High throughput 2D-IR protein screening - new technology for drug design and diagnostics

二维红外(2D-IR)是一种超快激光光谱学方法，它提供了生物大分子(如蛋白质和DNA)的3D结构和分子间相互作用的灵敏‘图谱’。2D-IR是对结晶学、低温EM或核磁共振等成熟的基于结构的生物物理工具的补充，它在室温下的溶液相中运行，并提供对分子结构的超快动态运动的访问，这些运动会影响配体结合，因此是下一代药物分子的目标。这意味着2D-IR有可能作为一种工具用于药物设计或测量复杂混合物中的蛋白质含量，如生物体液(如血清)。然而，2D-IR商用的一个主要障碍是在光谱的红外区对水(H2O)的强吸收，这掩盖了蛋白质和核酸的重要特征。来自溶剂和生物分子的信号的这种重叠需要在测量之前用D2O昂贵且劳动密集型的同位素替换溶剂，使得2D-IR不能作为制药业的商业工具并且无法测量生物流体。这项建议建立在中央激光设施的世界领先的STFC资助的研究基础上，该研究使用于测量2D-IR的激光脉冲序列能够首次抑制H2O信号(《化学科学》10,6448(2019)；编辑精选一周和热门文章集)。这意味着蛋白质的信号可以在没有溶剂替代的情况下被干净地测量；蛋白质光谱学相当于在大海捞针。我们的发明，具有成熟的知识产权(PCT/GB2019/051585)，意味着我们可以在生理相关(基于水的)溶剂和生物流体中对蛋白质的结构和动力学进行2D-IR光谱测量，这消除了2D-IR商业化实施的主要障碍，从而为制药和生物医学部门打开了2D-IR的能力。为了实现这一商业潜力，我们与全球生物制药公司UCB Pharma和STFC中央激光设施建立了合作伙伴关系，以创建世界上第一个高通量2D-IR筛选设施。通过克服商业2D-IR实施面临的下一系列挑战，如快速、准确的样品传输和用户友好的数据可视化，该设施将使2D-IR发展到更高的TRL水平(3-7)，最终能够测量标准96孔亚微升体积的药物蛋白质样品，并在按下按钮不到两小时内提供可供分析的光谱。该设施将为最终用户提供直接访问生物分子靶标和候选药物之间相互作用的结构和动态细节的途径，并为药物设计带来重要的新视角。在我们开发药物设计筛选平台的主要目标的同时，我们将进一步开发这项技术，与其他合作伙伴，如SME ClinSpec DX合作，推进生物液的2D-IR筛查，为疾病诊断和医疗保健应用提供生物液的定量蛋白质分析。该项目的成功完成将为CLF提供商业上可行的按样品收费的2D-IR筛查服务奠定基础，并建立2D-IR分析的最佳上市途径。

Two-dimensional infrared (2D-IR) is an ultrafast laser spectroscopy method that provides a sensitive 'map' of the 3D structures and intermolecular interactions of biomacromolecules, such as proteins and DNA. 2D-IR complements well-established structure-based biophysical tools like crystallography, cryo-EM or NMR by operating in the solution phase at room temperature and by providing access to ultrafast dynamic motions of molecular structures, which influence ligand binding and so are targets for next-generation drug molecules. This means that 2D-IR has potential as a tool for use in drug design or in measuring protein content in complex mixtures such as biofluids (e.g. blood serum). However, a major barrier to the commercial uptake of 2D-IR has existed in the strong absorption of water (H2O) in the infrared region of the spectrum, which obscured important signatures from proteins and nucleic acids. This overlap of signals from solvent and biomolecule necessitated expensive and labour-intensive isotopic replacement of the solvent with D2O before measurements, making 2D-IR non-viable as a commercial tool for the pharmaceutical industry and incapable of measuring biofluids.This proposal builds on world leading STFC-funded research at the Central Laser Facility that enables the laser pulse sequence used to measure 2D-IR to suppress H2O signals for the first time (Chem Sci 10, 6448 (2019); Editors' Pick of the Week and HOT article collections). This means that the signals from proteins can be cleanly measured without solvent substitution; the protein spectroscopy equivalent of locating the needle in the haystack. Our invention, with established IP (PCT/GB2019/051585) means that we can perform 2D-IR spectroscopy measurements of protein structure and dynamics in physiologically relevant (H2O-based) solvents and biofluids, which removes the major obstacle to commercial implementation of 2D-IR and so opens up the power of 2D-IR to the pharmaceutical and biomedical sectors. To realise this commercial potential, we have formed a partnership with global biopharmaceutical company UCB Pharma and the STFC Central Laser Facility to create the world's first high throughput 2D-IR screening facility. By overcoming the next set of challenges facing commercial 2D-IR implementation, such as rapid, accurate sample delivery and user-friendly data visualisation, the facility will progress 2D-IR to higher TRL levels (3-7), ultimately being able to measure a standard 96-well plate of sub-microlitre volume drug-protein samples and deliver analysis-ready spectra in less than two hours at the press of a button.This facility will deliver straightforward access for end-users to the structural and dynamic details of interactions between biomolecule target and drug candidate and bring a vital new perspective to drug design. In parallel with our primary aim of developing a screening platform for drug design, we will develop the technology further, working with other partners, such as SME ClinSpec DX, to advance 2D-IR screening of biofluids, delivering quantitative protein analysis of biofluids for disease diagnosis and healthcare applications.Successful completion of the project will lay the groundwork for a commercially viable fee-per-sample 2D-IR screening service at the CLF and establish the optimum route to market for 2D-IR analysis.