Tipping William J, Merchant Andrew S, Fearon Rebecca, Tomkinson Nicholas C O, Faulds Karen, Graham Duncan
Centre for Molecular Nanometrology, WestCHEM, Department of Pure and Applied Chemistry, Technology and Innovation Centre, University of Strathclyde Glasgow G1 1RD UK
Department of Pure and Applied Chemistry, University of Strathclyde Glasgow G1 1XL UK
RSC Chem Biol. 2022 Aug 9;3(9):1154-1164. doi: 10.1039/d2cb00160h. eCollection 2022 Aug 31.
Stimulated Raman scattering (SRS) microscopy is a powerful technique for visualising the cellular uptake and distribution of drugs and small molecules in live cells under biocompatible imaging conditions. The use of bio-orthogonal groups within the drug molecule, including alkynes and nitriles, has enabled the direct detection of a plethora of bioactive molecules in a minimally perturbative fashion. Limited progress has been made towards real-time detection of drug uptake and distribution into live cells under physiological conditions, despite the accordant potential it presents for preclinical drug development. SRS microscopy has been applied to the study of cellular dynamics of the drug 7RH, which is a potent inhibitor of dicoidin domain receptor 1 (DDR1) and prevents cellular adhesion, proliferation and migration . The uptake of 7RH into a variety of mammalian cell models was shown to be independent of DDR1 expression. Using a perfusion chamber, the recurrent treatment of live cancer cells was achieved, enabling 7RH uptake to be visualised in real-time using SRS microscopy, after which the viability of the same cellular population was assessed using commercially available fluorescent markers in a multimodal imaging experiment. The effect of 7RH treatment in combination with the chemotherapeutic, cisplatin was investigated using sequential perfusion and time-lapse imaging in the same live cell population, to demonstrate the application of the approach. SRS microscopy also identified potent inhibition of cellular adhesion and migration in breast cancer cell models with increasing 7RH treatment concentrations, thus representing a novel read-out methodology for phenotypic assays of this kind. The direct assessment of drug-cell interactions under physiological conditions offers significant potential for the preclinical drug development process.
受激拉曼散射（SRS）显微镜是一种强大的技术，可在生物相容性成像条件下可视化活细胞中药物和小分子的细胞摄取及分布情况。在药物分子中使用生物正交基团，包括炔烃和腈，能够以最小程度的干扰直接检测大量生物活性分子。尽管其在临床前药物开发方面具有潜在优势，但在生理条件下实时检测药物进入活细胞的摄取和分布方面进展有限。SRS显微镜已应用于对药物7RH细胞动力学的研究，7RH是盘状结构域受体1（DDR1）的强效抑制剂，可阻止细胞粘附、增殖和迁移。研究表明，7RH在多种哺乳动物细胞模型中的摄取与DDR1表达无关。使用灌注室实现了对活癌细胞的反复处理，从而能够利用SRS显微镜实时观察7RH的摄取情况，之后在多模态成像实验中使用市售荧光标记物评估同一细胞群体的活力。在同一活细胞群体中，通过顺序灌注和延时成像研究了7RH与化疗药物顺铂联合治疗的效果，以证明该方法的应用。SRS显微镜还发现，随着7RH处理浓度的增加，乳腺癌细胞模型中的细胞粘附和迁移受到强效抑制，因此代表了一种用于此类表型分析的新型读出方法。在生理条件下直接评估药物与细胞的相互作用为临床前药物开发过程提供了巨大潜力。
