Polyphosphoramidate gene carriers: effect of charge group on gene transfer efficiency.
Cationic polymeric carriers have been widely used for gene delivery. However, the structure-function relationship, especially the effect of charge groups of cationic polymeric carriers on the transfection activity, is poorly understood. To examine this important parameter, a series of cationic polymers, polyphosphoramidates (PPAs) with an identical backbone, same side chain spacer, similar molecular weights but different charge groups containing primary to quaternary amino groups (PPA-EA, PPA-MEA, PPA-DMA and PPA-TMA, Figure 1) were synthesized. The DNA-binding affinity of these four PPAs increased in the order of PPA-EA<PPA-MEA<PPA-DMA approximately PPA-TMA. The cytotoxicity decreased in the order of PPA-EA>PPA-MEA>PPA-DMA>PPA-TMA. Particle size and zeta potential of four different types of PPA/DNA nanoparticles did not show significant correlation with PPA structure. These PPAs did not show significant buffering capacity within pH 5-7, even though transfection mediated by PPA-EA was the only one that seemed to be limited by endolysomal escape. Endocytosis of DNA mediated by PPAs was also similar (17-22%) for all four PPAs. However, the transfection efficiency of these PPAs varied significantly. In vitro transfection efficiency of PPAs decreased in the order of PPA-EA>PPA-MEA>PPA-DMA approximately PPA-TMA. Nanoparticles with PPA-EA containing primary amino groups gave the highest transfection efficiency in cell lines at the charge ratios from 6/1 to 20/1 (+/-). Matching the trend of transfection efficiency observed in vitro, PPA-EA mediated the highest transgene expression, comparable to that of polyethylenimine, in the spinal cord following intrathecal injection of the nanoparticles. These results establish that PPA gene carriers with primary amino group side chains are more potent than those with secondary, tertiary or quaternary amino groups in vitro and in the intrathecal gene delivery model.
Department of Biomedical Engineering, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
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Zhang
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Mao
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Leong
D000818:Animals; D019556:COS Cells; D002412:Cations; D002460:Cell Line; D002850:Chromatography, Gel; D004247:DNA / Q000008:administration & dosage*; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage* / Q000138:chemical synthesis; D007278:Injections, Spinal; D008156:Luciferases / Q000235:genetics; D008297:Male; D036103:Nanotechnology; D011108:Polymers; D051381:Rats; D017208:Rats, Wistar; D013116:Spinal Cord / Q000201:enzymology*; D014162:Transfection / Q000379:methods*; D019076:Transgenes
D002412:Cations; D011108:Polymers; D004247:DNA; D008156:Luciferases