Noninvasive site-specific measurement of gene expression in deep tissues with secreted reporters

使用分泌报告基因对深层组织中的基因表达进行无创位点特异性测量

• 批准号: 10704050
• 负责人: Jerzy Olgierd Szablowski
• 金额: $ 18.29万
• 依托单位: RICE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-13 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10704050
• 关键词: Address; Animals; Area; Autopsy; Bacterial Proteins; Biochemical; Biological Markers; Biological Models; Blood; Blood Circulation; Blood Tests; Brain; Brain Diseases; Brain Injuries; Brain imaging; Brain region; Cells; Cerebrovascular system; Characteristics; Chemicals; Color; Cytochrome P450; Democracy; Detection; Development; Diffusion; Disease; Engineering; Equipment; Euthanasia; Eye; FOS gene; Ferritin; Focused Ultrasound; Future; Gene Delivery; Gene Expression; Gene Expression Profiling; Genes; Goals; Histology; Human; Image; Imaging Device; Immobilization; Kinetics; Label; Laboratory Research; Light; Luciferases; Magnetic Resonance Imaging; Measurement; Measures; Methods; Molecular; Monitor; Mus; Neurons; Optical Methods; Organ; Organ Size; Patients; Penetration; Physiological; Process; Production; Public Health; Reporter; Reporting; Research; Serum; Signal Transduction; Site; Specificity; Spinal Cord; Structure; Technology; Testing; Thick; Time; Tissues; Toxic effect; Translations; Vascular Permeabilities; Work; blood-brain barrier permeabilization; brain cell; brain size; cell type; cranium; design; gene therapy; immunogenic; improved; in vivo; interstitial; millimeter; new technology; non-invasive monitor; promoter; protein biomarkers; sensor; side effect; therapeutic gene; therapeutic protein; transcytosis; transgene expression; ultrasound

Project Summary Monitoring gene delivery in deep tissues is critical for in vivo studies and translation of gene therapies, but available methods for achieving such measurements have limited capabilities. Ideally, levels of gene expression could be noninvasively measured anywhere in the body of both small and large animals. Ideally, multiple genes could be measured at once, without the need for expensive equipment that would put such a method out of financial reach for academic research groups. However, no such method exists as of now. In this proposal, we will develop technologies for monitoring gene expression that, when fully developed, will be: 1) noninvasive, 2) site-specific, 3) could view multiple types of molecules at once (high-multiplexity of imaging), 4) are not impacted by the depth of penetration, skull thickness, or organ size; 5) will use inexpensive equipment allowing democratized access. To achieve this, we will attempt a drastically different approach – instead of imaging reporters within the deep tissue, we will design reporters that can be easily transported from a known region of the brain into the blood, where they can be easily measured. We will develop two independent methods for achieving this goal. First, will express gene expression reporters that exit the cells into brain interstitial space. Then we will temporarily enhance vascular permeability in selected ~millimeter-sized brain regions to allow for free diffusion of these reporters from the brain into the blood. In the second method, we will develop a new class of gene expression reporters that can cross through an intact brain vasculature to continuously report on gene expression within the transduced brain cells. Spatial precision could be provided for this method using noninvasive gene delivery that genetically labels cells with site-specificity. With each of these methods, we will be able to measure gene expression in a specific brain region with a simple blood draw. Our technologies will be particularly beneficial in the studies of large animal species, where monitoring gene expression in the brain, in most cases, is not feasible without euthanasia. While initially focused on the brain as a test-case with a high need for such methods, the same concepts can be applied to other tissues.

项目摘要 监测深层组织中的基因传递对于体内研究和基因疗法的翻译至关重要，但 实现这种测量的现有方法能力有限。理想情况下，基因表达水平 可以在小动物和大动物体内的任何地方进行非侵入性测量。理想情况下，多个基因 可以一次测量，而不需要昂贵的设备，这将使这种方法 学术研究团体的财务覆盖范围。然而，到目前为止还没有这样的方法。在这项提案中，我们 将开发监测基因表达的技术，当完全开发时，将是：1)非侵入性，2) 特定部位，3)可以一次查看多种类型的分子(成像的高多解性)，4)不受影响 根据穿透深度、头骨厚度或器官大小；5)将使用廉价的设备 民主化接入。为了实现这一点，我们将尝试一种截然不同的方法--而不是成像 在记者的深层组织中，我们将设计出可以方便地从已知区域运送的记者 将大脑注入血液，在那里它们可以很容易地被测量。我们将开发两个独立的方法来 实现这一目标。首先，将基因表达报告退出细胞，进入脑组织间隙。 然后我们将暂时增强选定的~毫米大小脑区的血管通透性，以允许 这些记者从大脑自由扩散到血液中。在第二种方法中，我们将开发一个新类 可以通过完整的脑血管系统连续报告基因的基因表达报告 在转导的脑细胞中表达。可以使用以下方法来提供空间精度 非侵入性的基因传递，用基因标记细胞的位置特异性。使用这些方法中的每一种，我们将 能够通过简单的抽血来测量特定大脑区域的基因表达。我们的技术将 在大型动物物种的研究中特别有用，在那里监测大脑中的基因表达， 在大多数情况下，没有安乐死是不可行的。虽然最初专注于大脑作为测试用例，但具有高 需要这样的方法，同样的概念也可以应用于其他组织。