一氧化氮及其下游产物过氧亚硝基新型有机荧光探针的构建及生物传感研究

Nitric oxide (NO) and its downstream product peroxynitrite (ONOOˉ) play key roles in various physiological systems, and their down-regulation are closely linked with various pathophysiological conditions including the endothelial dysfunction, cancer, neurodegenerative disease, Alzheimer’s disease and other disorders. In recent ten years, much attention has been paid to detecting and imaging the two important species in biological systems by fluorescence techniques. However, due to their very short lifetime, low steady-state concentration as well as the potential interference from other reactive oxidative species, selectively, sensitively and fastly probing NO or ONOOˉ in biological systems has been a focal point. It has been recognized that mitochondrial is the major site for the formation of intracellular NO and ONOOˉ, and lysosomal functions are also closely related to the two species. However, only a handful of mitochondria or lysosomal-targetable fluorescent probes for NO or ONOOˉ were reported to date. In this project, we present a new type of aromatic secondary amine-based fluorescent probes for NO and a new type of aromatic tertiary amine-based fluorescent probes for ONOOˉ for the purpose of addressing the limitations of poor selectivity and slow response for most of the reported NO or ONOOˉ fluorescent probes. Natobly, most of the fluorescent probes in the project are designed to be mitochondria or lysosomal-targetable, and thus are highly valuable for in-depth studying the NO/ONOOˉ-related physiological and pathological roles. The project feasibility has been supported by our preliminary studies. As far as we know, the design strategies in the project have never been reported previously.

一氧化氮（NO）和它的下游产物过氧亚硝基（ONOOˉ）在许多生理过程中扮演了重要角色，它们在生物体内平衡的失调可导致内皮机能障碍、癌症、神经退化、老年痴呆等疾病。由于NO和ONOOˉ低的稳态浓度和短的寿命，如何在复杂的生物系统内实现它们的高选择性且快速的荧光传感，是当前荧光探针领域研究的重要的挑战。线粒体是NO和ONOOˉ合成的主要细胞器，溶酶体功能也与NO和ONOOˉ密切相关，然而能靶向线粒体及溶酶体的NO和ONOOˉ荧光探针屈指可数。本项目拟系统开发一类芳香二级胺型NO荧光探针和一类芳香三级胺型ONOOˉ荧光探针，重点解决大部分已报到的相关荧光探针在选择性和响应速度方面的限制。另外，本项目所设计的NO和ONOOˉ荧光探针主要是线粒体和溶酶体靶向型荧光探针，将为深层次研究NO和ONOOˉ的生理病理功能奠定重要的化学基础。本项目的可行性已得到初步研究的证实。本项目的设计策略未见报到。

在本项目的支持下，开发了基于芳香一级胺、二级胺、三级胺反应基团的生物活性氮分子（RNS）荧光探针及其线粒体、溶酶体靶向衍生物，提高了生物信使分子一氧化氮（NO）及生物强氧化剂过氧亚硝基（ONOO-）的选择性，并将传感时间提高到了秒级，相关探针已在糖尿病和缺血再灌注损伤发病机理研究、癌细胞荧光诊断、巨噬细胞M1/M2表型的区分以及M1型巨噬细胞对癌细胞免疫响应研究中成功应用，并有望进一步应用于抗癌免疫药物的筛选和评价；发展了激发态d*-pi*相互作用介导的、发射波长大于700nm的“砜罗丹明”荧光染料，发展了溶酶体靶向的、发射波长位于远红外区的“胺基硅罗丹明”双光子荧光染料，并通过对分子内扭曲的电荷转移机制（TICT）的调控大幅度提高了罗丹明及其近红外荧光染料在水中的荧光量子收率；开发了基于取代-重排级联反应的生物硫醇荧光探针，实现了细胞内半胱氨酸（Cys）和谷胱甘肽（GSH）的选择性双通道荧光传感以及癌细胞/肿瘤组织的特异性荧光诊断，相关诊断试剂已在癌症患者的肿瘤组织切片上得到验证，并有望进一步应用于术中肿瘤边界的快速评估。相关研究成果分别发表在Chem. Sci.、Biomaterials、ACS Appl. Mater. Interfaces、Chem. Commun.、J. Mater. Chem.等学术期刊，共计23篇，其中Sci收录21篇。