Vandenbroucke Roosmarijn E, De Geest Bruno G, Bonné Stefan, Vinken Mathieu, Van Haecke Tamara, Heimberg Harry, Wagner Ernst, Rogiers Vera, De Smedt Stefaan C, Demeester Joseph, Sanders Niek N
Laboratory of General Biochemistry and Physical Pharmacy, Ghent University, Ghent, Belgium.
J Gene Med. 2008 Jul;10(7):783-94. doi: 10.1002/jgm.1202.
Small interfering (si)RNA mediated inhibition of oncogenes or viral genes may offer great opportunities for the treatment of several diseases such as hepatocellular carcinoma and viral hepatitis. However, the development of siRNAs as therapeutic agents strongly depends on the availability of safe and effective intracellular delivery systems. Poly(beta-amino esters) (PbAEs) are, in contrast to many other cationic polymers evaluated in siRNA delivery, biodegradable into smaller, nontoxic molecules.
We show for the first time that PbAE : siRNA complexes, containing 1,4-butanediol (PbAE1) or 1,6-hexanediol (PbAE2) diacrylate-based polymers, induced efficient gene silencing in both hepatoma cells and primary hepatocytes without causing significant cytotoxicity. Furthermore, carriers that slowly release the siRNA into the cytoplasm and hence induce a prolonged gene silencing are of major clinical interest, especially in fast dividing tumour cells. Therefore, we also studied the duration of gene silencing in the hepatoma cells and found that it was maintained for at least 5 days after siRNA delivery with PbAE2, the polymer with the slowest degradation kinetics.
From the time-dependent cellular distribution of these PbAE : siRNA complexes, we suggest that the slowly degrading PbAE2 causes a sustained endosomal release of siRNA during a much longer period than PbAE1. This may support the hypothesis that the endosomal release mechanism of PbAE : siRNA complexes is based on an increase of osmotic pressure in the endosomal vesicles after polymer hydrolysis. In conclusion, our results show that both PbAEs, and especially PbAE2, open up new perspectives for the development of efficient biodegradable siRNA carriers suitable for clinical applications.
小干扰（si）RNA介导的癌基因或病毒基因抑制作用可能为治疗多种疾病（如肝细胞癌和病毒性肝炎）提供巨大机遇。然而，将siRNAs开发为治疗剂在很大程度上取决于安全有效的细胞内递送系统的可用性。与许多其他用于siRNA递送评估的阳离子聚合物不同，聚（β-氨基酯）（PbAEs）可生物降解为更小的无毒分子。
我们首次表明，含有基于1,4-丁二醇（PbAE1）或1,6-己二醇（PbAE2）二丙烯酸酯聚合物的PbAE:siRNA复合物在肝癌细胞和原代肝细胞中均能诱导有效的基因沉默，且不会引起明显的细胞毒性。此外，能将siRNA缓慢释放到细胞质中从而诱导长时间基因沉默的载体具有重要的临床意义，尤其是在快速分裂的肿瘤细胞中。因此，我们还研究了肝癌细胞中基因沉默的持续时间，发现用降解动力学最慢的聚合物PbAE2递送siRNA后，基因沉默至少维持了5天。
根据这些PbAE:siRNA复合物随时间变化的细胞分布情况，我们认为降解缓慢的PbAE2比PbAE1能在更长时间内持续从内体释放siRNA。这可能支持以下假设：PbAE:siRNA复合物的内体释放机制是基于聚合物水解后内体囊泡渗透压的增加。总之，我们的结果表明，两种PbAEs，尤其是PbAE2，为开发适用于临床应用的高效可生物降解siRNA载体开辟了新的前景。
