The Novel Fluorescent Probes For β-Amyloid Protein And Brain Imaging In Mice

Alzheimer’s disease(AD)is an irreversible,high-incidence and senile dementia-like disease.With the increasing and aging of the population,the incidence of AD increases year by year.AD has a long asymptomatic incubation period and is difficult to diagnose in the early stage of the disease.When the patient shows behavioral abnormalities and structural changes in the brain,the course of the disease has entered the middle and late stages and has become irreversible and uncontrollable.At present,there is no effective drug to cure AD or effectively reverse the disease process.β-Amyloid protein(Aβ)is closely related to the pathogenesis and pathological process of AD.Aβ is an important biomarker of AD,an important cause of neurotoxicity,and also an important pathogenic factor to promote the pathological process of AD.Therefore,Aβ is an important target for the diagnosis and treatment of AD.In situ imaging of brain Aβ in AD model animals is helpful to study the Aβ level changes of Aβ in the pathological process of AD and reveal the interaction between Aβ aggregation and disease progression,which is of great significance to elucidate the pathogenesis of AD and develop effective drugs for the AD treatment.Fluorescence imaging has been widely used to in situ monitor the bioactive substances in cells and living organisms due to its advantages of non-invasive,time-space controllability and simple operation.Among them,near-infrared optical imaging and two-photon fluorescence imaging have been widely used in in-situ imaging of bioactive substances in living animals due to their advantages such as strong tissue penetration,less autofluorescence interference and less light damage.In recent years,a variety of fluorescent probes have been reported for the detection of Aβin biological samples.However,there are still some problems to be solved.(1)The excitation and emission wavelengths of the probes are short,so it is unable to image Aβ in vivo.(2)Most of the probes can only respond to insoluble Aβ(aggregates,fibers and plaques)in the later stage of the disease,but almost have no fluorescence response to soluble Aβ(monomers and oligomers)in the early stage.The design and synthesis of fluorescent probes that can respond to soluble Aβ plays an important role in the early diagnosis of AD and in elucidating the early pathological mechanism of AD.(3)Most probes can only detect Aβ by fluorescence imaging,but can not inhibit Aβmonomer aggregation.The development of novel fluorescent probes that simultaneously image Aβ and inhibit its aggregation will promote the design and development of novel AD theranostic reagents targeting Aβ.To this end,based on biaryl and benzopyranium structures,we designed and synthesized BA and BP series of fluorescent probes.The fluorescence response,binding affinity,aggregation inhibition and biological imaging applications of the probes were investigated.1.Based on the biaryl structure,a series of BA probes BA-1 and BA-2 were designed and synthesized by using intramolecular charge transfer and rotational relaxation of the molecular rotor to regulate the fluorescence changes.The synthesis of the probe is simple,needs only one step reaction,has high yield and is easy to purify.BA-1 and BA-2 showed strong fluorescence response to Aβ and the response was not interfered by other bioactive substances.The binding affinity of the probes to Aβ aggregates were stronger than that of the commercial dye Th T.In addition,the probes were able to respond to various forms of Aβ,including the monomer,aggregate and fiber.In particular,the probes showed a high fluorescence response to Aβ monomer,which has potential application value in the early diagnosis and pathological study of AD.Both probes can be used for fluorescence imaging of Aβ plaque in AD transgenic mice.This study will promote the further design and synthesis of novel Aβ probes,especially fluorescent probes response to Aβ monomer.2.BP-1,BP-2 and BP-3 probes were designed and synthesized based on benzopyronium structure skeleton by controlling hydrophilicity and rotational degree of freedom.The probes showed significant fluorescence responses to different Aβ forms such as the monomer,aggregate and fiber.The probes showed strong binding ability to Aβ aggregates,and their binding affinities are obviously stronger than that of commercial Aβ dye Th T.In addition,BP-2 and BP-3 showed the ability to inhibit the further aggregation of Aβ monomers.This series of probes have the properties of near-infrared fluorescence emission and two-photon excitation,both of which are capable of fluoresce imaging of the Aβ plaques in mouse brain tissue.The probe BP-3 can enter the brain of mice through the blood-brain barrier by intravenous injection.With the help of twophoton confocal microscopy,BP-3 can perform in situ fluorescence imaging of Aβ plaques in the brain of living mice.In addition,a significant difference in brain fluorescence intensity was observed between AD model mice and wild-type mice of the same age by IVIS Imaging System.This study provides a new kind of fluorescent probes capable of in situ imaging Aβ plaques in the brain of living animals and inhibiting the Aβ aggregation,which will promote the subsequent design and synthesis of new AD theranostic reagents targeting Aβ.