Integrated drug-in-cell therapy of in-stent restenosis

 描述(由申请人提供)：由于缺乏有效和安全相结合的治疗方法，通过血管成形术治疗的动脉损伤引发的再狭窄(再狭窄)以缓解动脉粥样硬化的阻塞仍然是一个挑战。旨在加速血管愈合和内皮再生的策略有可能预防再狭窄，同时避免目前使用的治疗方法的副作用。然而，为了实现这一潜力，需要显著提高它们的效率。这项建议涉及一种综合的细胞内药物抗再狭窄策略，该策略基于内皮细胞与载药磁性纳米载体的双重功能化，以提供支架靶向输送能力和增强的抗再狭窄效果。后者是通过诱导和激活内皮型一氧化氮合酶和以其前体苯甲酸雌二醇酯(E2Bz)的形式形成的磁性纳米颗粒(MNP)中的雌二醇来实现的。本研究将在复制人类疾病主要特征的动脉粥样硬化大鼠模型中建立使用该方法预防支架内再狭窄和促进再内皮化的可行性。具体目标如下：目标1：细胞内药物方法在培养和共培养研究中的评价和优化。将制备E2Bz负载的生物可降解MNP，并对其尺寸、组成、磁性和药物释放特性进行表征。MNP的细胞相容性、MNP的内化和降解动力学，以及E2Bz诱导MNP功能化内皮细胞合成一氧化氮及其对共培养的平滑肌细胞的生长抑制作用将是Aim 1实验的主要终点。目的2：靶向性、再内皮化和疗效研究。在大鼠动脉粥样硬化/颈动脉支架模型中，将研究磁靶向输送功能化细胞后的动脉定位、组织分布和内皮恢复。广度 再狭窄的发生率将在产后两周通过计算机形态计量学确定，细胞功能化的药理作用将以负载空白MNP的细胞作为对照。

 DESCRIPTION (provided by applicant): Injury-triggered renarrowing (restenosis) of arteries treated with angioplasty to relieve atherosclerotic obstruction remains a challenge due to a lack of therapies combining adequate efficacy and safety. Strategies aimed at accelerated vascular healing and endothelium regeneration have potential to prevent restenosis while avoiding the side effects of the currently used therapies. However, in order to realize this potential a significant improvement in their efficiency is required. This proposal is concerned with an integrated drug-in-cell antirestenotic strategy based on dual functionalization of endothelial cell with drug-loaded magnetic nanocarriers to provide them with capacity for stent-targeted delivery and enhanced antirestenotic effectiveness. The latter is achieved by inducing and activating endothelial nitric oxide synthase with estradiol formulated in magnetic nanoparticles (MNP) in the form of its precursor, estradiol benzoate (E2Bz). The present studies will establish feasibilit of using this approach for preventing in-stent restenosis and achieving facilitated reendothelialization in a validated rat model of atherosclerosis reproducing key features of human disease .The following specific aims will be addressed: AIM 1: Drug-in-cell approach evaluation and optimization in culture and co-culture studies. E2Bz-loaded biodegradable MNP will be formulated, and their size, composition, magnetic properties and drug release profile will be characterized. Cell compatibility, internalization and degradation kinetics of MNP, as well as E2Bz-induced nitric oxide synthesis by MNP-functionalized endothelial cells and its growth inhibitory effect on co-cultured smooth muscle cells will be the main endpoints of the Aim 1 experiments. AIM 2: Targeting, reendothelialization and efficacy studies. Arterial localization, tissue distribution and endothelium recovery after magnetically targeted delivery of functionalized cells will be studied in the rat atherosclerosis/ carotid stenting model. The extent of restenosis will be determined two weeks post-delivery by computerized morphometry, and the pharmacological effect of cell functionalization will be delineated using animals treated with cells loaded with blank MNP as controls.