Small interfering RNA (siRNA) is a powerful gene silencing tool and has been considered a potential agent for
the treatment of many diseases. However, development of safe and effective siRNA delivery systems still
remains a great challenge. In this study, we developed a new siRNA delivery system based on the
electrostatic encapsulation of siRNA/cationic vector complexes with dendrimer-like polymeric DNAs
(YY-DNAs). The binary complexes of siRNA with cationic vectors such as lipofectamine (LP),
polyethylenimine (PEI) and poly-L-lysine (PLL) were first constructed. Then, the encapsulation was
performed by the addition of YY-DNAs to the binary complexes in order to form stable complexes of
siRNA/LP/YY-DNAs, siRNA/PEI/YY-DNAs and siRNA/PLL/YY-DNAs. The encapsulated siRNA complexes
showed nearly spherical morphology with about 13–37 nm average hydrodynamic size and their
z-potentials were negative. The cationic complexes of siRNA/LP, siRNA/PEI and siRNA/PLL showed a
high cytotoxicity towards the cells and strong aggregation with erythrocytes, while their encapsulation
into YY-DNAs dramatically decreased the toxicities of complexes. Furthermore, these anionic
encapsulated siRNA complexes were highly taken up by the HeLa cells and showed extremely high
cellular uptake efficiencies and gene silencing effects without such cytotoxicity and aggregation. The
stability of these complexes in 10% FBS and human serum was investigated and they showed high
stability even after incubation for 72 h and 48 h, respectively. Therefore, we have newly identified safe
and efficient anionic complexes of siRNA for clinical uses.