Design And Synthesis Of Novel Near-infrared Excited Fluorescent Probe For Bioimaging Of Furin And Hypochlorous Acid

The real-time detection and monitoring of biomarkers generated during the disease process facilitates us to explore its internal mechanisms and will greatly contribute to the prevention,diagnosis and treatment of diseases and cancers.As a result of high sensitivity,specificity,fast response,real-time and in-situ imaging of analytes,fluorescence imaging technology plays a pivotal role in the fields of environmental monitoring,food analysis and disease diagnosis.However,the traditional small-molecule fluorescent probes are equipped with a short-term excitation wavelength,resulting in a series of shortcomings such as poor light stability,biological background fluorescence,and low penetration depth of sample tissue,which greatly limit their application in biological imaging.Therefore,it is urgent to design and develop long-wavelength excited small-molecule fluorescent probes.Based on the excellent characteristics of two-photon and near-infrared fluorescence imaging technology,a series of small-molecule fluorescent probes with novel structure and excellent properties were constructed.They have been successfully applied to real-time monitoring and imaging of active small molecules,proteases and microenvironments in the physiology and pathology processes.The details are as follows:(1)In Chapter 2,We designed and synthesized a turn-on two-photon fluorescent probe for intracellular furin detection and hypoxia imaging.Furin is one of the most characteristic enzymes in the family of proprotein convertases(PCs),which plays a vital role in the activation of biologically active substances.Similarly,the incidence and the severity of cancer are positively correlated with the expression of furin.A self-eliminating linker(4-methylaminopiperidine)was used to achieve the integration of two-photon fluorophore-naphthalimide and the furin-specific substrate sequence(RVRR).Then the two-photon fluorescent probe Nap-F was constructed,which was specific for furin.Based on the change in the charge distribution of this probe before and after the action of furin,Nap-F exhibits a turn-on fluorescence signal.In addition,since the expression level of furin protease is closely related to the oxygen content of the intracellular physiological environment,Nap-F realized the imaging and detection of cellular hypoxia successfully.(2)Due to the excitation or emission wavelengths of most existing furin fluorescent probes were in the visible light range,the analysis of furin in living animal and studying the mechanism of furin in the disease were hindered.Accordingly,in Chapter 3,we designed and synthesized a near-infrared fluorescent probe HD-F for the detection of furin activity.The probe showed excellently spe cific and sensitive to furin andcould detect furin in various cell lysates successfully.At the same time,it was able to reveal the difference of furin expression in various lysates.More importantly,HD-F visualized the divergence of furin expression in various cells and clearly distinguish between normal cells and cancer cells.In addition,HD-F has also been successfully applied to real-time imaging and dynamic monitoring of furin in tumor tissues and in vivo.(3)Hypochlorous acid(HClO),a class of classical reactive oxygen species(ROS),plays a significant role in various life activities.Existing HClO fluorescent probes suffered the interference of other ROS in the detection process,especially ONOO~-.In order to solve this problem,in this chapter,we designed and synthesized a novel near-infrared fluorescent probe Lyso-SiR-2S with lysosomal targeting by integrating the dithiobenzoic acid lactone group into the silicon rhodamine B(Si-Rh B).Compared with the traditional(thio)carboxyl group,the probe was more reactive and selective to HClO,and avoided the interference of ONOO~-in the detection process of HClO.In addition,Lyso-SiR-2S achieved the detection of exogenous and endogenous HClO in the cells successfully and showed well lysosome-targeting.The design and development of this probe provides an effective tool for clinical testing of HClO.