Abstract
AbstractSmall interfering RNAs (siRNAs) are a new class of promising therapeutic molecules that can be used for sequence-specific downregulation of disease-causing genes. However, endosomal entrapment of siRNA is a key hurdle for most delivery strategies, limiting the therapeutic effect. Here, we use live-cell microscopy and cytosolic galectin-9 as a sensor of membrane damage, to probe fundamental properties of endosomal escape of cholesterol-conjugated siRNA induced by endosome-disrupting compounds. We demonstrate efficient release of ligand-conjugated siRNA from vesicles damaged by small molecules, enhancing target knockdown up to ∼47-fold in tumor cells. Still, mismatch between siRNA-containing and drug-targeted endolysosomal compartments limits siRNA activity improvement. We also show widespread endosomal damage in macroscopic tumor spheroids after small molecule treatment, substantially improving siRNA delivery and knockdown throughout the spheroid. We believe the strategy to characterize endosomal escape presented here will be widely applicable, facilitating efforts to improve delivery of siRNA and other nucleic acid-based therapeutics.
Publisher
Springer Science and Business Media LLC
Subject
General Physics and Astronomy,General Biochemistry, Genetics and Molecular Biology,General Chemistry
Reference44 articles.
1. Brummelkamp, T. R., Bernards, R. & Agami, R. Stable suppression of tumorigenicity by virus-mediated RNA interference. CCELL 2, 243–247 (2002).
2. Barton, G. M. & Medzhitov, R. Retroviral delivery of small interfering RNA into primary cells. Proc. Natl. Acad. Sci. USA 99, 14943–14945 (2002).
3. Zimmermann, T. S. et al. RNAi-mediated gene silencing in non-human primates. Nature 441, 111–114 (2006).
4. Soutschek, J., Akinc, A., Bramlage, B. & Charisse, K. Therapeutic silencing of an endogenous gene by systemic administration of modified siRNAs. Nature 432, 173–178 (2004).
5. Wolfrum, C. et al. Mechanisms and optimization of in vivo delivery of lipophilic siRNAs. Nat. Biotechnol. 25, 1149–1157 (2007).
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