Nanoparticle Brain Delivery of Iron Chelators for AD

描述(由申请人提供)：强有力的证据表明，铁螯合疗法可以通过消耗体内多余的金属而使阿尔茨海默病(AD)患者受益。遗憾的是，铁络合剂的毒性、给药途径和跨越血脑屏障的能力受限等问题阻碍了这一方法的进一步发展。我们的长期目标是利用纳米药物输送技术开发新型铁络合治疗剂，用于AD的预防和治疗。具体的假设是，与铁络合剂结合的纳米颗粒不仅可以作为载体将螯合剂送入大脑，还可以将铁(和其他一些金属)-螯合剂络合物带出大脑，从而防止与过量金属相关的脑损伤。这一假说是基于：1)纳米颗粒可以模仿选定的脂蛋白颗粒，并通过脂蛋白受体介导的机制进入大脑。2)一些脂蛋白颗粒也可以通过同样的受体介导的机制离开大脑。3)我们的初步研究表明，铁络合剂可以与纳米颗粒偶联，形成的颗粒有可能通过模仿脂蛋白进入大脑。更重要的是，连接到纳米颗粒上的螯合剂保持了金属结合能力，金属-螯合剂-纳米颗粒复合体能够模仿一些可以离开大脑的脂蛋白颗粒。为了验证我们的假设，我们提出了具体的目标。1.合成铁络合剂并将其偶联到纳米粒子上，然后测试形成的体系对金属的结合能力。铜络合剂还将用于偶联和测试，以深入了解不同金属在AD中的作用。我们或根据文献已经开发了合成和偶联的方法。2.对形成的体系和与金属络合的体系的血浆蛋白吸收图谱进行表征。PPAP将通过二维凝胶分析来确定，并表明系统模拟脂蛋白的能力。将使用体外共培养的脑血屏障模型对粒子跨脑血屏障(BBB)的双向输送系统进行动力学研究。3.使用改进后的程序进行体内动力学研究，检查系统进入和离开大脑的能力。分别用层析法、石墨炉原子吸收光谱仪(GFAAS)测定小鼠脑内纳米粒子和金属铁的含量。其他器官、血流和排泄物中的这些含量也将被检查。4.证明该系统是否从阿尔茨海默氏症转基因小鼠和帕金森病大鼠的大脑中去除铁或其他金属，并防止大脑受到氧化损伤。除了大脑，像AIM 3中的其他隔间将被检查纳米颗粒、金属和氧化损伤水平。这些终点将通过生物、组织和免疫化学方法以及石墨炉原子吸收光谱等仪器分析来确定。 本研究不仅将为研究过量金属离子导致阿尔茨海默病的发病机制提供依据，也将为AD的治疗提供潜在的治疗方法。此外，这种方法也可以应用于其他由过量金属介导的神经退行性疾病，以及神经成像。

DESCRIPTION (provided by applicant): Strong evidence has shown that iron chelation therapy can benefit Alzheimer's disease (AD) patients by depleting excess metals from the body. Unfortunately, problems with toxicity, route of administration and restricted ability of the iron chelators to cross the BBB have impeded the further development of this approach. Our long-term goal is to develop novel iron chelation therapeutic agents using nanoparticle drug delivery technology for AD prevention and treatment. The specific hypothesis is that nanoparticles conjugated with iron chelators can serve as vehicles not only to send chelators into the brain but also bring the iron (and some other metals)-chelator complexes out of the brain, hence preventing excess metal associated brain damage. This hypothesis is based on 1) nanoparticles can mimic selected lipoprotein particles and enter the brain via lipoprotein receptor-mediated mechanisms. 2) Some lipoprotein particles can also leave the brain via the same receptor-mediated mechanisms. 3) Our preliminary studies show that iron chelators can conjugate with nanoparticles and the formed particles have the potential to enter the brain by mimicking lipoprotein. More important, the chelators conjugated to nanoparticles retain metal binding ability and the metal-chelator-nanoparticle complexes are capable of mimicking some lipoprotein particles that can leave the brain. To demonstrate our hypothesis, the specific aims are proposed. 1. Synthesize iron chelators and conjugate them to nanoparticles, then test the metal-binding ability of the formed systems. Copper chelators will also be used for conjugation and testing to obtain insights into the roles of different metals in AD. The methods for synthesis and conjugation have already been developed by us or according to literature. 2. Characterize the plasma protein absorption patterns (PPAP) on the formed systems and the systems complexed with metals. The PPAP will be determined by the 2-dimensional gel analysis, and indicate the capabilities of the systems to mimic lipoproteins. Kinetic studies of the particle delivery systems bi-directionally across Brain Blood Barrier (BBB) will be conducted using an in vitro coculture BBB model. 3. Examine the ability of the systems to enter and leave the brain using in vivo kinetic studies with a modified procedure. The contents of nanoparticles and metal irons in mouse brain will be assessed using chromatography, Graphite Furnace Atomic Absorption Spectrometry (GFAAS), respectively. These contents in other organs, bloodstream and excretions will also be examined. 4. Demonstrate whether the systems remove iron or other metals from the brain of Alzheimer transgenic mice and Parkinson's disease rats, and prevent the brain from oxidative damage. In addition to brain, other compartments like in aim 3 will be examined for the nanoparticle, metal and oxidative damage levels. These endpoints will be determined by bio-, histo- and immunochemical methods and instrumental analyses such as GFAAS. This study will not only provide the insights into the mechanisms of AD development associated with excess metal ions, but also provide potential therapeutics. Moreover, this approach may be applied to other neurodegenerative diseases mediated by excess metals, and to neuro-imaging.