Bioengineering of the blood-brain barrier permeability

血脑屏障通透性的生物工程

• 批准号: 6972939
• 负责人: SERGUEI V VINOGRADOV
• 金额: $ 22.71万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6972939
• 关键词: 6 thioguanine; P glycoprotein; RNA interference; biological models; biotechnology; blood brain barrier; brain cell; cerebrovascular system; crosslink; drug delivery systems; gene delivery system; gene induction /repression; laboratory mouse; luciferin monooxygenase; multidrug resistance; neuropeptides; neuropharmacology; pharmacokinetics; plasmids; polyethylene glycols; small interfering RNA; tissue /cell culture; transfection /expression vector; vascular endothelium; zidovudine

DESCRIPTION (provided by applicant): Permeability of various drugs across the blood-brain barrier (BBB) is significantly dependent on the expression and functional activity of specific efflux transporters located in the membrane of brain capillary endothelial cells (BCEC). Selective transient downregulation of the transporters will lead to the application of more effective and less toxic doses of therapeutic drugs against brain tumors or viral infections in CNS. Previously, antisense inhibitors have been shown to temporarily arrest the synthesis of major multidrug resistance agent, membrane P-glycoprotein, and promote reversal of the resistant cell phenotype. The more effective RNA interference mechanism has been recently discovered for selective switching off expression of various genes. Short hairpin RNA (siRNA) could be introduced into target cells through a plasmid DNA precursor using methods of non-viral gene therapy. However, targeted delivery of the pDNA to the cells of the BBB requires a good systemic carrier and selective vectors that bind to the BCEC. As such a carrier, polymer crosslinked Nanogel particles modified with the brain-specific homing peptides (BSHP) have been chosen for tranfection of the BBB by shRNA-encoding plasmid DNA with an ultimate goal suppressing the specific membrane proteins, drug efflux transporters, in the BBB. Specific BSHPs to be attached to the surface of the Nanogel and target delivery to the BBB have been selected in vivo from a vast amount of peptides in the phage display library. Nanogel is non-toxic and highly effective as a transfection agent in many cell lines and evidently, one of the carriers with great potential for systemic administration. The vectorized RNA Interference-Producing system (RIP system) could be used for bioengineering of the BBB permeability for therapeutic agents whose brain accessibility was hampered by specific drug efflux transporters. The central hypothesis of the proposal is that transient suppression of selected drug efflux transporters in the BBB via systemic transfection of brain endothelium using targeted RNAi-producing systems can result in significant enhancement of drug transport to the brain during chemotherapy of the CNS-related diseases. Our Specific aim 1 is to develop the BCEC-targeted Nanogel carriers for systemic delivery of plasmid DNA to the BBB. Specific aim 2 is the enhance transfection efficacy of the BCEC-targeted Nanogel carriers in vitro and in vivo. Specific aim 3 is to suppress selected drug efflux transporters in the BBB in vivo and temporary increase drug transport into the brain. In this Aim brain transport of several representive nucleoside analogue drugs will be assessed in animal model following the transient downregulation of drug efflux transporters in the BBB by the Nanogel-based RNAi-producing systems.

描述（由申请方提供）：各种药物穿过血脑屏障（BBB）的渗透性显著依赖于位于脑毛细血管内皮细胞（BCEC）膜中的特异性外排转运蛋白的表达和功能活性。 选择性瞬时下调转运蛋白将导致更有效和更低毒性剂量的治疗药物用于治疗脑肿瘤或CNS中的病毒感染。 以前，反义抑制剂已被证明可以暂时阻止主要的多药耐药剂，膜P-糖蛋白的合成，并促进耐药细胞表型的逆转。 最近发现了更有效的RNA干扰机制，用于选择性关闭各种基因的表达。 短发夹RNA（Short hairpin RNA，siRNA）可以通过质粒DNA前体使用非病毒基因治疗的方法引入靶细胞。 然而，将pDNA靶向递送至BBB的细胞需要良好的全身性载体和结合BCEC的选择性载体。 作为这样的载体，已选择用脑特异性归巢肽（BSHP）修饰的聚合物交联纳米凝胶颗粒用于通过编码shRNA的质粒DNA转染BBB，最终目标是抑制BBB中的特异性膜蛋白，药物外排转运蛋白。 已经从噬菌体展示文库中的大量肽中在体内选择了要附着到纳米凝胶表面并靶向递送到BBB的特异性BSHP。 纳米凝胶作为转染剂在许多细胞系中是无毒且高效的，并且显然是具有全身给药的巨大潜力的载体之一。 载体化的RNA干扰产生系统（RIP系统）可用于生物工程的血脑屏障通透性的治疗药物，其脑可及性受到特定的药物外排转运蛋白。 该提案的中心假设是，通过使用靶向RNAi产生系统全身转染脑内皮，在BBB中瞬时抑制选定的药物外排转运蛋白，可导致CNS相关疾病化疗期间药物向脑转运的显著增强。 我们的具体目标1是开发靶向BCEC的纳米凝胶载体，用于将质粒DNA系统性递送至BBB。 具体目标2是增强靶向BCEC的纳米凝胶载体在体外和体内的转染功效。 具体目标3是抑制体内BBB中选定的药物外排转运蛋白，并暂时增加药物转运到大脑中。 在这个目的中，几个代表性的核苷类似物药物的脑转运将在动物模型中进行评估，随后通过基于纳米凝胶的RNAi产生系统瞬时下调血脑屏障中的药物外排转运蛋白。