Fusion-dependent formation of lipid nanoparticles containing macromolecular payloads
Fusion-dependent formation of lipid nanoparticles containing macromolecular payloads
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DOI：
10.1039/c9nr02004g
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发表时间：
2019-05-14
期刊：
影响因子：
6.7
通讯作者：
Cullis, Pieter R.
中科院分区：
文献类型：
--
作者：
Kulkarni, Jayesh A.;Witzigmann, Dominik;Cullis, Pieter R.
Onpattro（patisiran）的成功清楚地证明了脂质纳米颗粒（LNP）系统用于实现基因治疗的实用性。这些系统由可电离的阳离子脂质、磷脂、胆固醇和聚乙二醇（PEG）-脂质组成，通过快速混合乙醇-脂质溶液与酸性水溶液，然后透析至中和缓冲液中制备。详细了解LNP的形成机制对于改进LNP设计至关重要。在这里，我们使用低温透射电子显微镜和荧光技术，以进一步证明，LNP是通过融合的前体，pH敏感的脂质体成大的电子致密的核心结构的pH值被中和。接下来，我们表明，融合过程是有限的聚乙二醇-脂质上出现的颗粒的积累。最后，我们表明，融合依赖的形成机制也适用于LNP含有大分子有效载荷，包括mRNA，DNA载体，和金纳米粒子。
The success of Onpattro (patisiran) clearly demonstrates the utility of lipid nanoparticle (LNP) systems for enabling gene therapies. These systems are composed of ionizable cationic lipids, phospholipid, cholesterol, and polyethylene glycol (PEG)-lipids, and are produced through rapid-mixing of an ethanolic-lipid solution with an acidic aqueous solution followed by dialysis into neutralizing buffer. A detailed understanding of the mechanism of LNP formation is crucial to improving LNP design. Here we use cryogenic transmission electron microscopy and fluorescence techniques to further demonstrate that LNP are formed through the fusion of precursor, pH-sensitive liposomes into large electron-dense core structures as the pH is neutralized. Next, we show that the fusion process is limited by the accumulation of PEG-lipid on the emerging particle. Finally, we show that the fusion-dependent mechanism of formation also applies to LNP containing macromolecular payloads including mRNA, DNA vectors, and gold nanoparticles.