Imaging small molecule-induced endosomal escape of siRNA.,Nature Communications

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Imaging small molecule-induced endosomal escape of siRNA.
Nature Communications ( IF 14.7 ) Pub Date : 2020-04-14 , DOI: 10.1038/s41467-020-15300-1
Hampus Du Rietz _{1,

2} , Hampus Hedlund _{1,

2} , Sten Wilhelmson _{1,

2,

3} , Pontus Nordenfelt ₄ , Anders Wittrup _{1,

2,

3}

Affiliation

Small 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.

中文翻译：

成像小分子诱导的siRNA内体逃逸。

小干扰RNA（siRNA）是一类有前景的新型治疗分子，可用于引起疾病的基因的序列特异性下调。但是，siRNA的内体包埋是大多数递送策略的关键障碍，限制了治疗效果。在这里，我们使用活细胞显微镜和胞质半乳糖凝集素9作为膜损伤的传感器，以探测破坏内体的化合物诱导的胆固醇结合的siRNA的内体逃逸的基本特性。我们证明了从受小分子破坏的囊泡中有效释放配体缀合的siRNA，增强了肿瘤细胞中的靶基因敲低至约47倍。仍然，含siRNA的和靶向药物的溶酶体区室之间的不匹配限制了siRNA活性的提高。我们还显示小分子治疗后宏观肿瘤球体中的广泛的内体损伤，大大改善了整个球体的siRNA传递和敲低。我们相信，此处介绍的表征内体逃逸的策略将广泛适用，从而有助于改善siRNA和其他基于核酸的治疗剂的递送。

更新日期：2020-04-24

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