Proj 1: In Vivo Optical Molecular Imaging with Fluorescence Peptides

项目1：这项研究计划的广泛、长期目标是开发一种集成光学系统 以荧光肽为探针靶向检测癌前病变的分子成像策略 (发育不良的)体内组织。研制了宏观和微观荧光仪器。 作为多肽检测的互补成像方法。这一方法可用于监测 生物标记物在小动物模型中的表达及其对人类患者癌症的早期检测 放映。在本应用中，结肠腺瘤被用作异型增生的模型。具体目标是 1)利用噬菌体展示技术鉴定5个候选多肽，用于对培养细胞进行生物筛选，并 新鲜切除的人结肠粘膜标本，2)开发新的光学成像仪器，使用 双轴共聚焦结构，以可视化与多肽结合相关的组织微结构，以及3) 在形成息肉的基因工程小鼠体内验证与异型增生的优先肽结合 体细胞激活Cre重组酶对远端结肠的影响及常规筛查 结肠镜检查。多肽由于其高度的多样性和小的尺寸而被选为开发的探针 可提供与深层组织渗透的高亲和力结合，以及其低免疫原性和低风险 毒性便于翻译为临床使用。噬菌体展示是一种强大的组合技术， 提供了一种无偏见的方法来识别与发育异常具有亲和力的独特多肽 粘膜AS非常适合适应与体内微环境相关的细微变化。这个 双轴共聚焦结构提供了一种可以实现长时间亚细胞分辨率的仪器 利用离轴荧光采集技术，可实现工作距离远，并克服组织散射的影响。 此外，为了与内窥镜兼容，这种设计可以缩小尺寸。直径5毫米的仪器 包装足够小，可以通过治疗性内窥镜中直径为6 mm的仪器通道。 高速扫描是用微型MEMS(微型机电系统)镜进行的，它收集 以足够的帧速率进行光学切片以生成3D体积图像。候选多肽将是 外用于结肠腺瘤及周围正常粘膜，肉眼可见 荧光成像定位增强的多肽结合区域以用于随后的显微荧光 成像以评估发育不良隐窝微结构内结合多肽的空间分布。 公共卫生：拟议的研究将导致开发新型光学成像探头和 可以在临床前模型中进行评估并在实际应用中转化为临床的工具 用于早期发现中空器官癌症的筛查工具。

PROJECT 1: The broad, long-term objectives of this research plan are to develop an integrated optical molecular imaging strategy that uses fluorescence peptides as probes to target the presence of premalignant (dysplastic) tissue in vivo. Macroscopic and microscopic fluorescence instruments are developed as complementary imaging methods for peptide detection. This methodology can be used for monitoring biomarker expression in small animal models and for the early detection of cancer in human patients undergoing screening. In this application, the colonic adenoma is used as a model for dysplasia. The specific aims are to 1) identify 5 candidate peptides using techniques of phage display for biopanning against cultured cells and freshly excised specimens of human colonic mucosa, 2) develop new optical imaging instrumentation using the dual axes confocal architecture to visualize the tissue microarchitecture relevant to peptide binding, and 3) validate preferential peptide binding to dysplasia in vivo in a genetically engineered mouse that forms polyps in the distal colon by somatic activation of Cre-recombinase and in human subjects undergoing routine screening colonoscopy. Peptides are chosen for development as probes because their high diversity and small size can provide high affinity binding with deep tissue penetration, and their low risk of immunogenicity and toxicity facilitate translation for use in the clinic. Phage display is a powerful combinatorial technique that provides an unbiased approach for identifying unique peptides that exhibit affinity binding to dysplastic mucosa as is well suited to accommodate subtle changes associated with the in vivo microenvironment. The dual axes confocal architecture provides an instrument that can achieve sub-cellular resolution with long working distance and overcomes the effects of tissue scattering using off-axis fluorescence collection. Moreover, this design can be scaled down in size for endoscope compatibility. A 5 mm diameter instrument package is sufficiently small to pass through a 6 mm diameter instrument channel in a therapeutic endoscope. High speed scanning is performed with a tiny MEMS (micro-electro-mechanical systems) mirror that collects optical sections at a sufficient frame rate for generating 3D volumetric images. The candidate peptides will be topically applied to colonic adenomas and surrounding normal appearing mucosa followed by macroscopic fluorescence imaging to localize regions of increased peptide binding for subsequent microscopic fluorescence imaging to assess the spatial distribution of peptide binding within the dysplastic crypt microarchitecture. Public Health: The proposed studies will result in the development of novel optical imaging probes and instruments that can be evaluated in pre-clinical models as well as translated to the clinic as practical screening tool for the early detection of cancer in hollow organs.