Yu Zhili, Ye Junxiao, Pei Xing, Sun Lu, Liu Ergang, Wang Jianxin, Huang Yongzhuo, Lee Seung Jin, He Huining
Tianjin Key Laboratory on Technologies Enabling Development of Clinical Therapeutics and Diagnostics, School of Pharmacy, Tianjin Medical University, Tianjin 300070, China.
College of Pharmacy, Tsinghua University, Beijing 100084, China.
Acta Pharm Sin B. 2018 Jan;8(1):116-126. doi: 10.1016/j.apsb.2017.11.011. Epub 2017 Dec 19.
RNAi technology has aroused wide public interest due to its high efficiency and specificity to treat multiple types of diseases. However, the effective delivery of siRNA remains a challenge due to its large molecular weight and strong anionic charge. Considering their remarkable functions and features that are often desired in drug delivery carriers, biomimetic systems for siRNA delivery become an effective and promising strategy. Based on this, covalent attachment of synthetic cell penetrating peptides (CPP) to siRNA has become of great interest. We developed a monomeric covalent conjugate of low molecular weight protamine (LMWP, a well-established CPP) and siRNA a cytosol-cleavable disulfide linkage using PEG as a crosslinker. Results showed that the conjugates didn't generate coagulation, and exhibited much better RNAi potency and intracellular delivery compared with the conventional charge-complexed CPP/siRNA aggregates. Three different synthetic and purification methods were compared in order to optimize synthesis efficiency and product yield. The methodology using hetero-bifunctional NHS-PEG-OPSS as a crosslinker to synthesize LMWP-siRNA simplified the synthesis and purification process and produced the highest yield. These results pave the way towards siRNA biomimetic delivery and future clinical translation.
RNA干扰（RNAi）技术因其高效性和特异性可用于治疗多种疾病，已引起广泛的公众关注。然而，由于小干扰RNA（siRNA）分子量较大且带强负电荷，其有效递送仍然是一个挑战。考虑到它们在药物递送载体中通常所需的显著功能和特性，用于siRNA递送的仿生系统成为一种有效且有前景的策略。基于此，将合成的细胞穿透肽（CPP）共价连接到siRNA上已引起了极大的关注。我们使用聚乙二醇（PEG）作为交联剂，开发了一种低分子量鱼精蛋白（LMWP，一种成熟的CPP）与siRNA的单体共价缀合物，通过可在细胞质中裂解的二硫键连接。结果表明，与传统的电荷复合CPP/siRNA聚集体相比，该缀合物不会产生凝聚，并且表现出更好的RNA干扰活性和细胞内递送能力。为了优化合成效率和产物收率，比较了三种不同的合成和纯化方法。使用异双功能N-羟基琥珀酰亚胺-聚乙二醇-邻吡啶二硫醚（NHS-PEG-OPSS）作为交联剂合成LMWP-siRNA的方法简化了合成和纯化过程，并产生了最高的收率。这些结果为siRNA仿生递送和未来的临床转化铺平了道路。
