Bioorthogonal probe development for highly parallel in vivo imaging

用于高度并行体内成像的生物正交探针开发

基本信息

批准号：
10596786
负责人：
RALPH WEISSLEDER, MD, PHD
金额：
$ 41.58万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-04-21 至 2026-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10596786
关键词：
3-Dimensional Affect Affinity Antibodies Benchmarking Biological Biology Biomedical Research Biotechnology Cancer Biology Cell Mobility Cells Chemistry Color Complex Data Development Drug Delivery Systems Dyes Environment Feasibility Studies Fingerprint Flow Cytometry Fluorochrome Future Goals Gold Image Imaging technology Immune In Vitro Investigational Therapies Kinetics Label Ligands Magnetic Resonance Imaging Malignant Neoplasms Methods Modernization Molecular Profiling Molecular Target Noise Optics Polymers Positron-Emission Tomography Proteins Reaction Reporter Residual state Resolution Science Signal Transduction Stains Stromal Cells Techniques Technology Testing Time Tissues Viola Visualization cancer therapy cell behavior cell motility cellular targeting drug action experimental study imaging agent imaging capabilities improved in vivo in vivo evaluation in vivo imaging insight intravital imaging intravital microscopy microscopic imaging mouse model multiplexed imaging new technology novel real-time images resistance mechanism response small molecule success technology validation transcriptomics tumor tumor heterogeneity tumor microenvironment two-dimensional validation studies

项目摘要

ABSTRACT Multiplexed analytical techniques such as flow cytometry, scRNAseq and spatial transcriptomics have revolutionized modern biology. Unfortunately, there are still no reliable ways of performing highly multiplexed in vivo real-time imaging to understand the functional behavior of cells and molecules in their intact microenvironment over time. Based on advances in “click chemistries” we have developed a novel, scission based in vivo multiplexing technology (SAFE; Ko et al., 2022, Nat Biotech, in press; Ko et al., Adv Science, 2022 in press), that for the first time allows intravital imaging far beyond what is currently possible (~12-20 channel in vivo imaging in feasibility studies instead of 4-6 channels afforded by intravital microscopy). The new technology utilizes specially labeled fluorescent affinity ligands i) where an exogenously administered tetrazine cleaves a C2TCO containing linker between antibody and fluorochromes in seconds and at non-toxic concentrations and ii) allows antibody-tagged superbright polymeric and indocyanine dyes to be washed away between cycles of imaging and thus re-staining. The goal of this proposal is to further develop and validate the technology and then use it to gain deeper temporal and spatial insight into the tumor microenvironment. We propose two specific aims: i) Optimize the SAFE-IVM technology for broader (~30 channel) multiplexed in vivo imaging (new chemistry including novel iTCO, Tz-BHQ, expand colors, improve imaging chambers); and ii) perform multiplexed imaging of tumor microenvironment and its immune cells. Findings from latter studies will be important because they will i) establish the utility of the new technology benchmarked against gold standards and ii) open the door to broader applicability of the new technology.

抽象的流式细胞术、scRNAseq 和空间转录组学等多重分析技术已不幸的是，仍然没有可靠的方法来进行多重分析。体内实时成像，了解细胞和分子完整的功能行为随着时间的推移，基于“点击化学”的进步，我们开发了一种新颖的分裂。基于体内多重技术（SAFE；Ko 等人，2022，Nat Biotech，出版中；Ko 等人，Adv Science， 2022 年出版），首次允许活体成像远远超出目前的可能性（~12-20 可行性研究中的体内成像通道，而不是活体显微镜提供的 4-6 个通道）。新技术利用特殊标记的荧光亲和配体 i) 四嗪可在数秒内裂解抗体和荧光染料之间含有连接体的 C2TCO，并且无毒浓度和 ii) 允许抗体标记的超亮聚合染料和吲哚菁染料被洗掉该提案的目标是进一步开发和验证该技术。技术，然后利用它来更深入地了解肿瘤微环境的时间和空间。提出两个具体目标： i) 优化 SAFE-IVM 技术以实现更广泛（约 30 通道）的体内多路复用成像（新化学物质包括新型 iTCO、Tz-BHQ、扩展颜色、改进成像室）；后期研究的结果将进行肿瘤微环境及其免疫细胞的多重成像。很重要，因为它们将 i) 建立以黄金为基准的新技术的效用标准和 ii) 为新技术的更广泛应用打开了大门。