IN VIVO CELL TARGETING / IMAGING WITH NEAR-IR-EMITTING DOPED ZNSE QUANTUM DOTS

使用近红外发射掺杂锌硒量子点进行体内细胞靶向/成像

• 批准号: 8363215
• 负责人: KAM W LEONG
• 金额: $ 0.61万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8363215
• 关键词: Adverse effects; Animals; Antibodies; Binding; Biocompatible; Biodistribution; Biological; Biopolymers; C57BL/6 Mouse; Cadmium; Cell Survival; Cells; Clinical Trials; Diagnostic Neoplasm Staging; Disease; Drug Kinetics; Encapsulated; Environment; Exhibits; Fluorescence; Funding; Glutathione; Grant; Heparin; Human; Image; Implant; In Vitro; Infection; Injection of therapeutic agent; Investigation; Label; Lead; Ligands; Location; Malignant Neoplasms; Malignant neoplasm of prostate; Mesenchymal Stem Cells; Microscopy; Monitor; Mus; NOD/SCID mouse; National Center for Research Resources; Noise; Nude Mice; Optics; Oral; Organ; Photobleaching; Principal Investigator; Process; Property; Proteins; Quantum Dots; Rattus; Regenerative Medicine; Research; Research Infrastructure; Resistance; Resources; Route; Series; Signal Transduction; Signaling Molecule; Source; Sprague-Dawley Rats; Staging; Stem cells; Surface; Synthesis Chemistry; System; Tail; Techniques; Testing; Therapeutic Agents; Therapeutic Effect; Time; Tissues; Toxic effect; Tumor Necrosis Factor-alpha; Tumor Tissue; Uncertainty; United States National Institutes of Health; Veins; Water; amino group; aqueous; base; biological systems; cancer diagnosis; cost; fluorescence imaging; fluorophore; in vivo; intravenous injection; lead selenide; lymph nodes; mast cell; migration; nanoparticle; novel; particle; pre-clinical; research study; stem cell fate; success; trafficking; transmission process; tumor

