Polymer side-chain degradation as a tool to control the destabilization of polyplexes.
OBJECTIVE
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 degrees 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 degrees 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 degrees 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 degrees 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.
Funhoff
Arjen M
AM
Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands.
van Nostrum
Cornelus F
CF
Janssen
Adriënne P C A
AP
Fens
Marcel H A M
MH
Crommelin
Daan J A
DJ
Hennink
Wim E
WE
D000818:Animals; D001711:Biotransformation; D019556:COS Cells; D002522:Chlorocebus aethiops; D004247:DNA / Q000235:genetics / Q000493:pharmacokinetics; D004355:Drug Stability; D008689:Methacrylates / Q000737:chemistry / Q000493:pharmacokinetics; D011108:Polymers / Q000737:chemistry* / Q000493:pharmacokinetics*; D014162:Transfection / Q000379:methods
D008689:Methacrylates; D011108:Polymers; D004247:DNA; C049840:2-(dimethylamino)ethyl methacrylate; C032802:hydroxypropyl methacrylate