| RNA interference (RNAi) techniques hold forth great promise for therapeutic silencing of deleterious genes. However, clinical applications of RNAi require the development of safe and efficient methods for intracellular delivery of small interfering RNA (siRNA) oligonucleotides specific to targeted genes. We describe the use of a lipitoid, a cationic oligopeptoid-phospholipid conjugate, for non-viral transfection of synthetic siRNA oligos in cell culture. This peptidomimetic delivery vehicle allows for efficient siRNA transfection in a variety of human cell lines with negligible toxicity and promotes extensive downregulation of the targeted genes at both the protein and the mRNA level. We compare the lipitoid reagent to a standard commercial transfection reagent. The lipitoid is highly efficient even in primary IMR-90 human lung fibroblasts in which other commercial reagents are typically ineffective. Electron microscopy imaging data reveals that lipitoid molecules interact with siRNA oligonucleotides to form well-defined, spherical complexes in the nanometer range. The morphology of these complexes is shown to be dependent on various physiological parameters, which can be readily controlled in the lipitoid-siRNA delivery system. We associate the physical properties to the biological activity of the complexes, concluding that the charge ratio and size of the complexes are of critical importance to attain RNAi effects. These findings are critical to the development of well-controlled, efficient delivery systems. Due to the tunable nature of the peptidomimetic region, this system can be further elaborated to display ligands for the targeting of the complexes to specific tissues. |
