A high-throughput Galectin-9 imaging assay for quantifying nanoparticle uptake, endosomal escape and functional RNA delivery
Journal article, 2021

RNA-based therapies have great potential to treat many undruggable human diseases. However, their efficacy, in particular for mRNA, remains hampered by poor cellular delivery and limited endosomal escape. Development and optimisation of delivery vectors, such as lipid nanoparticles (LNPs), are impeded by limited screening methods to probe the intracellular processing of LNPs in sufficient detail. We have developed a high-throughput imaging-based endosomal escape assay utilising a Galectin-9 reporter and fluorescently labelled mRNA to probe correlations between nanoparticle-mediated uptake, endosomal escape frequency, and mRNA translation. Furthermore, this assay has been integrated within a screening platform for optimisation of lipid nanoparticle formulations. We show that Galectin-9 recruitment is a robust, quantitative reporter of endosomal escape events induced by different mRNA delivery nanoparticles and small molecules. We identify nanoparticles with superior escape properties and demonstrate cell line variances in endosomal escape response, highlighting the need for fine-tuning of delivery formulations for specific applications.


Michael J. Munson

AstraZeneca AB

Gwen O’Driscoll

AstraZeneca AB

Andreia M. Silva

AstraZeneca AB

Elisa Lázaro-Ibáñez

AstraZeneca AB

Audrey Gallud

Chalmers, Biology and Biological Engineering, Chemical Biology

John T. Wilson

Vanderbilt University

Anna Collén

AstraZeneca AB

Elin Esbjörner Winters

Chalmers, Biology and Biological Engineering, Chemical Biology

Alan Sabirsh

AstraZeneca AB

Communications Biology

23993642 (eISSN)

Vol. 4 1 211

Subject Categories

Cell Biology

Pharmacology and Toxicology

Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)





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