RRC ID 71948
著者 Munson MJ, O'Driscoll G, Silva AM, Lázaro-Ibáñez E, Gallud A, Wilson JT, Collén A, Esbjörner EK, Sabirsh A.
タイトル A high-throughput Galectin-9 imaging assay for quantifying nanoparticle uptake, endosomal escape and functional RNA delivery.
ジャーナル Commun Biol
Abstract 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.
巻・号 4(1)
ページ 211
公開日 2021-2-16
DOI 10.1038/s42003-021-01728-8
PII 10.1038/s42003-021-01728-8
PMID 33594247
PMC PMC7887203
MeSH Endosomes / metabolism* Galectins / genetics Galectins / metabolism* Gene Transfer Techniques* Genes, Reporter HeLa Cells Hep G2 Cells Humans Lipids / chemistry* Luminescent Proteins / genetics Luminescent Proteins / metabolism Microscopy, Fluorescence Nanoparticles* RNA Transport* RNA, Messenger / genetics RNA, Messenger / metabolism* Recombinant Fusion Proteins / metabolism Time Factors Time-Lapse Imaging
IF 4.165
リソース情報
ヒト・動物細胞 HuH-7(RCB1366)