Thermally poly-condensed lysine and lysine-co-histidine for siRNA delivery

IntroductionSmall interfering RNA (siRNA) has been intensively investigated over the last decades as a promising approach for addressing several diseases. However, the clinical applications of this approach have been hindered... [ view full abstract ]

Introduction

Small interfering RNA (siRNA) has been intensively investigated over the last decades as a promising approach for addressing several diseases. However, the clinical applications of this approach have been hindered due to several biological and technical obstacles that face siRNA delivery. In this context, highly branched and easily functionalized polymers have emerged as attractive materials to develop a safe, efficient and cost-effective delivery system. Here, we aimed to prepare a non-viral vector for siRNA based on hyperbranched polymers due to their advantages in terms of synthesis cost that make them more feasible and applicable for scale-up and manufacturing.

Method

Hyperbranched polylysine was synthesised and modified with histidine during thermal poly-condensation of amino acids. The resultant materials were characterised and their capability to condense and deliver siRNA were evaluated on A549 cells.

Results

The conditions of polymerisation were optimised to afford water-soluble hyperbranched polylysine and hyperbranched polylysine-co-histidine with average molecular weight up to 30 kDa. The physicochemical characterisation indicated that the incorporation of histidine produced polymers of a higher degree of branching, glass transition temperature and buffer capacity (Table 1). The size measurements and protection assay (Figure 1) indicated that the polymers of high molecular weight (~30 kDa) have better ability to condense and protect siRNA into nanoparticles than the lower molecular weights polymers (~15 kDa). Also, a negative impact of histidine incorporation can be seen clearly on the results. Biologically, hb-pK-33kDa achieved the best transfection efficiency (Figure 2) in comparison with other polymers.

Discussion  Structurally, the results revealed that the incorporation of histidine modulates the structures of polymers this more likely through occupying the more reactive ε-amine of lysine and directs the polymerisation towards the α-amine of the monomers; consequently, more dendritic polymers were prepared, with a more rigidity as indicated by the DB and Tg values. As a result, the high molecular weight polylysine interacted efficiently with siRNA and afforded polyplexes with a higher surface charge and a lower tendency to aggregate in comparison with the other polymers.