Ma Lijuan, Ma Chaofu, Wang Zijian, Wei Yunan, Li Nan, Wang Jing, Li Mingshuang, Wu Zhisheng, Du Yang
Kobilka Institute of Innovative Drug Discovery, School of Medicine, The Chinese University of Hong Kong, Shenzhen 518172, China.
School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing 100029, China.
Anal Chem. 2025 Jan 21;97(2):1199-1209. doi: 10.1021/acs.analchem.4c04442. Epub 2025 Jan 9.
The development of multitargeted drugs is urgent for ischemic stroke. TRPV1 and TRPM8 are important targets of ischemic stroke. Previous drug candidate screening has identified that muscone, l-borneol, and ferulic acid may target TRPV1 and TRPM8 for ischemic stroke. However, the mechanisms of these drug candidates on targets were ill-informed. Therefore, firstly, a tongue-tissue biosensor was constructed. It explored the activation or inhibition mechanisms of drug candidates targeting TRPV1 and TRPM8 in a near-physiological environment. It was found that muscone could specifically inhibit TRPM8 and selectively activate TRPV1, while l-borneol exhibited the opposite effect. It suggested a synergistic network between these two drug candidates. Furthermore, more selective protein biosensors were developed to delve deeper into the synergistic mechanisms. A strong synergistic effect of muscone and l-borneol was proved. Molecular docking revealed that the synergistic effect was caused by different action sites, respectively. Subsequently, the synergistic effect of muscone and l-borneol was further confirmed by hypoxic nerve injury models of () and antithrombus and anti-ischemic models of zebrafish. Ultimately, through nontargeted metabolomics, it was found that muscone and l-borneol mainly regulated Ca concentration and energy metabolism by pathways such as purine and amino acid metabolisms. In conclusion, this research identified critical targets and synergistic drug candidates for multitarget neuroprotection of ischemic stroke. In addition, it has systemically demonstrated the feasibility of the integration of tissue/protein biosensors and metabolomics for the research and development of multitarget drugs. Compared to other screening and validation methods for drugs and targets, the biosensors we developed not only achieved higher sensitivity and specificity in complex physiological environments, ensuring a wider detection range, but also greatly saved biological samples. Simultaneously, they could be extended to other complex systems, such as biomarker screening in clinical samples and exosomes isolated from stem cells.
开发多靶点药物对缺血性中风治疗至关重要。瞬时受体电位香草酸亚型1（TRPV1）和瞬时受体电位 melastatin 8型（TRPM8）是缺血性中风的重要靶点。先前的候选药物筛选已确定麝香酮、l-冰片和阿魏酸可能针对TRPV1和TRPM8用于缺血性中风治疗。然而，这些候选药物作用于靶点的机制尚不明确。因此，首先构建了一种舌组织生物传感器。它在接近生理的环境中探索了候选药物靶向TRPV1和TRPM8的激活或抑制机制。研究发现，麝香酮可特异性抑制TRPM8并选择性激活TRPV1，而l-冰片则表现出相反的效果。这表明这两种候选药物之间存在协同网络。此外，还开发了更具选择性的蛋白质生物传感器以深入研究协同机制。结果证明了麝香酮和l-冰片具有很强的协同作用。分子对接显示，这种协同作用分别是由不同的作用位点引起的。随后，通过（）的缺氧神经损伤模型以及斑马鱼的抗血栓和抗缺血模型进一步证实了麝香酮和l-冰片的协同作用。最终，通过非靶向代谢组学发现，麝香酮和l-冰片主要通过嘌呤和氨基酸代谢等途径调节钙浓度和能量代谢。总之，本研究确定了缺血性中风多靶点神经保护的关键靶点和协同候选药物。此外，它系统地证明了组织/蛋白质生物传感器与代谢组学相结合用于多靶点药物研发的可行性。与其他药物和靶点的筛选及验证方法相比，我们开发的生物传感器不仅在复杂的生理环境中实现了更高的灵敏度和特异性，确保了更广泛的检测范围，而且极大地节省了生物样本。同时，它们可扩展到其他复杂系统，如临床样本中的生物标志物筛选以及从干细胞中分离的外泌体。
