Polymer Side-Chain Degradation as a Tool to Control the Destabilization of Polyplexes

Purpose. We purposed to design a cationic polymer that binds to pDNA to form polyplexes and that subsequently degrades within a few days at physiological pH and temperature, releasing the DNA in the cytosol of a cell. Methods. We synthesized a new monomer carbonic acid 2-dimethylamino-ethyl ester 1-methyl-2-(2-methacryloylamino)-ethyl ester (abbreviated HPMA-DMAE) and the corresponding polymer. Hydrolysis of the carbonate ester of both the monomer and the polymer was investigated at 37°C. The DNA condensing properties of the pHPMA-DMAE was studied using dynamic light scattering (DLS) and zeta potential measurements. Degradation of the polyplexes at 37°C and pH 7.4 was monitored with DLS and gel electrophoresis. In vitro transfections were performed in COS-7 cell line. Results. pHPMA-DMAE is able to condense DNA into small particles (110 nm) with a positive zeta potential. The half-life of the polymer and monomer at 37°C and pH 7.4 was around 10 h whereas at pH 5, the half-life was 380 h. In line with this, due to hydrolysis of the side groups, pHPMA-DMAE-based polyplexes dramatically increased in size at 37°C and pH 7.4 whereas at pH 5.0, only a very small increase was observed. Interestingly, intact DNA was released from the polyplexes after 48 h at pH 7.4 whereas all DNA remained bound to the polymer at pH 5.0. Polyplexes were able to transfect cells with minimal cytotoxicity if the endosomal membrane-disrupting peptide INF-7 was added to the polyplex formulation. Conclusions. Degradation of the cationic side-chains of a polymer is a new tool for time-controlled release of DNA from polyplexes, preferably within the cytosol and/or nucleus.