Assessing vascular permeability and nanobubble extravasation with contrast-enhanced ultrasound imaging

项目摘要 在许多慢性炎性疾病中，血管内皮细胞由于其自身的炎症反应而变得病理性可渗透。 诸如血管生成和生长因子和炎性细胞因子（例如组胺， 缓激肽等）。在癌症中，这一过程可用于通过增强的纳米颗粒向肿瘤递送纳米颗粒。 渗透和保留（EPR）效应。然而，基于纳米颗粒的疗法导致了不一致的 结果病人。这是由于许多因素，其中一个主要因素是异质性肿瘤血管结构 在患者之间和单个肿瘤内。纳米颗粒向肿瘤的运输和进入肿瘤细胞。 薄壁组织由于免疫系统的摄取、无效的边缘化和无效的免疫抑制而变得复杂。 外渗需要指导来告知临床医生对每个患者可能最有效的治疗方法。 有效的指导可以减少保健费用和药物的负面副作用。一个便宜，安全， 一种非侵入性的实时成像方法可能能够对血管渗透性的程度进行分类 一旦得到验证，即可为患者提供个性化的治疗方案。这种工具不仅可以用于 对于肿瘤，但是对于涉及病理可渗透脉管系统的所有疾病。考虑到这一目标， 本申请中提出的研究的目的是致力于开发一种实时方法， 使用基于纳米气泡（NB）的新型对比增强成像评估整个肿瘤的血管通透性 超声（CEUS）。该方法将建立在临床上使用的动态CEUS协议的基础上， 微泡（MB）。直径为100-400 nm的NB已被证明可外渗到肿瘤中 薄壁组织在开发这种方法时使用临床超声将确保最终转化为 患者是安全的，具有成本效益，非侵入性，广泛可及。为了测试这一目标，Aim 1实验 将侧重于基于NB尺寸并与MB进行比较来识别NB动态CEUS动力学。文件还将提出 动力学参数，边缘化，和外渗的NB在流动环境中的存在下，人类整体 血液和透化内皮。这些结果将有助于确定外渗NB的大小， 当应用于目标2中更复杂的体内设置时，内皮渗透性的程度。目标2a将使用 鸡胚绒毛尿囊膜（CAM）模型与控制水平的渗透性，以测试NB动态 体内CEUS参数。在已知渗透率水平的情况下，CAM结果将提供重要信息 用于评估体内渗透性的程度。该方法将应用于不受控制的肿瘤环境 目标2b。这项拟议的研究将产生：1）关于造影剂大小如何影响相互作用的新信息 与红细胞和动态CEUS参数，2）肿瘤相关EPR效应的实时分析， 产生新的生理数据，和3）一种非侵入性的方法，用于确定病理的程度， 体内渗透性。这一建议将提供至关重要的知识血管渗透性和外渗 纳米颗粒的潜力，这是必不可少的，以改善病人的护理。

PROJECT SUMMARY In many chronic inflammatory diseases, vascular endothelial cells become pathologically permeable due to conditions like angiogenesis and production of growth factors and inflammatory cytokines (e.g. histamine, bradykinin, etc.). In cancer, this process can be exploited for delivery of nanoparticles to tumors via the enhanced permeability and retention (EPR) effect. However, nanoparticle-based therapeutics have led to inconsistent results in patients. This is due to many factors, with a main one being heterogeneous tumor vascular architecture both between patients and within a single tumor. Transport of the nanoparticle to the tumor and into the parenchyma is complicated by uptake by the immune system, ineffective margination, and inefficient extravasation. Guidance is needed to inform clinicians on what therapies may be most effective for each patient. Effective guidance could reduce health-care costs and negative side effects of medication. An inexpensive, safe, non-invasive, and real-time imaging method may be capable of categorizing the extent of vascular permeability in tumors and once validated, personalize therapeutic regimens for patients. Such a tool could be used not only for tumors, but for all diseases involving pathologically permeable vasculature. With this goal in mind, the objective of the proposed research in this application is to work toward development of a real-time method for evaluating vascular permeability over the entire tumor using novel nanobubble (NB)-based contrast-enhanced ultrasound (CEUS) in vivo. This method will build upon dynamic CEUS protocols used clinically with microbubbles (MBs). NBs, which are 100-400 nm in diameter, have been shown to extravasate into the tumor parenchyma. The use of clinical ultrasound in developing this method will ensure that eventual translation to patients is safe, cost-effective, non-invasive, and widely accessible. To test this objective, Aim 1 experiments will focus on identifying NB dynamic CEUS kinetics based on NB size and compared to MBs. It will also identify kinetic parameters, margination, and extravasation of NBs in a flow environment in the presence of human whole blood and permeabilized endothelium. These results will help identify the size of extravasated NBs and the degree of endothelial permeability when applied to a more complicated in vivo setup in Aim 2. Aim 2a will use a chick embryo chorioallantoic membrane (CAM) model with controlled levels of permeability to test NB dynamic CEUS parameters in vivo. With known levels of permeability, the CAM results will provide essential information for assessing the extent of permeability in vivo. This method will be applied to an uncontrolled tumor environment in Aim 2b. This proposed research will yield: 1) new information on how contrast agent size affects interaction with red blood cells and dynamic CEUS parameters, 2) a real-time analysis of the tumor-associated EPR effect, yielding new physiological data, and 3) a non-invasive method for determining the extent of pathological permeability in vivo. This proposal will provide crucial knowledge on vascular permeability and extravasation potential of nanoparticles, which is essential to improve patient care.