The modification of siRNA with 3′ cholesterol to increase nuclease protection and suppression of native mRNA by select siRNA polyplexes

• Published in: Biomaterials 2011-02, Vol.32 (5), p.1404-1411
• Main Authors: Ambardekar, Vishakha V, Han, Huai-Yun, Varney, Michelle L, Vinogradov, Serguei V, Singh, Rakesh K, Vetro, Joseph A
• Format: Article
• Language: English
• Subjects: Advanced Basic Science; Animals; Cells, Cultured; Cholesterol - analogs & derivatives; Cholesterol - chemistry; Dentistry; Female; H-2K b-ts-A58; Immortomouse; Mammary fat pad; Mice; PEGss; PEGylated cationic polymers; Polyethylene Glycols - chemistry; Polyethyleneimine - chemistry; Polymers - chemistry; PPIB; Ribonucleases - metabolism; RNA, Messenger - metabolism; RNA, Small Interfering - chemistry; RNA, Small Interfering - metabolism
• ISSN: 0142-9612; 1878-5905
• DOI: 10.1016/j.biomaterials.2010.10.019

Abstract Polymer-siRNA complexes (siRNA polyplexes) are being actively developed to improve the therapeutic application of siRNA. A major limitation for many siRNA polyplexes, however, is insufficient mRNA suppression. Given that modifying the sense strand of siRNA with 3′ cholesterol (chol-siRNA) increases the activity of free nuclease-resistant siRNA in vitro and in vivo , we hypothesized that complexation of chol-siRNA can increase mRNA suppression by siRNA polyplexes. In this study, the characteristics and siRNA activity of self assembled polyplexes formed with chol-siRNA or unmodified siRNA were compared using three types of conventional, positively charged polymers: (i) biodegradable, cross-linked nanogels (BDNG) (ii) graft copolymers (PEI-PEG), and (iii) linear block copolymers (PLL10-PEG, and PLL50-PEG). Chol-siRNA did not alter complex formation or the resistance of polyplexes to siRNA displacement by heparin but increased nuclease protection by BDNG, PLL10-PEG, and PLL50-PEG polyplexes over polyplexes with unmodified siRNA. Chol-CYPB siRNA increased suppression of native CYPB mRNA in mammary microvascular endothelial cells (MVEC) by BDNG polyplexes (35%) and PLL10-PEG polyplexes (69%) over comparable CYPB siRNA polyplexes but had no effect on PEI-PEG or PLL50-PEG polyplexes. Overall, these results indicate that complexation of chol-siRNA increases nuclease protection and mRNA suppression by select siRNA polyplexes. These results also suggest that polycationic block length is an important factor in increasing mRNA suppression by PLL-PEG chol-siRNA polyplexes in mammary MVEC.