Department of Medical Biochemistry, University of Shizuoka, School of Pharmaceutical Sciences, 52-1 Yada, Suruga-ku, Shizuoka 422-8526, Japan.
Nippon Fine Chemical Co. Ltd., 5-1-1 Umei, Takasago, Hyogo 676-0074, Japan.
Int J Pharm. 2020 Jul 30;585:119479. doi: 10.1016/j.ijpharm.2020.119479. Epub 2020 May 27.
RNA interference induced by small interfering RNA (siRNA) is a promising strategy for the treatment of various intractable diseases including cancer. Lipid nanoparticles (LNP) composed of ionizable lipids and siRNA are known as a leading siRNA delivery system. However, LNPs composed of conventional ionizable lipids will be aggregated in the physiological environment because of loss of ionization. Therefore, the inclusion of hydrophilic polymer-conjugated lipids such as polyethylene glycol (PEG)-conjugated lipid is required to improve the LNP stability. Herein, we synthesized a novel charge-reversible lipid derivative, dioleoylglycerophosphate-diethylenediamine conjugate (DOP-DEDA). The surface of LNP composed of DOP-DEDA (DOP-DEDA LNP) was constantly ionized and positively charged at pH 6.0, almost neutral at pH 7.4, and negatively charged at pH 8.0. Importantly, DOP-DEDA LNP were stable in the physiological milieu without PEG-conjugated lipid. DOP-DEDA LNP disrupted the red blood cells only under the low-pH condition in a hemolysis assay, suggesting that the interaction between DOP-DEDA LNP and biological membranes is pH-dependent. DOP-DEDA LNP encapsulating siRNA against polo-like kinase 1 (siPLK1) highly suppressed the expression of PLK1 mRNA and its protein. The cellular uptake of DOP-DEDA LNP was increased in an apolipoprotein E3 (apoE3) dose-dependent manner. In addition, DOP-DEDA LNP was taken up into cancer cells via both clathrin- and caveola-mediated endocytosis pathways. These findings indicate that LNP composed of this charge-reversible lipid should be a highly stable and potent siRNA delivery vector.
小干扰 RNA（siRNA）诱导的 RNA 干扰是治疗各种难治性疾病（包括癌症）的一种有前途的策略。由可离子化脂质和 siRNA 组成的脂质纳米颗粒（LNP）被认为是一种领先的 siRNA 递送系统。然而，由于失去电离，由常规可离子化脂质组成的 LNPs 在生理环境中会聚集。因此，需要包含亲水性聚合物缀合脂质（如聚乙二醇（PEG）缀合脂质）来提高 LNP 的稳定性。在此，我们合成了一种新型电荷可逆脂质衍生物，二油酰基甘油磷酸二乙二胺缀合物（DOP-DEDA）。由 DOP-DEDA 组成的 LNP（DOP-DEDA LNP）的表面在 pH 6.0 时不断电离并带正电荷，在 pH 7.4 时几乎呈中性，在 pH 8.0 时带负电荷。重要的是，DOP-DEDA LNP 在没有 PEG 缀合脂质的生理环境中是稳定的。在溶血试验中，DOP-DEDA LNP 仅在低 pH 条件下破坏红细胞，表明 DOP-DEDA LNP 与生物膜的相互作用是 pH 依赖性的。包封针对 Polo 样激酶 1（siPLK1）的 siRNA 的 DOP-DEDA LNP 高度抑制了 PLK1 mRNA 和其蛋白的表达。DOP-DEDA LNP 的细胞摄取呈载脂蛋白 E3（apoE3）剂量依赖性增加。此外，DOP-DEDA LNP 通过网格蛋白和小窝蛋白介导的内吞途径被摄取到癌细胞中。这些发现表明，由这种电荷可逆脂质组成的 LNP 应该是一种高度稳定和有效的 siRNA 递送载体。
