Nonviral Gene Delivery: Gemini Bispyridinium Surfactant-Based DNA Nanoparticles

Abstract

The interaction with a model membrane, the formation of DNA nanoparticles, and the transfection ability of a homologous series of bispyridinium dihexadecyl cationic gemini surfactants, differing in the length of the alkyl spacer bridging the two pyridinium polar heads in the 1 and 1' positions (P16-n with n = 3, 4, 8, 12), have been studied by means of differential scanning calorimetry (DSC), atomic force microscopy, electrophoresis mobility shift assay, and transient transfection assay measurements. The results presented here show that their performance in gene delivery is strictly related to their structure in solution. For the first time the different transfection activities of the compounds can be explained by referring to their thermodynamic properties in solution, previously studied. The compound with a spacer formed by four carbon atoms, showing unexpected enthalpic properties vs concentration in solution, is the only one giving rise to a transfection activity comparable to that of the commercial reagent, when formulated with L-α-dioleoylphosphatidylethanolamine. We suggest that P16-4 behaves like molecular tongs able to grip basic groups near each other, allowing the formation of compact and nearly spherical DNA particles. The compound with the longest spacer gives rise to loosely condensed structures by forming a sort of bow, not able to give rise to transfection notwithstanding the double positive charge of the molecule. On the other hand, DSC measurements on synthetic membranes show that the compounds with the shortest spacers (three and four methylene groups) practically do not interact with the 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine membrane, while compounds P16-8 and, particularly, P16-12 induce the formation of surfactant-rich and surfactant-poor domains in the membrane, without showing any peculiarity for compound P16-4. This could suggest that the mechanisms involved in the interaction with the model membrane and in gene delivery are substantially different and could strike a blow for an endocytosis mechanism for the internalization in the cell of the DNA nanoparticles.

Synthesis, Physicochemical Characterization, and Interaction with DNA of Long‐Alkyl‐Chain Gemini Pyridinium Surfactants

“…[64] Higher activity has been observed when the transfecting agent hasf our or eight methylene groups,w hich suggestsapossible correlation of biological activity with counterion binding (b)a nd A min parameter and also with the visual inspection of AFM images for which only those surfactants showeda na bility to compactDNA. [64] The variation of the spacer length is known to modifyt he packing parameter [46,59] and to tune the aggregation properties [84,85] (CMC, micellar aggregation number,m icellar shape, etc.) and these preliminar resultsi ndicatet hat it affects the properties of the surfactant-DNA lipoplexes.…”

“…For am ore detailed discussion about the gene-delivery ability of the compounds presented here, see Ref. [64].…”

“…This structural feature, which determines the peculiar behavior of surfactants 2b and 3b for A min and the degree of counterion binding b,a lso seems to be highly relevant for their activity in transfection experiments as suggested in our recent paper where the DNA shift and the interaction with membranes of presents eries of surfactants are described. [64] In view of these results, it appears interesting to explore similar structures that bear ah ydrolyzable bond such as amide or ester, [86] to improve the biodegradability and possibly the viability to the cells, since usually quaternary compounds show bactericidal activity and limited biodegradability that, however, do not hamper their use as gene carriers.…”

Recent progress in gene therapy to deliver nucleic acids with multivalent cationic vectors

“…The cationic groups commonly used are ammonium [53,54], imidazolium [55][56][57], pyridinium [58,59], lysine or arginine [60][61][62], and/or amine/polyamines [63][64][65][66][67].…”

“…[45,75,78,117,174]. Isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC) and termogravimetric analysis (TGA) have been also used to analyze the interaction energy and related thermodynamic information during the formation of lipoplexes formed by some GCLs with pDNA [59,91,95,96,117,137,175], or polyplexes with cationic polymers and pDNA [95,162,176] or siRNA [153]. NMR spectroscopy has been used to analyze the lipoplex structure and the CL diffusion during the formation of lipoplexes formed by GCLs [53,111,112] and also for polyplexes with cationic polymers [95,98] or polyrotaxanes [89,90].…”