Exploration of Linking Chemistry in the Design of Aptamer-Molecularly Imprinted Polymer Hybrids (aptaMIPs)

分子印迹涉及在聚合物中制造一个结合袋，该结合袋在化学上和形状上对目标化合物具有特异性。与抗体和酶等生物分子识别元件相比，这些“智能塑料”具有更强的稳健性。它们还具有在极端环境条件下工作的能力。然而，它们有时可能缺乏必要的特异性/亲和力。适配体是DNA/RNA的小片段，具有靶向蛋白质和小分子的能力，并以高特异性和亲和力与它们结合。它们无毒，是抗体的有吸引力的替代品。由于它们对酶和化学降解的敏感性，它们主要用于研究，尽管这种情况正在慢慢改变，它们正变得具有商业意义。到2020年，全球适配体市场预计将从2015年的11亿美元增长到24亿美元。在EPSRC的支持下，PI（分子印迹专家）领导了一项为期12个月的概念验证研究，通过将适体掺入分子印迹聚合物（MIPs）中，创造出了新型杂交材料。简而言之，适体结构被修改，使其能够直接与聚合物结合，因此它将在不受环境条件影响的情况下保持其形状。创造了新颖、高亲和力和稳定的材料。这些“aptaMIPs”表现出卓越的分子识别能力，并在稳定性和特异性靶标识别方面对mip和适配体进行了重大改进，有效地保持了两类材料的最佳性能。该提案旨在通过对用于创建这些新材料的核心化学物质进行为期两年的研究来探索aptaMIPs的潜力。我们将以试点研究的结果为基础，创造有用、有效、具有高商业潜力的材料。本建议的研究将集中于：(i)确定正确的连接剂化学；开发所有四种碱基的可聚合改性；确定需要多少联系单位；确定这些联系的最佳位置。对这四点的深入研究将使我们能够充分理解适配体如何将自身融入聚合物的关键化学过程，并通过这一点，使我们能够理解是什么造就了一个好的aptaMIP，以及为什么。除此之外，还将分析所使用的综合策略，以确保这些杂交品种的创造简单有效。已经选择了两个目标来研究这些化学物质。它们在大小和用途上有所不同：一种是蛋白质，一种是生物活性药物，但两者都有巨大的商业潜力。通过这些模型系统，我们旨在证明aptaMIP材料的有效性和潜力。除了PI之外，还有两个项目合作伙伴组成了研究小组：Watts小组是试点研究的合作者，他们位于马萨诸塞大学RNA治疗研究所（一所世界领先的新型适体合成学校）。他们将通过获得最先进的合成设备，结合低聚物合成和应用的专业知识来支持该提案。Aptamer Group是一家位于约克郡的商业适配体开发公司。他们的专业知识将通过提供已知的寡聚物序列使该项目受益，这些序列将作为我们研究和获得专业仪器的基础。他们对合适的油田和商业前景的详细了解将支持该项目的影响。整个团队的经验将使这个跨学科的提案，涵盖聚合物，核酸，蛋白质和分析化学领域的成功。我们将从现有的概念验证阶段开始，开发aptaMIPs及其合成过程，使其成为人工分子识别领域可行的竞争对手，准备在其功能可以利用的系统中应用。

Molecular imprinting involves making a binding pocket in a polymer which is chemically and shape specific for the target compound. These "smart plastics" offer robustness compared to biological molecular recognition elements such as antibodies and enzymes. They also have the ability to work in extreme environmental conditions. However, they can sometimes lack the necessary specificity/affinity. Aptamers are small pieces of DNA/RNA that have the ability to target proteins and small molecules and bind to them with high specificity and affinity. They are not toxic and are attractive alternatives to antibodies. They have been used primarily in research due to their susceptibility to enzymatic and chemical degradation, though this is slowly changing and they are becoming commercially relevant. The global aptamers market is projected to reach $2.4 billion by 2020, up from $1.1 billion in 2015.A 12-month proof-of-concept study, supported by the EPSRC and led by the PI (a molecular imprinting specialist), created novel hybrid materials made by incorporating aptamers into molecularly imprinted polymers (MIPs). In simple terms, the aptamer structure is modified to allow it to be directly incorporated into a polymer, so it will hold its shape while being protected from environmental conditions. Novel, high affinity and stable materials were created.These "aptaMIPs" demonstrated exceptional molecular recognition and offer significant improvements on both MIPs and aptamers in terms of stability, and specific target recognition, effectively maintaining the best properties of both classes of materials. This proposal seeks to explore the potential of aptaMIPs through a two year study into the core chemistry used to create these novel materials. We will build on the results of the pilot study and create useful, effective materials with high commercial potential.The research in this proposal will focus on: (i) Identifying the right linker chemistry;(ii) Developing polymerisable modifications for all four bases;(iii) Identifying how many linkers are needed;(iv) Identifying the best position for these linkers. An in-depth study on these four points will enable a full understanding of the key chemistry of how the aptamer incorporates itself into the polymer and, through this, allow us to understand what makes a good aptaMIP and why. Alongside these the synthetic strategies used will be analysed to ensure the creation of these hybrids is simple and effective.Two targets have been selected to study these chemistries. These differ in size and application: a protein and a bioactive drug, but both targets have significant commercial potential. Through these model systems we aim to demonstrate the validity and potential of aptaMIP materials.Alongside the PI, two project partners form the research team:The Watts group were collaborators on the pilot study and are based at the University of Massachusetts RNA Therapeutics Institute (a world leading school in novel aptamer synthesis). They will support the proposal through access to state-of-the-art synthesis equipment, combined with know-how in oligomer synthesis and application.Aptamer Group are a commercial aptamer development company based in York. Their expertise will benefit the project by providing the known oligomer sequences which will act as the basis for our studies and access to specialised instrumentation. The impact of the project will be supported by their detailed knowledge of the aptamer field and commercial outlook.The experience of the whole team will allow this interdisciplinary proposal, covering the fields of polymer, nucleic acid, protein and analytical chemistries to succeed. We will take aptaMIPs from the existing proof-of-concept stage and develop them, and their synthetic process, into viable competitors in artificial molecular recognition, ready for application in systems where their functionality can be exploited.