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Quantum dots (QDs) are appealing as in vivo fluorophores in a variety of biological investigation due to their unique optical properties such as strong fluorescence signal, high resistance to photobleaching, and broad excitation spectra that allows multiplexing application with a single excitation source. To date, the most widely studied QDs that are readily available from commercial sources are CdSe, CdTe and PbSe. However, the intrinsic toxicity of cadmium and lead sheds a doubt on its applicability for long term in vivo studies. In view of this issue, we have developed a novel green chemistry synthetic route to produce a series of cadmium and lead free QDs in aqueous environment. The as-synthesized QDs that are stabilized with glutathione (GSH) exhibit low toxicity as demonstrated by in vitro cell viability test. In addition, the emission of these QDs are tuanable in the red to near-infrared range (650-800 nm), that coincide with the biological window of transmission to offer high signal-to-noise for fluorescence imaging of cells and small animals. Above all, these highly water-soluble, biocompatible QDs are readily amendable to interfacing with biological systems through biomolecules conjugated with the carboxyl/amino groups on GSH or encapsulated in biopolymers. In the proposed study, we will explore the applications of the as-synthesized QDs for long-term in vivo imaging, tracking and targeting. We will do so within the framework of the following specific aims: Aim 1: To study the short-term in vivo biodistribution, clearance, and potential toxicity of the as-synthesized QDs/QDs encapsulated in biopolymers/QDs conjugated with targating ligands over a period of 1 week. Biodistribution dynamics of QDs is a vital aspect for their specific effects on target tissue as well as to identify any undesired side effects after systemic application. In this study, QDs will be injected into the tail vein of nude mice and imaged at various time points post-injection using a fluorescent imaging system. The mice will be sacrificed towards the end of the study and their major organs will be surgically exposed and imaged. Aim 2: To monitor the route of trafficking of mast cell-derived particles to draining lymph nodes (DLNs). During infection, signalling molecules could traverse significant distances to reach the DLNs. However, it is unclear how these molecules could go through the route without dilution or degradation. Mast cells, upon activation, have shown to release stable heparin-based particles containing tumor necrosis factor and other proteins. We propose to label these MC partlces with our QDs, inject into the footpads of MC deficient KitW-sh/W-sh mice / C57BL/6 mice and visualize the trafficking process using the fluorescence imaging system for 2 h. The mice will be sacrificed at the end of the experiment and histological anaylsis will be done on their major organs. (This study is also applicable for monitoring the of QDs labeled nanoparticles introduced into Sprague Dawley rat through oral route or intravenous injection. For this case, the rat will be monitored for 1 week using the fluorescence imaging system, sacrificed at the end of the study and their major organs will be surgically exposed and imaged). Aim 3: To perform long-term in vivo tracking of the migration/differentiation of QDs-labeled stem cells. One of the potential applications of stem cells is for regenerative medicine; however for pre-clinical and clinical trials, it is important to have a noninvasive technique to evaluate the therapeutic effect and location of the implanted stem cells to rule out potential side effects. For this experoment we will conjugate specific antibodies to QDs that will target the surface markers on human mesenchymal stem cells (hMSCs). The QD-labeled hMSCs will be injected intravenously in NOD/SCID mice and the mice will be monitored using the fluorescent imaging system over a period of 8 weeks to access the fate of the stem cells. Aim 4: To develop QDs as highly sensitive probes for in vivo cancer diagnosis. The success of cancer diagnosis is related to the stage at which the malignancy is detected. However, at present there are very few sensitive tests that could detect early-stage cancers. To further illustrate the vast applications of QDs as in vivo targeting and tracking capabilities, we propose to conduct this study by first conjugate our aqueous QDs with active tumor targeting ligands/therapeutic agents to bind to early-stage deep tissue tumor and to establish real-time monitoring of pharmacokinetics and disease treatment through fluorescence imaging. In vivo targeting/tracking studies would be carried out on nude mice grown with human prostate cancer for a period of 3 months. Towards the end of the study, the mice would be sacrificed and histological examination would be carried out.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的主要研究者可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 量子点 (QD) 作为体内荧光团在各种生物学研究中颇具吸引力，因为它们具有独特的光学特性，例如强荧光信号、高抗光漂白性以及允许使用单一激发源进行多重应用的宽激发光谱。迄今为止，研究最广泛且可从商业来源获得的量子点是 CdSe、CdTe 和 PbSe。然而，镉和铅的内在毒性使其在长期体内研究中的适用性受到质疑。鉴于这个问题，我们开发了一种新颖的绿色化学合成路线，在水环境中生产一系列无镉和铅的量子点。体外细胞活力测试表明，用谷胱甘肽（GSH）稳定的合成量子点表现出低毒性。此外，这些量子点的发射可在红色至近红外范围（650-800 nm）内调节，这与生物传输窗口一致，为细胞和小动物的荧光成像提供高信噪比。最重要的是，这些高度水溶性、生物相容性的量子点很容易通过与谷胱甘肽上的羧基/氨基缀合或封装在生物聚合物中的生物分子与生物系统连接。 在拟议的研究中，我们将探索合成的量子点在长期体内成像、跟踪和靶向方面的应用。我们将在以下具体目标的框架内这样做： 目标 1：研究合成后的量子点/封装在与目标配体缀合的生物聚合物/量子点中的量子点在 1 周内的短期体内生物分布、清除和潜在毒性。量子点的生物分布动力学是其对靶组织产生特定作用以及识别全身应用后任何不良副作用的重要方面。在这项研究中，量子点将被注射到裸鼠的尾静脉中，并使用荧光成像系统在注射后的不同时间点进行成像。研究结束时将处死小鼠，并通过手术暴露其主要器官并进行成像。 目标 2：监测肥大细胞衍生颗粒向引流淋巴结 (DLN) 的运输途径。在感染过程中，信号分子可以穿越很长的距离到达 DLN。然而，目前尚不清楚这些分子如何在不稀释或降解的情况下通过该路线。 肥大细胞在激活后会释放出稳定的肝素颗粒，其中含有肿瘤坏死因子和其他蛋白质。 我们建议用我们的 QD 标记这些 MC 颗粒，注射到 MC 缺陷 KitW-sh/W-sh 小鼠/C57BL/6 小鼠的足垫中，并使用荧光成像系统可视化运输过程 2 小时。实验结束时处死小鼠，对其主要器官进行组织学分析。 （本研究也适用于监测通过口服或静脉注射引入斯普拉格道利大鼠的量子点标记纳米颗粒。对于这种情况，将使用荧光成像系统监测大鼠1周，在研究结束时处死，并通过手术暴露其主要器官并成像）。 目标 3：对量子点标记干细胞的迁移/分化进行长期体内跟踪。干细胞的潜在应用之一是再生医学。然而，对于临床前和临床试验来说，重要的是采用非侵入性技术来评估治疗效果和植入干细胞的位置，以排除潜在的副作用。在本实验中，我们将特定抗体与 QD 结合，以人间充质干细胞 (hMSC) 的表面标记为目标。 QD 标记的 hMSC 将被静脉注射到 NOD/SCID 小鼠中，并使用荧光成像系统在 8 周的时间内对小鼠进行监测，以了解干细胞的命运。 目标 4：开发量子点作为体内癌症诊断的高灵敏度探针。 癌症诊断的成功与否与检测到恶性肿瘤的阶段有关。然而，目前能够检测早期癌症的敏感测试很少。为了进一步说明量子点在体内靶向和跟踪能力方面的广泛应用，我们建议首先将我们的水性量子点与活性肿瘤靶向配体/治疗剂结合以结合早期深部组织肿瘤，并通过荧光成像建立药代动力学和疾病治疗的实时监测来进行这项研究。体内靶向/追踪研究将在人类前列腺癌裸鼠身上进行，为期 3 个月。在研究结束时，将处死小鼠并进行组织学检查。