Bioorthogonal time-controlled intramitochondrial elimination

Bioorthogonal transformations are chemical reactions that can be carried out in a controlled manner in biological systems. The substances that react with each other must therefore not only be biocompatible and thus not affecting the biological system, but must also react selectively and rapidly under these conditions. In this way, for example, two compounds can be connected inside a living cell and thereby labeled, for example to visualize biomolecules by molecular imaging. In recent years, it has moreover become possible to bioorthogonally cleave compounds, as a result of which substances can be released selectively at certain locations inside a living system or organism. For example, this is being used more and more for the selective release of drugs in cancer cells. Within this project, bond-cleavage reactions are to be developed and investigated, which enable a time-controlled release of compounds inside cells. The aim of this approach is to selectively incorporate a modified substance into the DNA of mitochondria, the cell`s power plants. This substance can subsequently be used to make the mitochondrial DNA (mtDNA) visible enabling investigations of this important biomolecule. The targeted release inside the mitochondria prevents incorporation into the DNA inside the nucleus, which currently makes visualization of the mtDNA considerably more difficult using state-of-the-art procedures.

The research carried out within this project has significantly advanced the field of bioorthogonal chemistry, which allows for precise chemical reactions within living cells. We first developed a new strategy to achieve the efficient release of phenols through bioorthogonal click-to-release mechanisms, successfully demonstrating this by controlling the activation of a caged prodrug. In addition, we created new bioorthogonal tools, specifically a set of tetrazines that act as highly effective chemical scissors. These tools overcome previous limitations by significantly accelerating the click-to-release process, which is crucial for applications requiring rapid response times. The enhanced speed and efficiency of these tetrazines enable more precise control over chemical reactions within living cells, opening new avenues for research and medical applications. Furthermore, we devised innovative methods to synthesize modified trans-cyclooctenes, which serve as click-cleavable linkers, providing versatile tools for various bioorthogonal applications. Our new approaches to incorporating linkers that decompose in a preprogrammed timely manner into bioorthogonally cleavable compounds enable the precise, time-controlled release of payloads upon accelerated click-triggered bond-cleavage. This level of control is essential for applications such as the study of dynamic cellular processes. Moreover, we achieved targeted bioorthogonal click-to-release within mitochondria, allowing for the specific delivery of substances inside these cellular powerhouses. While the project was successfully concluded, ongoing studies are building on our methods. We are currently investigating the selective incorporation of modified substances into mitochondrial DNA (mtDNA) using time-controlled release. This line of research could pave the way for future studies that further explore the role of mtDNA in cellular function and disease. In summary, our findings represent significant progress in bioorthogonal chemistry, providing powerful tools and strategies for time-controlled chemical reactions in living cells. The developed tools and methods not only expand the repertoire of techniques to study cellular processes but also offer practical solutions for several challenges in the fields of bioorthogonal chemistry and chemical biology.