Albumin hitchhiking siRNAs for gene targeting in aged brain

目前治疗阿尔茨海默病(AD)的一个障碍是在大脑中使用有效剂量的生物药物。AD的最大风险因素是衰老，这与血脑屏障(BBB)的进行性功能障碍相吻合，导致正常情况下被排除在健康大脑之外的血清蛋白进入和滞留。其中一种蛋白质是白蛋白，在年轻的脑组织中没有检测到，但随着年龄的增长逐渐积累在大脑中。在这里，我们提出了一种治疗阿尔茨海默病的新策略，该策略将利用白蛋白作为“天然”载体来增强siRNA介导的老年大脑中的基因靶向。我们假设，将siRNA“搭便车”到白蛋白上将改善静脉给药后老年大脑中siRNA的积累和基因沉默活性，从而提供一种可定制的靶向AD相关基因的策略。这种方法是由我们实验室开发的一种新的二酰基脂肪酸载体(“EG18”)促进的，它可以直接连接到siRNA(“siRNA-EG18”)。我们有相关的初步数据表明：1)与我们先前发表的siRNA-L2载体相比，siRNA-EG18对白蛋白的亲和力和特异性增强，循环半衰期延长； 2)siRNA-EG18在骨性关节炎小鼠模型中持续基因敲除，导致白蛋白在发炎的膝关节积聚；3)siRNA-EG18在静脉注射适量1 mg/kg剂量后24小时在老年小鼠的大脑中积聚，而不是在幼年小鼠的大脑中积聚。这项建议的目标1将以这些结果为基础，通过检查siRNA-EG18的药代动力学、生物分布和毒性随年龄和剂量方案的变化而变化。目的2将检测siRNA-EG18在老年人大脑中的生物活性，重点针对APOE，这是已知的AD最强的遗传风险因素。总之，这项提议将为通过搭乘siRNA的白蛋白在老年脑中实现基因沉默建立工作参数，从而为AD的个性化药物治疗提供新的机会。

A current hurdle for the treatment of Alzheimer’s disease (AD) is administration of efficacious doses of biological drugs in the brain. The strongest risk factor for AD is aging, which coincides with progressive dysfunction of the blood-brain barrier (BBB) leading to entry and retention of serum proteins that are normally excluded from healthy brain. One of these proteins is albumin, which is not detected in young brain tissue but gradually accumulates in the brain with age. Here, we propose to a novel strategy for treating AD that will leverage albumin as a “natural” carrier to enhance siRNA-mediated gene targeting in the aged brain. We hypothesize that “hitchhiking” siRNA onto albumin will improve siRNA accumulation and gene silencing activity in the aged brain after intravenous administration, thereby providing a customizable strategy to target genes associated with AD. This approach is facilitated by a novel diacyl fatty acid carrier developed in our lab (“EG18”) that can be directly conjugated to siRNA (“siRNA-EG18”). We have relevant preliminary data showing that: 1) siRNA-EG18 has enhanced affinity and specificity for albumin and increased circulation half-life relative to our previously published siRNA-L2 carrier; 2) siRNA-EG18 exerts sustained gene knockdown in a mouse model of osteoarthritis that leads to albumin accumulation in the inflamed knee joint; 3) siRNA-EG18 accumulates in the brains of old but not young mice 24 hours after intravenous delivery of a modest 1 mg/kg dose. Aim 1 of this proposal will build on these results by examining the pharmacokinetics, biodistribution, and toxicity of siRNA-EG18 as a function of age and dosing scheme. Aim 2 will examine the bioactivity of siRNA-EG18 in the aged brain, with a focus on targeting APOE, the strongest known genetic risk factor for AD. Collectively, this proposal will establish working parameters for achieving gene silencing in the aged brain via albumin hitchhiking of siRNA, thereby providing new opportunities for personalized medicine in AD.