Shape Specific, Enzyme-Responsive, Nano-Imprinted Particles for Drug Delivery

描述（由申请人提供）：纳米技术是开发先进药物递送载体的最有前途的途径之一。经典地，该领域集中于基于自组装或基于乳液的概念合成纳米载体。尽管聚合物或脂质体药物递送系统已经取得了显著进展，但仍然存在一些关键的基本限制。这些包括不能（a）精确控制纳米载体的形状、纵横比、尺寸和多分散性，和（B）将多种疾病特异性触发释放机制整合到纳米颗粒设计中。我们的目标是使用自上而下，高通量的纳米纤维技术，特别是一个修改的步骤和闪光压印光刻（S-FIL）的方法，合成高度单分散的聚合物纳米载体的各种形状，尺寸和纵横比。通过将疾病响应元件（例如肽）直接掺入颗粒基质中，我们提出将酶响应释放性质赋予这些纳米载体，使得药物或造影剂主要响应于肿瘤相关信号而释放。这两年的具体目标是：目标1：开发一种高通量，自上而下的纳米制造工艺，用于制造精确尺寸和几何形状（形状或纵横比）的肿瘤靶向，酶反应纳米载体。在这个目标中，修改的步骤和闪光压印光刻（S-FIL）技术将被用来生产各种尺寸，横截面形状和纵横比的纳米尺寸的颗粒。用于颗粒合成的基本材料将是聚乙二醇二丙烯酸酯（PEGDA）和丙烯酸化五肽GFLGK（肽-DA），其对溶酶体半胱氨酸蛋白酶（例如组织蛋白酶B）敏感。组织蛋白酶B在包括非小细胞肺癌（NSCLC）在内的多种癌症中高度过表达。目标二：评价尺寸、形状、纵横比和大分子单体浓度对（a）印迹纳米载体的体外细胞摄取和（B）模型药物和造影剂从酶响应性纳米颗粒的细胞内释放和转染的影响。我们推测，颗粒的形状，长宽比以及纳米尺寸应显着影响颗粒内化的效率。此外，将使用成纤维细胞和肺癌细胞研究包封药物的细胞内递送以及模型SiRNA药物的转染功效。目的3：研究不同大小、形状和长径比的纳米印迹颗粒在动物体内的生物分布。在研究各种横截面形状和纵横比的纳米颗粒中的一个主要假设是，这些几何参数可能对血管中的颗粒运输性质具有强烈影响，因此应该对颗粒生物分布以及肿瘤积聚功效具有显著影响。总的来说，这些结果应提供一个强有力的基础，为未来的研究提供治疗和诊断剂在动物模型中的各种癌症。这个为期两年的探索性项目的目标是开发一种用于制造智能药物递送纳米颗粒的纳米压印光刻方法。这是一种独特的自上而下的纳米纤维颗粒合成方法。这些纳米颗粒被设计成具有特定的尺寸和形状，可以影响它们的生物分布。此外，颗粒将能够靶向肿瘤细胞并主要响应于疾病特异性信号（例如酶上调）将货物（药物或成像剂）递送至肿瘤。

DESCRIPTION (provided by applicant): Nanotechnology is one of the most promising avenues for the development of advanced drug delivery vehicles. Classically, the field has focused on synthesizing nanocarriers based on self assembly or emulsion based concepts. Although significant progress has been made in polymeric or liposomal drug delivery systems, there remain some key fundamental limitations. These include inability to (a) precisely control shape, aspect ratios, size and polydispersity of the nanocarriers and (b) integrate a variety of disease-specific triggered release mechanisms into the nanoparticle design. Our objective is to use top-down, high throughput nanofabrication technology, specifically a modified step and flash imprint lithography (S-FIL) method, to synthesize highly monodisperse polymer nanocarriers of various shapes, sizes and aspect ratios. By incorporating disease-responsive elements (e.g. peptides) directly into the particle matrix we propose to impart enzyme-responsive release properties into these nanocarriers such that drugs or contrast agents are released primarily in response to tumor-associated signals. The specific aims for this two year period are: Aim 1: To develop a high throughput, top-down nano manufacturing process for fabrication of tumor-targeted, enzyme responsive nanocarriers of precise size and geometry (shape or aspect ratio). In this aim, a modified Step and Flash Imprint Lithography (S-FIL) technique will be employed to produce nanometer size particles of various sizes, cross-sectional shapes, and aspect ratios. The basic material for particle synthesis will be polyethylene glycol diacrylates (PEGDA) and the acrylated penta peptide GFLGK (peptide-DA), which is sensitive to lysosomal cysteine proteases (e.g. Cathepsin B). Cathepsin B is highly over expressed in a variety of cancers including non-small cell lung cancer (NSCLC). Aim 2: To evaluate the effects of size, shape, aspect ratios and macromer concentration on (a) in-vitro cellular uptake of imprinted nanocarriers and (b) intracellular release and transfection of model drugs and contrast agents from enzyme-responsive nanoparticles. We hypothesize that particle shapes, aspect ratio as well as nanoscale dimensions should significantly influence the efficiency of particle internalization. In addition, intracellular delivery of the encapsulated drugs as well as transfection efficacy of a model SiRNA drug will be studied using both fibroblast and lung cancer cells. Aim 3: To study bio- distribution of nanoimprinted particles of various size, shapes and aspect ratios in naove as well as tumor bearing animals. A major hypothesis in studying nanoparticles of various cross sectional shapes and aspect ratios is that these geometric parameters might have strong influence on particle transport properties in blood vessels and therefore should have significant impact on particle bio-distribution as well as tumor accumulation efficacy. Collectively these results should provide a strong basis for future studies on the delivery of therapeutic and diagnostic agents in animal models of various cancers. The goal of this two year exploratory project is to develop a nanoimprint lithography method for fabrication of smart drug delivery nanoparticles. This is a unique top down nanofabrication method for particle synthesis. These nanoparticles are designed to have specific size and shape that can influence their bio distribution. In addition the particles would be able to target tumor cells and deliver the cargo (drugs or imaging agents) to the tumor primarily in response to a disease-specific signal, e.g. enzyme up regulation.