A Universal Protein Tag for Delivery of SiRNA-Aptamer Chimeras

Liu, Hong Yan; Gao, Xiaohu

Our protein tag does not rely on high positive charge to interact with RNA molecules. In fact, it only recognizes relatively long dsRNAs (.16 bp) such as the siRNA segment and the short stem region of the aptamer in our chimera molecule. Extensive biochemistry investigations have shown that for the current length of the chimera, maximum two copies of dsRBD can bind to it with differential affinity (the first copy binds much stronger than the second copy). The gene silencing experiments conducted here reflect this effect since mixing chimera with 13 or 23 protein tags does not affect the silencing efficiency. Considering the molecular weights of the chimera (28.8 kDa) and the protein tag (24.8 kDa), molecular weight of the final complex at 151 binding will become 53.6 kDa. Based on well-documented size effect for in vivo drug delivery 49 , this size is sufficiently large to reduce premature renal clearance while still small enough for deep tissue penetration. For example, by tagging siRNA-aptamer chimera with a 20 kDa PEG, its in vivo circulating half-life has been shown to increase from approximately 30 min to 30 hours 15 ; whereas large nanoparticles (.30 nm) have been shown to be ineffective in tumor treatment except for some hyperpermeable tumors 50 .