Establishing a chemoenzymatic flow-based strategy for nucleoside synthesis

建立基于化学酶流的核苷合成策略

• 批准号: 2597126
• 金额: --
• 依托单位: University of Strathclyde
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2021
• 资助国家: 英国
• 起止时间: 2021 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2597126
• 关键词: Establishing; chemoenzymatic; flow; based; strategy

Nucleoside analogues are essential building blocks used throughout the biotechnology and pharmaceutical industries. Applications of nucleoside analogues are diverse, ranging from their incorporation into oligonucleotides for diagnostics (e.g., PCR) and antisense therapeutics, through to their utility as small molecule anti-virals. With the advent of the Covid-19 pandemic, the global market for novel nucleosides is growing rapidly, expecting to reach US$809.3M by 2022. Underpinning this growth (from US$113.3M in 2014) is the need for innovative strategies to enable their efficient and sustainable synthesis, and large scale manufacture. Despite the pervasiveness of nucleosides used throughout industry and in academia, methods of synthesis have not kept abreast of their downstream applications. This is predominantly due to conventional synthetic methodology suffering from the formation of side-products as well as the use of toxic reagents which results in downstream scale-up being environmentally unsustainable. In contrast to the challenging nature of using pure synthetic routes, enzymatic methods of nucleoside synthesis using for example, purine/pyrimidine nucleoside phosphorylases (PNPs), are far more sustainable, scalable and predominantly produce the desired stereoisomer. This provides considerable opportunities to integrate enzymatic and chemical syntheses to produce nucleoside analogues not readily accessible by the use of these strategies in isolation. In order to integrate each of the respective processes (i.e., synthesis of precursors by chemical synthesis and enzymatic glycosylation to form the nucleoside core), we propose to develop a facile synthetic 'production line' using flow chemistry. Flow chemistry offers the potential to telescope reactions in a far more efficient and cost-effective manner relative to conventional batch-based processes. Furthermore, by exploiting experience of the Burley and Hoskisson groups in developing enzymes for applications in biocatalysis will enable our collaboration to tailor these enzymes for flow-based synthesis. In addition, using the Burley group's existing experience and links with a flow-based industrial partner (Vaportec) will provide additional opportunities to streamline the development of both the enzymatic and synthetic chemistry parts of the project. OVERALL OBJECTIVES OF THE STUDENTSHIPThe overall objective of this collaborative studentship proposal is to establish a new flow-based chemoenzymatic strategy to synthesise nucleoside analogues. Central to this work is delivering a sustainable synthetic method of synthesis in which the building blocks will feed directly into our larger BBSRC-funded network (strategic longer-larger grant, sLoLa, 2020-2025) in which modified RNA oligonucleotides are essential tools to interrogate how gene expression is regulated.The specific aims of the proposal are to: (i) identify novel glycosylation enzymes to prepare nucleoside analogues. (ii) determine the compatibility of these enzymes for use in a flow-based reactor system. (iii) establish the utility of the flow-based approach for the chemoenzymatic synthesis of novel nucleosides for their incorporation into oligonucleotides.

核苷类似物是整个生物技术和制药行业使用的基本构件。核苷类似物的用途多种多样，从用于诊断(例如，聚合酶链式反应)和反义治疗的寡核苷酸，到作为小分子抗病毒药物的用途。随着新冠肺炎大流行的到来，全球对新型核苷的市场正在迅速增长，预计到2022年将达到8.093亿美元。支撑这一增长(2014年为1.133亿美元)的是对创新战略的需求，以实现高效和可持续的合成和大规模制造。尽管核苷在工业和学术界中广泛使用，但合成方法并没有跟上它们在下游的应用。这主要是因为传统的合成方法存在副产物的形成以及有毒试剂的使用，这导致下游放大在环境上是不可持续的。与使用纯合成路线的挑战性相比，使用例如嘌呤/嘧啶核苷磷酸化酶(PNPs)的酶促合成核苷的方法更可持续、可扩展，并且主要产生所需的立体异构体。这为整合酶和化学合成来生产核苷类似物提供了相当大的机会，这些策略很难通过孤立地使用这些策略来获得。为了将各个过程(即通过化学合成合成前体和通过酶促糖基化形成核苷核心)相结合，我们建议使用流动化学开发一条简便的合成‘生产线’。与传统的间歇过程相比，流动化学提供了以更有效和更具成本效益的方式观察反应的潜力。此外，通过利用Burley和Hoskison团队在开发用于生物催化的酶方面的经验，我们的合作将使我们能够为基于流动的合成定制这些酶。此外，利用Burley集团现有的经验和与基于流程的工业合作伙伴(Vaportec)的联系，将提供更多机会来简化该项目的酶和合成化学部分的开发。这项合作学生计划的总体目标是建立一种新的基于流动的化学酶策略来合成核苷类似物。这项工作的核心是提供一种可持续的合成方法，其中构建块将直接馈入我们更大的BBSRC资助的网络(战略性更长-更大的赠款，sLoLa，2020-2025年)，其中修饰的RNA寡核苷酸是询问基因表达如何调控的重要工具。该提案的具体目标是：(I)鉴定新的糖基化酶以制备核苷类似物。(2)确定用于流动反应器系统的这些酶的配伍性。(3)确定以流动为基础的方法用于化学酶法合成新的核苷，以便将其并入寡核苷酸。