Endocytic Profiling of Cancer Cell Models Reveals Critical Factors Influencing LNP-Mediated mRNA Delivery and Protein Expression

Sayers, Edward J.; Peel, Samantha; Schantz, Anna; England, R.; Beano, Maya; Bates, Stephanie M.; Desai, Arpan; Puri, Sanyogitta; Ashford, Marianne; Jones, Arwyn Tomos

doi:10.1016/j.ymthe.2019.07.018

Cited by 65 publications

(61 citation statements)

References 47 publications

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“…Interestingly, by integrating the mCherry-Gal9 reporter into multiple cell lines, we can also begin to explore cell to cell differences in response to LNPs. Our study showed that liver-derived cell models demonstrated greater sensitivity to endosomal disruption from MC3-LNPs than non-liver cell models without equivalent differences in the level of particle uptake, providing a more detailed understanding to the observation by Sayers et al that LNPs are differently effective in liver-derived Huh7 compared to lung-derived NCI-H358 45 . Examination of a wider range of LNP formulations and cationic lipids will be important to determine whether this is an intrinsic sensitivity of these cell models to nanoparticle delivery or whether this is indicative of a composition-driven LNP selectivity for certain cell types.…”

Section: Discussionsupporting

confidence: 58%

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

Munson

O’Driscoll

Silva

et al. 2020

Preprint

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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. Advances in the development and rational optimisation of delivery vectors, such as lipid nanoparticles (LNPs), are impeded by the limited availability of screening methods that 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. This assay has, furthermore, been integrated with a screening platform for nanoparticle formulation to optimise LNPs. 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 also identify nanoparticles with superior escape properties and demonstrate significant cell line variances in endosomal escape response, highlighting the need for fine-tuning of delivery formulations for specific applications.

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Section: Discussionsupporting

confidence: 58%

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

Munson

O’Driscoll

Silva

et al. 2020

Preprint

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“…17 Nevertheless, considering only the fusogenic properties of LNPs might not be the best approach, because these vehicles interact with cell membranes which might have ''incompatible'' fusogenic properties. According to Sayers et al 85 one shall investigate the properties of cell membranes when working on the delivery of nucleic acids. Moreover employing more than one helper lipid in LNPs can also improve their properties.…”

Section: Discussionmentioning

confidence: 99%

DOPC versus DOPE as a helper lipid for gene-therapies: molecular dynamics simulations with DLin-MC3-DMA

Ermilova

Swenson

2020

Phys. Chem. Chem. Phys.

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“…Generating more potent (ionizable cationic) lipids and improved understanding of nano-bio interactions in vivo are continuously fueling the optimization of LNP systems for delivering nucleic acid therapeutics. Deciphering intracellular trafficking pathways and mechanism (s) of endosomal escape will facilitate efforts to boost LNP potency [274][275][276][277]. Elucidating the nature and dynamic of the biomolecular corona formed on LNPs (following intravenous injection) and understanding its implications for biodistribution will be crucial to develop gene therapies beyond the liver [162][163][164]278,279].…”

Section: Future Perspectivesmentioning

confidence: 99%

Lipid nanoparticle technology for therapeutic gene regulation in the liver

Witzigmann

Kulkarni

Leung

et al. 2020

Advanced Drug Delivery Reviews

241

168

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Hereditary genetic disorders, cancer, and infectious diseases of the liver affect millions of people around the globe and are a major public health burden. Most contemporary treatments offer limited relief as they generally aim to alleviate disease symptoms. Targeting the root cause of diseases originating in the liver by regulating malfunctioning genes with nucleic acid-based drugs holds great promise as a therapeutic approach. However, employing nucleic acid therapeutics in vivo is challenging due to their unfavorable characteristics. Lipid nanoparticle (LNP) delivery technology is a revolutionary development that has enabled clinical translation of gene therapies. LNPs can deliver siRNA, mRNA, DNA, or gene-editing complexes, providing opportunities to treat hepatic diseases by silencing pathogenic genes, expressing therapeutic proteins, or correcting genetic defects. Here we discuss the state-of-the-art LNP technology for hepatic gene therapy including formulation design parameters, production methods, preclinical development and clinical translation.

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Endocytic Profiling of Cancer Cell Models Reveals Critical Factors Influencing LNP-Mediated mRNA Delivery and Protein Expression

Cited by 65 publications

References 47 publications

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

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

DOPC versus DOPE as a helper lipid for gene-therapies: molecular dynamics simulations with DLin-MC3-DMA

Lipid nanoparticle technology for therapeutic gene regulation in the liver

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