Preparation And Application Of The Imaging Agent DDNP-SPIONs For Targeting β-amyloid Protein

Alzheimer’s disease (AD) is a neurodegenerative disease of the brain withsymptoms including dementia, cognitive deficits and memory impairment with a highincidence among the old. With population aging, AD is becoming one of the seriousproblems for human health and living quality. The two characteristic neuropathology-logical features in AD are senile plaques (SPs) composed of aggregates of β-Amyloid(Aβ) peptides and intracellular neurofibrillary tangles (NFTs) in the brain4-6. Formationand accumulation of aggregates of Aβ peptides in the brain are critical factorscontributing to the development and progression of AD. Although a battery ofneuropsychological tests are often used in making a clinical diagnosis of AD, definitivediagnosis still relies on pathological evaluation at autopsy. In recent years, with thedevelopment of medical technology and the deepening research for AD pathogenicprinciple, molecular imaging technologies, for example, positron emission tomography(PET), single-photon emission computed tomography (SPECT) imaging and magneticresonance imaging (MRI) are widely used in the early diagnosis of AD, so that thedesign of molecular imaging probes to target β-amyloid as markers of AD becomesscientific research hot spot.In order to research and develop a MRI contrast agent of superparamagnetic ironoxide nanoparticles which can target β-amyloid protein in vivo, we first develop ahydrophobic fluorescent probe,1,1-dicyano-2-[6-(dimethylamino) naphthalen-2-yl]propene carboxyl derivative, which can not only target β-amyloid protein but alsocombine with superparamagnetic iron oxide nanoparticles(SPIONs). In this paper,1,1-dicyano-2-[6-(dimethylamino)naphthalen-2-yl]propene carboxyl derivativefluorescent probe of β-Amyloid was synthesized by acylation in the catalyst DMAPbetween succinic anhydride and1-{6-[(2-hydroxyethyl)(methyl) amino]-2-naphthyl}ethan-1-one. The UV-vis and fluorescence spectra of the intermediate product1-{6-[(2-Hydroxyethyl)(methyl)amino]-2-naphthyl}ethan-1-one,(1-{6-[(2-Hydroxy-ethyl)(methyl)amino]-2-naphthyl}ethylidene)malononitrile and objective product1,1-dicyano-2-[6-(dimethylamino)naphthalen-2-yl]propene carboxyl derivative in differentpolar solvents were investigated and the effects of the solvent polarity on the opticalproperties of the three compounds were studied. In order to know the effect ofDDNP-SPIO nanoparticles as MRI contrast agent, we further measured the relaxation efficacy of DDNP-SPIO nanoparticles by a clinical MRI scanner, and then the bindingcapacity of DDNP-SPIO with Aβ(1~40) was quantitatived by fluorescence method invitro.The following works are completed:①The oleic acid modified SPIO nanoparticles was synthesized by thermaldecomposition method. The structure and properties of oleic acid modified SPIOnanoparticles was comfired by IR, TGA-DTA, TEM, Zetasizer Nano, XRD, VSM andother analytical instruments.②We synthesised DDNP carboxyl derivative which the6-of naphthalene ringwas with the carboxyl activity functional groups. The structure and properties of DDNPcarboxyl derivative was studied by IR, TGA-DTA and1H-NMR.③The UV-vis and fluorescence spectra of the intermediate product1-{6-[(2-Hydroxyethyl)(methyl)amino]-2-naphthyl}ethan-1-one,(1-{6-[(2-Hydroxy-ethyl)(methyl)amino]-2-naphthyl}ethylidene)malononitrile and objective product1,1-dicyano-2-[6-(dimethylamino)naphthalen-2-yl]propene carboxyl derivative indifferent polar solvents were investigated and the effects of the solvent polarity on theoptical properties of the three compounds were studied by UV-visiblespectrophotometry and fluorescence emission spectroscopy. The results of the effcts ofthe solvent polarity have shown that the λmax, λex and λem of the compounds3,4,5were red shift with the solvent polarity increase, according to the Lippert equation, thedifference of dipole moment between the before and after the excitation of thecompound5was7.25D. The change of dipole moment made the charge rearrangementof the solvent molecules, and the energy gap between the ground state and excited statebecomes narrow. The reasons of above explain the red-shifted of the fluorescenceemission spectra with the solvent polarity increase.④The DDNP-SPIO nanoparticles were synthesized by ligand exchange with oleicacid, the structure and properties of DDNP-SPIO nanoparticles were studied by IR,TGA-DTA, TEM, Zetasizer Nano, XRD and VSM, and comparative analysis theproperties of oleic acid and DDNP modified the SPIO nanoparticles.⑤In order to know the effect of DDNP-SPIO nanoparticles as MRI contrast agent,we further measured the relaxation efficacy of DDNP-SPIO nanoparticles by a clinicalMRI scanner, and then the binding capacity of DDNP-SPIO with Aβ(1~40) wasquantitatived by fluorescence method in vitro. The results of the MRI scanning wereshown that as-synthesis nanopaticles possessed excellent MRI sensitivity. DDNP-SPIO nanoparticles were significantly shorter the T2of protons. The DDNP-SPIOnanoparticles which have higher magnetic moment than the common iron oxide powderin the same iron concentration could increase the inhomogeneity of the local magneticfield. The results of the vitro binding experiment displayed that DDNP-SPIOnanpoparticles showed high binding affinities towards Aβ(1-40) aggregates in vitro(Kd=30.9nmol/L) and indicated that DDNP-SPIO nanoparticles can specificallycombinate with Aβ(1~40) fibres,and so that it can specifically recognize Aβ(1~40)fibres. Therefore, DDNP-SPIO nanoparticles can be used as magnetic resonancecontrast for early detection of AD.