From Pulmonary Surfactant, Synthetic KL4 Peptide as Effective siRNA Delivery Vector for Pulmonary Delivery,Molecular Pharmaceutics

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From Pulmonary Surfactant, Synthetic KL4 Peptide as Effective siRNA Delivery Vector for Pulmonary Delivery
Molecular Pharmaceutics ( IF 4.5 ) Pub Date : 2017-11-17 00:00:00 , DOI: 10.1021/acs.molpharmaceut.7b00725
Yingshan Qiu ₁ , Michael Y. T. Chow ₁ , Wanling Liang ₁ , Winnie W. Y. Chung ₁ , Judith C. W. Mak _{1,

2} , Jenny K. W. Lam ₁

Affiliation

Pulmonary delivery of small interfering RNA (siRNA) has huge potential for the treatment of a wide range of respiratory diseases. The ability of naked siRNA to transfect cells in the lungs without a delivery vector has prompted the investigation of whether an endogenous component is at least partially responsible for the cellular uptake of siRNA, and whether a safe and efficient delivery system could be developed from this component to further improve the transfection efficiency. Surfactant protein B (SP-B), a positively charged protein molecule found in lung surfactant, is one of the possible candidates. While the role of SP-B in siRNA transfection remains to be determined, the SP-B mimic, synthetic KL4 peptide, was investigated in this study as a potential siRNA carrier. KL4 is a 21-residue cationic peptide that was able to bind to siRNA to form nanosized complexes. It mediated siRNA transfection effectively in vitro on human lung epithelial cells, A549 cells, and BEAS-2B cells, which was comparable to Lipofectamine 2000. When commercial pulmonary surfactant (Infasurf) was added in the transfection medium, the gene silencing effect of siRNA in cells transfected with Lipofectamine 2000 was completely abolished, whereas those transfected with KL4 remained unaffected. At 4 °C, KL4 failed to deliver siRNA into the cells, indicating that an energy-dependent process was involved in the uptake of the complexes. Chlorpromazine (inhibitor of chathrin-mediated endocytosis), but not nystatin (inhibitor of caveolae-mediated endocytosis), inhibited the uptake of KL4/siRNA complexes, suggesting that they entered cells through clathrin-mediated endocytosis. There was no sign of cytotoxicity or immune response caused by KL4 and KL4/siRNA complexes. Overall, this study demonstrated that synthetic KL4 peptide is a promising candidate for siRNA carrier for pulmonary delivery and could be a potential platform for delivering other types of nucleic acid therapeutics.

中文翻译：

肺表面活性剂合成的KL4肽作为有效的siRNA肺递送载体

肺部传递的小干扰RNA（siRNA）具有治疗多种呼吸系统疾病的巨大潜力。裸siRNA在没有递送载体的情况下转染肺细胞的能力促使人们研究内源性组分是否至少部分负责细胞对siRNA的摄取，以及是否可以从该组分开发出安全有效的递送系统进一步提高转染效率。表面活性剂蛋白B（SP-B）是在肺表面活性剂中发现的带正电的蛋白分子，是可能的候选物之一。虽然尚需确定SP-B在siRNA转染中的作用，但在这项研究中将SP-B模拟的合成KL4肽作为潜在的siRNA载体进行了研究。KL4是21个残基的阳离子肽，能够与siRNA结合形成纳米复合物。有效介导siRNA转染体外对人肺上皮细胞，A549细胞和BEAS-2B细胞的作用与Lipofectamine 2000相当。当在转染培养基中添加商业肺表面活性剂（Infasurf）时，siRNA在用Lipofectamine 2000转染的细胞中具有完全的基因沉默作用。废除，而那些用KL4转染的仍不受影响。在4°C下，KL4无法将siRNA传递到细胞中，这表明能量依赖过程参与了复合物的摄取。氯丙嗪（chathrin介导的内吞作用的抑制剂），但制霉菌素（制霉菌素介导的小孔介导的胞吞作用的抑制剂）却不抑制KL4 / siRNA复合物的摄取，表明它们通过网格蛋白介导的内吞作用进入细胞。没有由KL4和KL4 / siRNA复合物引起的细胞毒性或免疫应答的迹象。全面的，

更新日期：2017-11-19

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