The rodent eye as a non-invasive window for understanding cancer nanotherapeutics

 描述(申请人提供)：啮齿类动物的眼睛作为了解癌症纳米疗法的非侵入性窗口。摘要：我们建议使用小鼠的眼睛作为非手术窗口，使用最先进的眼成像仪“EyePod”对视网膜下异种移植模型和纳米传输进行高效的光学研究。EyePod使用单细胞分辨率活体共聚焦显微镜和光学相干断层扫描，通过眼睛的自然光学完全非侵入性地执行。这项技术可以在几周甚至几个月内实现可重复的活体成像，定量跟踪肿瘤的发展和治疗纳米颗粒的输送，以及测量肿瘤和组织的反应。我们最近报道的新型胶束基纳米卟啉将用于这一项目。这是一种令人兴奋的、用途广泛的基于卟啉/胆酸的热敏剂。 胶束纳米粒子可用于(I)高效包裹疏水性化疗药物或荧光染料，(Ii)通过卟啉的固有荧光对肿瘤进行近红外荧光(NIRF)检测，(Iii)光动力学治疗(PDT)和光热治疗(PTT)，分别通过肿瘤部位高效的自由基和热产生，(Iv)Gd(Iii)负载用于MRI成像，(V)64Cu负载用于PET成像，以及(Vi)方便地将癌症靶向配体连接到胶束表面以进行癌症靶向输送。彻底了解这种纳米载体如何在肿瘤微环境中分布，以及它如何对受控的光刺激做出反应，将使我们能够最大限度地发挥其作为纳米治疗试剂的治疗潜力。我们最近还开发了一种短肽，这种短肽专门被小鼠脑血管内皮细胞摄取。1.建立裸鼠眼内胶质母细胞瘤和乳腺癌移植瘤模型，并利用无创光学技术(EyePod)研究这两种肿瘤模型在几天和几周的纵向细胞分辨率下的发展及其对纳米阿霉素治疗的反应。2.利用肿瘤模型和EyePod研究肿瘤靶向和非靶向纳米卟啉在体内肿瘤微环境中的生物分布和光响应，并利用冷冻电子显微镜在超微结构水平上对纳米递送进行解剖。3.优化脑血管内皮细胞穿透配体，并利用体内EyePod成像和冷冻电子显微镜研究其纳米转运进入视网膜血管，以及跨越与血脑屏障非常相似的血视网膜屏障。假设和影响：我们假设小鼠的眼睛提供了一个独特和方便的窗口来研究：(I)肿瘤的发展，(Ii)纳米药物或显像剂在肿瘤微环境中的生物分布，以及(Iii)肿瘤对纳米疗法的生物学反应。通过将先进的光学技术与最先进的免标记和荧光标记相结合，我们将能够在微米分辨率下实时检查纳米颗粒和封装药物的位置和命运。从这项研究中获得的信息不仅有助于了解纳米颗粒对肿瘤血管的渗透、在肿瘤间质中的分布以及对肿瘤细胞的摄取，而且还将 使我们能够建立可靠的方案，有效地研究肿瘤发展、肿瘤微环境、纳米载体包裹的药物在肿瘤内的生物分布和命运，以及跨BRB的纳米给药。

 DESCRIPTION (provided by applicant): The rodent eye as a non-invasive window for understanding cancer nanotherapeutics Abstract: We propose to use the mouse eye as a non-surgical window for highly efficient, optical investigation of subretinal xenograft models and nanodelivery, using a state-of-the-art ocular imager, "EyePod". The EyePod employs single-cell resolution intravital confocal microscopy and optical coherence tomography, performed completely non-invasively through the natural optics of the eye. This technology enables repeatable in vivo imaging over weeks and even months, quantitative tracking of tumor development and delivery of theranostic nanoparticles, and the measurement of tumor and tissue responses. The novel micellar-based nanoporphyrin that we have recently reported will be used in this project. This exciting and highly versatile theranostic porphyrin/cholic acid-based micellar nanoparticle allows (i) efficient encapsulation of hydrophobic chemotherapeutic drugs or fluorescent dyes, (ii) near- infra red fluorescent (NIRF) detection of the tumor via the intrinsc fluorescence of porphyrins, (iii) photodynamic therapy (PDT) and photothermal therapy (PTT) via efficient free radical and heat generation at the tumor site, respectively, (iv) Gd(III) loadin for MRI imaging, (v) 64Cu loading for PET imaging, and (vi) convenient ligation of cancer-targeting ligands to the surface of the micelle for cancer- specific targeted delivery. Thorough understanding of how this nanocarrier distributes within the tumor microenvironment, and how it responds to controlled optical stimulation will enable us to maximize its therapeutic potential as a nano-theranostic agent. We have also recently developed a short peptide that is specifically taken up by mouse brain vascular endothelial cells. Specific Aims are: 1. To develop intraocular glioblastoma and breast cancer xenograft models in eyes of nude mice, and to use non-invasive optical techniques (the EyePod) to study the development of these tumor models at cellular resolution longitudinally over days and weeks, and their response to treatment with nanodoxorubicin. 2. To use the tumor models and EyePod to study the biodistribution and photo-response of tumor targeting and non-targeting nanoporphyrins within the tumor micro-environment in vivo, and to use cryo-electron microscopy to "dissect" the nanodelivery at the ultrastructural level. 3. To optimize brain vascular endothelial cell-penetrating ligands, and to use in vivo EyePod imaging and cryo-electron microscopy to study their nanodelivery into retinal vasculature, and across the blood retinal barrier, which is very similar to blood brain barrier. Hypothesis & Impact: We hypothesize that the mouse eye provides a unique and convenient window to study: (i) tumor development, (ii) biodistribution of nanoparticle drugs or imaging agents inside the tumor microenvironment, and (iii) biology of tumor responses to the nanotherapeutics. By combining advanced optics with state-of-the art label-free and fluorescence labeling, we will be able to examine at micrometer resolution the location and fate of the nanoparticles and the encapsulated drug in real time over days and weeks. Information gained from this study will not only facilitate understanding of nanoparticle penetration into the tumor vasculature, distribution inside the tumor stroma, and uptake into tumor cells, but will also allow us to establish robust protocols for efficient study of tumor development, tumor microenvironment, in vivo intra-tumoral biodistribution and fate of drugs encapsulated by nanocarrier, and nanodelivery across the BRB.