Development of selective probes to highlight cellular organelles of human cell and tissue samples in X-ray fluorescence imaging

在医学研究领域，X射线荧光成像(XRFI)正在成为一种强大的分析技术，它允许同时检测包括人体细胞和组织切片在内的各种样本中几乎所有比铝重的元素。经过过去几十年的一系列技术进步，现代XRFI现在提供了足够的分辨率，可以从生物样本生成2D和3D元素分布图，使研究人员能够在单个分析步骤中以亚细胞精度定位和量化几乎所有与医学相关的金属和非金属。不幸的是，XRFI提供的这些前所未有的机会迄今因严重缺乏可用于突出相关生物结构(例如线粒体、溶酶体)的探针而受到破坏。因此，研究人员目前被迫对他们的生物样本进行额外的有害分析步骤，这些步骤既耗时又容易产生不需要的人工制品，最终限制了收集数据的可靠性，并减缓了研究进展。为了解决这些缺陷，并使XRFI更适用于医学研究和药物开发目的，我在此建议开发一套迫切需要的、易于使用的XRFI探针，使研究人员能够直接解释生成的XRFI数据。这些新颖的XRFI探针将由具有惰性Ir、Ru或Re核的荧光、XRFI活性、生物正交过渡金属络合物构建。荧光报告单元将通过一个强大的连接器连接到基于识别的靶向部分，该部分将探针的定位指向选定的细胞细胞器。这种模块化设计将实现一种通用的混合匹配合成方法，以生成分别具有不同双模报告单元和靶向部分的探针阵列，可以通过XRFI和传统的荧光活细胞成像进行追踪。这一策略将促进生成的XRFI探针的验证和优化，并为最终用户提供一系列可用的探针，以适应他们各自项目的各自范围。我在超分子和药物化学方面有丰富的背景，从荧光探针和化学传感器的开发，到新的细胞器靶向潜在化疗药物的设计、合成和生物学评估，我认为自己是开发这些迫切需要的探针的理想人选。此外，波鸿鲁尔大学的梅茨勒-诺尔特小组是促进这一有影响力的研究项目成功完成的完美、互补的主办机构，因为他们在金属络合物合成方面拥有广泛的知识基础，丰富的可用仪器，以及宝贵的外部同步加速器设施来验证最终的XRFI探测器。

In the arena of medicinal research, X-ray fluorescence imaging (XRFI) is emerging as a powerful analytical technique that allows for the simultaneous detection of virtually all elements heavier than aluminium in a wide range of samples including human cells and tissue sections. After a series of technological advances over the past decades, modern XRFI now offers sufficient resolving power to generate 2D and 3D elemental distribution maps from biological samples, providing researchers with the ability to localise and quantify virtually all medicinally relevant metals and non-metals with sub-cellular precision in a single analysis step. Unfortunately, these unprecedented opportunities offered by XRFI are thus far undermined by a severe lack of available probes to highlight relevant biological structures (e.g., mitochondria, lysosomes). Consequently, researchers are currently forced to subject their biological samples to additional deleterious analysis steps that are time consuming and prone to generating undesired artefacts, which ultimately limits the reliability of collected data and slows down research progress.To address these pitfalls and to render XRFI more useful for medicinal research and drug development purposes, I herewith propose the development of a set of urgently required, facile-to-use XRFI probes that will enable researchers to directly interpret generated XRFI data. These novel XRFI probes will be constructed with a fluorescent, XRFI-active, bio-orthogonal transition metal complex featuring an inert iridium-, ruthenium-, or rhenium core. The fluorescent reporter unit will be connected via a robust linker to a recognition-based targeting moiety that directs the probe’s localisation towards a selected cellular organelle.This modular design will enable a versatile mix-and-match synthetic approach to generate an array of probes with different dual-modal reporter units and targeting moieties, respectively, that can be traced by both XRFI and conventional fluorescence live-cell imaging. This strategy will facilitate the validation and optimization of the generated XRFI probes as well as provide end-users with a range of usable probes to suit the respective confines of their individual projects.With my diverse background in supramolecular- and medicinal chemistry, ranging from the development of fluorescent probes and chemosensors to the design, synthesis, and biological evaluation of new, organelle-targeted potential chemotherapeutics, I consider myself the ideal candidate for the development of these urgently required probes. Additionally, the Metzler-Nolte group at the Ruhr-University Bochum is the perfect, complementary host institution to facilitate the successful completion of this impactful research project due to their extensive knowledge base on the synthesis of metal complexes, wealth of available instrumentation, and valuable access to external synchrotron facilities to validate the final XRFI probes.