结合电生理学与主动靶向技术的抗癫痫电场敏感纳米水凝胶的研究

Epilepsy is one of the most prevalent neurological disorders. For most patients，antiepileptic drugs are the mainstay of the management of epilepsy. However, the lack of targeting ability of the drug, which results in systemic side effects and drug resistance, makes the complete seizure control failure. Epileptic seizure is the paroxysmal abnormalities synchronization electrical activity in brain neurons. Therefore, constraining the discharge of epilepsy in the local site and suppressing it timely is the key to prevent the seizure. With chitosan as the main material, the peptide-modified electric field responsive nano-hydrogel will be prepared based on electrophysiology together with active targeting technology in this project. With the functionalization of homing peptide, the nano-hydrogel drug delivery system will target to the brain efficiently, and at the beginning of the seizure, the nano-hydrogel will convert the electrical energy into mechanical energy immediately, which weaken the epileptiform discharges and release drug quickly leading to an efficient treatment of epilepsy. Our preliminary experiments demonstrated that the nano-hydrogel could release drug quickly under electric field in vitro, and could target to brain in vivo. Further study will be performed to investigate the mechanism and relationship between anti-epilepsy efficacy and the reduced epileptiform discharges, brain-targeting, triggered drug release of the nano-carrier. Takes advantages of the electric field response, nervous system biocompatibility, and brain-targeting, the nano-hydrogel will be a potential candidate for the epilepsy treatment.

癫痫是神经系统最常见的疾病之一，其药物治疗存在靶向性差，易引起毒副作用和耐药等问题。癫痫的发作是脑内神经元阵发性异常超同步化电活动的结果，将癫痫灶点异常放电限制在局部并及时抑制，是阻止癫痫发作的关键。本项目结合癫痫发作的电生理学机制和主动靶向技术，以壳聚糖为主要材料构建脑靶向电场敏感纳米水凝胶。通过脑靶向肽介导纳米载体靶向累积于脑部，并在癫痫发作之初，纳米载体将癫痫灶点异常放电转化为机械能，在消弱癫痫样放电的同时快速释放药物，实现癫痫的高效治疗。预实验已经发现该纳米载体具有微电场响应性药物释放及靶向肽修饰的脑靶向特性。因此，本项目拟进一步评价该纳米载体的脑靶向特性、癫痫发作放电所触发的药物快速释放和癫痫样放电的消弱与抗癫痫疗效之间的相关性，并初步阐明其作用机制。通过纳米载体的电场响应性、神经系统相容性、体内外靶向性、癫痫大鼠的药物动力学及抗癫痫疗效研究，探讨癫痫治疗的新策略。

癫痫的发作是脑内神经元阵发性异常超同步化电活动的结果，将癫痫灶点异常放电限制在局部并及时抑制，是阻止癫痫发作的关键。本项目结合癫痫发作的电生理学机制和主动靶向技术，构建脑靶向电场响应纳米水凝胶。通过脑靶向肽介导纳米载体靶向累积于脑部，并在癫痫发作之际，在消弱癫痫样放电的同时快速释放药物，实现癫痫的高效治疗。主要研究内容和重要结果如下：.1、电场响应纳米水凝胶的合成与评价。以甲基丙烯酸二甲胺基乙酯、苯乙烯和对苯乙烯磺酸钠为单体，以N,N′-亚甲基双丙烯酰胺为交联剂，通过无皂乳液共聚法合成具有电场响应功能的纳米水凝胶。所合成的纳米水凝胶荷正电、粒径在14~146 nm之间；纳米水凝胶在水性介质中可快速响应电场变化，纳米粒溶胀，粒径变大；以苯妥英钠（PHT）为模型药物，制备负载PHT纳米水凝胶，药物从纳米水凝胶中的释放具有电场响应特性。.2、脑靶向电场响应纳米水凝胶的靶向性研究。利用脑靶向多肽 Angiopep-2对纳米水凝胶进行修饰，制备Angiopep-2修饰纳米水凝胶（ANG-ERHNPs）。ANG-ERHNPs保持了电场快速响应功能； ANG-ERHNPs有较好的跨体外BBB模型转运的能力；ANG-ERHNPs经腹腔注射后，通过活体成像系统可观察到其在大鼠脑组织和脑区累积；苯妥英钠脑靶向电场响应纳米水凝胶（ANG-PHT-ERHNPs）大鼠体内的相对生物利用度为PHT溶液组的1.91倍，明显延长了PHT在体内的循环时间；且具有脑部转运能力，ANG-PHT-ERHNPs脑靶向指数为2.56；在癫痫相关脑区（皮层、海马、杏仁核、小脑和脑干），Cmax提高近1倍。 .3、脑靶向电场响应纳米水凝胶的抗癫痫药效研究。采用最大电休克、杏仁核电点燃和戊四唑三种大鼠癫痫模型，评价腹腔注射给予不同剂量的PHT溶液、ANG-PHT-ERHNPs、苯妥英钠脑靶向纳米水凝胶（ANG-PHT-HNPs）的抗癫痫作用。相比ANG-PHT-HNPs和PHT溶液，ANG-PHT-ERHNPs在三种癫痫模型中均具有更好的抗癫痫作用，不仅能降低有效治疗剂量而且可以延长药物的抗癫痫治疗时间窗；戊四唑诱导的癫痫模型上测得的游离PHT结果进一步证实了PHT从ANG-PHT-ERHNPs中快速释放与发作脑电的严重程度呈正相关。