍使用刺激响应性聚合物生物材料进行基因传递的基本概念；然后它将挑战学生在职业研究中发展创造性想法。具有最具创新性建议的学生将被邀请在PI的实验室测试他们的假设，并将帮助高中参与者参加一个使用酸转化多肽进行基因传递的暑期研究项目。这一外展计划将从国际和平协会的讲座和社会经济贫困地区当地高中的动手实验中发展而来。最有动力的高中生将参加一个暑期研究项目，然后在本科生导师的指导下开发一个科学博览会项目。这些教育活动的影响将通过学生的出版物和演讲以及他们对科学和工程专业职业的选择来评估。

ID: MPS/DMR/BMAT(7623) 0956091 PI: Kwon, Young Jik ORG: University of California-IrvineTitle: CAREER: Acid-Transforming Polypeptides as Stimuli-Responsive, Efficient, Biocompatible, and Tunable Nonviral Gene CarriersINTELLECTUAL MERIT: Synthetic gene carriers (or nonviral vectors) that efficiently deliver genes to desired biological targets with high biocompatibility and versatility are a technology in high demand in basic and translational gene therapy research. The extra- and intracellular performance of nonviral vectors remains relatively poor compared to viral vectors, which, though proven to be very efficient, are often immunogenic and difficult to modify in order to vary their structures and functions. The proposed CAREER research will synthesize novel, stimuli-transforming polypeptides that complex nucleic acids with a cationic arm under neutral pH, but which, upon hydrolysis in a cellular compartment, are converted to neutral, naturally occurring polypeptides, thus destabilizing the complex and releasing nucleic acids. Specifically, the project will investigate novel biomaterials that incorporate ketalized serine (kSer) residues, which are analogous to lysine (Lys) but which transform into serine (Ser) upon acid-hydrolysis. The kSer residues will be used along with lysine residues, polyethylene glycol (PEG) and other functional polypeptides to create versatile polymeric non-viral vectors. Preliminary in vitro studies have shown that poly(kSer-Lys)/DNA polyplexes and PEG-poly(kSer)/DNA micelles significantly enhanced gene delivery efficiency via several improved intracellular pathways. The PI proposes to develop these novel acid-transforming polypeptide (ATPP)-based gene carriers by better understanding key extra- and intracellular gene delivery pathways. He will (1) investigate hybridization of ATPPs with fusogenic and nuclear localization signal peptides in order to improve targeting, (2) explore the effects of stimuli-sensitive linkages between ATPPs and functional peptides and/or PEG that can be severed by such stimuli to improve efficacy, and (3) optimize the ATPP-containing polymers to improve DNA complexation, cellular internalization, intracellular targeting, DNA release, and transfection.BROADER IMPACTS: Efficiently delivering therapeutic genes, especially using nonviral methods, is of great interest not only in the clinic (e.g., gene therapy) but also in the genetic manipulation of biological phenomena (e.g., reprogramming somatic cells to be induced stem cells) in biocompatible and versatile ways. The novel biomaterials developed from this research will be available to undergraduate and high school students for a hands-on research experience and for a summer research program. The proposed project will motivate undergraduate students to think creatively and independently and give them a chance to test their hypotheses with the PI. The outreach program aims to stimulate interest in scientific and engineering careers among participating high school students, particularly those from underrepresented groups in nearby urban areas. The proposed research will be translated to classrooms and beyond through undergraduate students' direct participation in the CAREER research and an outreach research program. A new undergraduate course will introduce basic concepts of gene delivery using stimuli-responsive polymeric biomaterials; it will then challenge students to develop creative ideas on the CAREER research. Students with the most innovative suggestions will be invited to test their hypotheses in the PI's laboratory and will assist high school participants in a summer research program on gene delivery using the acid-transforming polypeptides. This outreach program will grow out of the PI's lectures in combination with hands-on experiments at local high schools in socio-economically disadvantaged areas. The most motivated high school students will participate in a summer research program, followed by developing a science fair project under the mentorship of the undergraduate student mentors. Impacts of these educational activities will be assessed by students' publications and presentations and by their choice of professional careers in science and engineering.