Incorporation of Intracellular NanoSIMS Tracers to Oligonucleotide Conjugates via Strain Promoted Sydnone-Alkyne
Loujahine Lincy-Bianchi1, Maximilian Häfner1, Cécile Becquart2
1Medicinal Chemistry, Research and Development, Early Cardiovascular, Renal and Metabolism, Biopharmaceuticals R&D, AstraZeneca, SE-431 83 Gothenburg, Sweden.
Abstract:
Extensive efforts have been dedicated to developing cell-specific targeting ligands that can be conjugated to therapeutic cargo, offering a promising yet still challenging strategy to deliver oligonucleotide therapeutics beyond the liver. Indeed, while the cargo and the ligand are crucial, the third component, the linker, is integral but is often overlooked. Here, we present strain-promoted sydnone-alkyne cycloaddition as a versatile linker chemistry for oligonucleotide synthesis, expanding the choices for bioconjugation of therapeutics while enabling subcellular detection of the linker and payload using nanoscale secondary ion mass spectrometry (NanoSIMS) imaging. This strategy was successfully applied to peptide and lipid ligands and profiled using the well characterized N-acetylgalactosamine (GalNAc) targeting ligand. The linker did not affect the expected activity of the conjugate and was detectable and distinguishable from the labeled cargo. Finally, this work not only offers a practical bioconjugation method but also enables the assessment of the linker's subcellular behavior, facilitating NanoSIMS imaging to monitor the three key components of therapeutic conjugates.
Insights
This study introduces a new linker chemistry for oligonucleotide therapeutics, enabling better delivery and detection. The novel method improves bioconjugation and allows subcellular tracking of therapeutic components.
Area of Science:
- Bioconjugation Chemistry
- Oligonucleotide Therapeutics
- Nanoscale Imaging
Background:
- Targeted delivery of oligonucleotide therapeutics beyond the liver remains a significant challenge.
- Linkers are critical components of therapeutic conjugates but are often overlooked in development.
- Existing bioconjugation methods may limit therapeutic options and detection capabilities.
Purpose of the Study:
- To develop and validate a versatile linker chemistry for oligonucleotide synthesis and bioconjugation.
- To enable subcellular detection and monitoring of therapeutic conjugate components using nanoscale secondary ion mass spectrometry (NanoSIMS).
- To assess the impact of the linker on conjugate activity and its own subcellular behavior.
Main Methods:
- Strain-promoted sydnone-alkyne cycloaddition was employed as the linker chemistry for oligonucleotide synthesis.
- The developed linker was conjugated to peptide and lipid ligands, including the N-acetylgalactosamine (GalNAc) targeting ligand.
- Nanoscale secondary ion mass spectrometry (NanoSIMS) imaging was utilized for subcellular detection and analysis of the linker and payload.
Main Results:
- The strain-promoted sydnone-alkyne cycloaddition proved to be a versatile linker chemistry for therapeutic conjugates.
- The linker did not impede the expected activity of the conjugated oligonucleotide therapeutics.
- NanoSIMS imaging successfully detected and distinguished the labeled linker from the therapeutic cargo within cells.
Conclusions:
- Strain-promoted sydnone-alkyne cycloaddition offers a practical and effective bioconjugation method for oligonucleotide therapeutics.
- This approach facilitates the assessment of linker behavior at the subcellular level, aiding in the optimization of targeted drug delivery.
- The study provides a valuable tool for monitoring all three key components—ligand, linker, and cargo—of therapeutic conjugates using NanoSIMS imaging.
