Novel organic-ion-based technology for long-term virus preservation at ambient temperature

项目概要 大多数目前可用的疫苗，特别是活的和基于mRNA的COVID-19疫苗，是温度敏感的， 需要严格的冷链维护，需要在推荐的温度下储存和分配， 从生产到管理。这一必要性给全球性的国际移民设置了最令人望而却步的障碍， 特别是在发展中国家，免疫计划占成本交付的80%。因此，在本发明中， 迫切需要一种技术来提供成本有效的和长期的环境温度存储， 无需冷链或复杂的样品回收方案。 该提案旨在开发一种用于在环境温度下长期储存病毒的有机离子平台， 在面对新疫苗需求不断增长的情况下， 与当前生物库技术相关的维护。离子液体（Ionic Liquids，ILs）是完全由无机盐组成的离子液体， 离子的选择提供了一个非常合适的平台，在该平台上，可以通过选择离子来改变性质，从而使 用于病毒稳定化的溶剂/介质的可调设计。我们假设，建议的离子液体的解决方案， 约20重量%的水可以防止病毒基因组和蛋白质衣壳的水解和酶促降解， 提供了一种可靠的方法来保存病毒。我们将使用来自肌病毒家族的噬菌体作为 裸蛋白颗粒的实例和登革病毒作为脂质包膜颗粒的实例。一是 通过杂环阳离子的系统变化开发一个深思熟虑的新型离子液体库， 亲液阴离子，并明智地将两个官能团（NH3+和SO 2F）引入IL结构中。 将通过配对新类型的生物相容性阳离子和阴离子来实现结构可变性。二是 通过评估其结构完整性、热稳定性， 保质期从六个月到四年。我们将监测病毒二级结构的变化，热变性- 以及颗粒形态。最后，我们将研究它们的经验结构-活性关系，以获得比较。 深入了解病毒之间相互作用的结合特征和分子机制， 粒子和目标的含水离子溶剂通过模拟，晶体学，和光谱方法。 该项目将为病毒的环境温度保存提供可行的解决方案， 通过开发对水解稳定的病毒载体和水基质， 和酶降解。所提出的方法的另一个重要特征是这些核酸-IL 溶液可以直接通过PCR扩增，而无需进行预先的提取、纯化或定量。 是的。这种方法具有简单的优点，这使得环境温度储存和 分布非常有效，增加了生物样品的长时间稳定性，减少了操作 成本和碳足迹，并改善病毒和基于病毒的技术的物流。 总结

PROJECT SUMMERY Most currently available vaccines, especially live and mRNA-based COVID-19 vaccines, are temperature sensi- tive and require stringent cold-chain maintenance, entailing their storage and distribution at recommended tem- peratures from production to administration. This necessity imposes the most prohibitive barrier to global im- munization programs, particularly in developing countries, accounting for up to 80% of the cost delivery. Thus, there is a critical need for a technology to provide cost-effective and long-term ambient temperature storage for viral samples without requiring cold chain or complicated sample recovery protocols. This proposal aims to develop an organic-ion platform for long-term storage of viruses at ambient tem- perature to potentially reduce costs in the face of growing needs for new vaccines and avoid labor-intensive maintenance associated with current biobanking technology. Ionic liquids (ILs)  organic salts comprised entirely of ions  offer a well-suited platform on which the properties can be altered by the selection of ions, enabling the tunable design of solvents/media for virus stabilization. We hypothesize that the solutions of proposed ILs with ca. 20 wt% water may prevent hydrolytic and enzymatic degradation of viral genomes and protein capsids, providing a reliable approach to preserve viruses. We will use a bacteriophage from the myovirus family as an example of a naked protein particle and dengue virus as an example of a lipid-enveloped particle. First, we will develop a thoughtfully conceived library of novel ILs through systematic variations of heterocyclic cations and kosmotropic anions, and judicious incorporation of two functionalities (NH3+ and SO2F) into the IL structures. Structural variability will be achieved by pairing new genre of biocompatible cations and anions. Second, we will examine their effectiveness for stabilizing viruses by evaluating their structural integrity, thermostability, and shelf-life from six months and four year. We will monitor changes in viral secondary structure, thermal denatur- ation, and particle morphology. Last, we will study their empirical structure-activity relationships to gain compre- hensive understanding of binding characteristics and molecular mechanisms of interactions between the viral particles and the targeted aqueous ionic solvents via simulation, crystallographic, and spectroscopic methods. This project will provide a viable solution for ambient temperature preservation of viruses for extended periods (potentially for decades) by developing the virusILwater matrices that are stable towards hydrolytic and enzymatic degradation. Another important feature of the proposed approach is that these nucleic acid-ILs solutions can be directly amplified by PCR without being subjected to prior extraction, purification or quantifica- tion. This approach has the merit of simplicity, which makes the process of ambient temperature storage and distribution profoundly efficient, increases the stability of biosamples for prolonged time, reduces operational costs and carbon footprint, and improves logistics for viruses and virus-based technologies. Summery