CAREER: Design and Characterization of Aptamer-Amphiphiles for Selective Binding

0846274Kokkoli智力优点：适体是短的单链寡核苷酸。它们是化学生产的，具有以高亲和力和特异性结合多种不同靶标的能力，并且已被证明可用作治疗剂和诊断工具。它们的亲和力通常与单克隆抗体观察到的亲和力相当，并且明显高于肽的亲和力。作为适配体两亲分子，适配体附着在疏水尾部，它们可以与其他两亲分子组装成各种不同的结构。它们与脂质组装形成适体功能化的脂质体将在生物技术应用中是理想的，例如靶向药物递送。 PI建议将适体作为适体两亲物进行研究，特别是结合fratalkine（一种仅在感染或炎症部位表达的粘附分子，例如癌症）的适体序列，并假设适体头基的方向以及疏水尾部和头基之间间隔物的使用将影响分子的组装行为和适体的二级结构，从而随后影响其与其结合的亲和力。目标。为了检验这一假设，设定了以下研究任务：在研究任务 1 中，目标是测试设计的适体两亲物（游离于溶液中和掺入脂质体中）的生物亲和力，从而评估哪些设计与 fractalkine 结合。 PI 将使用不同的技术（CD、熔解曲线、冷冻 TEM、SAXS 和 SANS）表征研究任务 2 中的适配体两亲物，从而评估适配体的二级结构和两亲物的组装行为。这些结果将使适配体两亲物的生物学功能与其结构和相应的相行为之间建立联系。这项研究将增进知识和理解，因为它不仅将为适体两亲物的设计提供有利的两亲特性（头基取向、间隔区设计、二级结构、组装行为），而且还将提供对为什么这些特性有益的基本理解。 更广泛的影响：-技术影响。 PI 提出了一种新工具“适体两亲物”的设计和表征，其基础是通过改变其结构片段可以控制其形态、表面化学和功能。这里首次提出了二级结构、两亲性和自组装特性的综合表征，以及这些特性与靶向运载工具工程之间的联系。这些研究活动代表了变革性研究，因为它们可能会导致“适配体两亲物”（文献中的一个新术语）研究新领域的起源，该领域将融合合理的两亲物设计原理、化学、生物学和工程学，以便为靶向药物输送提供先进的系统。 -社会/教育影响。 Fractalkine 是开发抗血管生成和抗癌药物的一个有吸引力的新靶点。 通过选择与 fractalkine 结合的适体作为模型系统，该工作概述了一种逻辑方法，可应用于未来设计适体两亲物，使纳米颗粒功能化并增加颗粒对 fractalkine 的特异性和亲和力，从而具有巨大的治疗潜力并显着造福社会。教育计划与研究计划紧密结合。 PI 正在与“DragonflyTV”（由 NSF 支持、双城公共电视台 (TPT) 制作、在全国公共广播服务 (PBS) 电视台播出的科学系列节目“DragonflyTV”）合作，旨在增强儿童和成人对科学技术的了解。 此外，PI 已经参加并将继续参加 TPT 组织的外展活动，其目标是吸引女性从事科学和工程领域的工作（PI 的长期目标），并且目前正在并将在未来继续与 MRSEC 合作。通过 MRSEC，PI 将在暑期接待全国申请者中的高中教师和少数族裔本科生，通过完善的高中网络以及来自一些少数族裔服务机构的少数族裔本科生。

0846274KokkoliIntellectual Merit: Aptamers are short single-stranded oligonucleotides. They are chemically produced, have the capability of binding with high affinity and specificity a variety of different targets, and have been shown to be useful as therapeutic agents and diagnostic tools. Their affinities are often comparable to those observed for monoclonal antibodies, and are significantly higher compared to those of peptides. As aptamer-amphiphiles, aptamers attached to a hydrophobic tail, they can assemble with other amphiphilic molecules into a variety of different structures. Their assembly with lipids to form aptamer-functionalized liposomes will be desirable in biotechnological applications, such as, targeted drug delivery. The PI proposes to study aptamers as aptamer-amphiphiles and, in particular, aptamer sequences that bind fractalkine (an adhesion molecule expressed only at sites of infection or inflammation, such as cancer), and hypothesizes that the orientation of the aptamer headgroup, and the use of a spacer between the hydrophobic tail and the headgroup, will affect the assembly behavior of the molecule, and the secondary structure of the aptamer, thus subsequently affecting its binding affinity for its target. To test this hypothesis the following research tasks have been set: In Research Task 1, the goal is to test the biological affinity of the designed aptamer-amphiphiles, free in solution, and incorporated in liposomes, thus evaluating which designs bind to fractalkine. The PI will characterize the aptamer-amphiphiles in Research Task 2 with different techniques (CD, melting curves, cryo-TEM, SAXS, and SANS) that will evaluate the secondary structure of the aptamer, and assembly behavior of the amphiphiles. These results will enable a connection between the biological function of aptamer-amphiphiles with their structure and corresponding phase behavior. This research will advance knowledge and understanding as it will not only furnish favorable amphiphilic properties (headgroup orientation, spacer design, secondary structure, assembly behavior) for the design of aptamer-amphiphiles, but will provide a fundamental understanding of why these properties are beneficial. Broader Impact: -Technical Impact. The PI proposes the design and characterization of a novel tool, "aptamer-amphiphiles", based on the expectation that by changing their structural segments one can control their morphology, surface chemistry, and function. A comprehensive characterization of secondary structure, amphiphilic and self-assembly properties, and the link between these properties and the engineering of targeted delivery vehicles is proposed here for the first time. The research activities represent transformative research as they may lead to the genesis of a new field of research on "aptamer-amphiphiles" (a new term in the literature) that will merge rational amphiphile design principles, chemistry, biology, and engineering in order to provide advanced systems for targeted drug delivery. -Societal / Educational Impact. Fractalkine is an attractive new target for the development of anti-angiogenesis and anti-cancer agents. By choosing aptamers that bind to fractalkine as the model system, the proposed work outlines a logical approach that can be applied to the future design of aptamer-amphiphiles that can functionalize nanoparticles and increase the particle's specificity and affinity for fractalkine, thus having great therapeutic potential and significantly benefiting society. The educational plan is strongly integrated with the research plan. The PI is collaborating with "DragonflyTV" (a science series that presents middle school students doing real science, supported by NSF and produced by Twin Cities Public Television (TPT), and seen on Public Broadcasting Service (PBS) stations nationwide) with the goal of enhancing scientific and technological understanding among children and adults. Furthermore, the PI has participated, and continue to participate, in outreach activities organized by TPT with the goal of attracting women in sciences and engineering (a long term goal for the PI), and is currently collaborating, and will continue to do so in the future, with MRSEC. Through MRSEC the PI will be hosting during the summer high school teachers, and minority undergraduates from a nationwide pool of applicants, through a well established network of high schools, and minority undergraduates from a number of Minority Serving Institutions.