Dicationic Amino Substituted Gemini Surfactants and their Nanoplexes: Improved Synthesis and Characterization of Transfe
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RESEARCH PAPER
Dicationic Amino Substituted Gemini Surfactants and their Nanoplexes: Improved Synthesis and Characterization of Transfection Efficiency and Corneal Penetration In Vitro Lokesh Narsineni 1 & Marianna Foldvari 1,2
Received: 29 February 2020 / Accepted: 29 April 2020 / Published online: 14 July 2020 # Springer Science+Business Media, LLC, part of Springer Nature 2020
ABSTRACT Purpose To formulate and characterize nanoparticles from m-7NH-m gemini surfactants, synthesized by a new improved method, for non-invasive gene delivery including optimization of composition for transfection efficiency and corneal penetration. Methods A one-pot, solvent-free, DMAP-free method was developed for the synthesis of m-7NH-m (m = 12– 18) gemini surfactant series. Lipoplexes (LPXs) and nanoplexes (NPXs) of gemini surfactant-plasmid DNA were formulated with and without DOPE helper lipid, respectively, at various charge ratios and characterized by dynamic light scattering and zeta potential measurements. Transfection efficiency, cellular toxicity, effect of DOPE and gene expression kinetic studies were carried out in A7 astrocytes by flow cytometry and confocal microscopy. Corneal penetration studies of 18-7NH-18 NPXs were carried out using 3D EpiCorneal® tissue model. Results The new synthesis method provides a two-fold improved yield and the production of a pure species of m7NH-m without DMAP and trimeric m-7N(m)-m surfactants as impurities. Structure and purity was confirmed by ESI-MS, 1 H NMR spectroscopy and surface tension measurements. Particle size of 199.80 ± 1.83 nm ± S.D. and a zeta potential value of +30.18 ± 1.17 mV ± S.D. was obtained for 18-7NH-
Guest Editor: Sheng Qi * Marianna Foldvari [email protected] 1
School of Pharmacy, University of Waterloo, 200 University Avenue West, Waterloo, Ontario N2L 3G1, Canada
2
School of Pharmacy, Center for Bioengineering and Biotechnology, Waterloo Institute of Nanotechnology, University of Waterloo, 200 University Avenue West, Waterloo, Ontario N2L 3G1, Canada
18 5:1 ratio NPXs showed optimum transfection efficiency (10.97 ± 0.11%) and low toxicity (92.97 ± 0.57% viability) at the 48-h peak expression. Inclusion of DOPE at 1: 0.5 and 1:1 ratios to gemini surfactant reduced transfection efficiency and increased toxicity. Treatment of EpiCorneal® tissue model showed deep penetration of up to 100 μm with 18-7NH-18 NPXs. Conclusion Overall, 18-7NH-18 NPXs are potential gene delivery systems for ophthalmic gene delivery and for further in vivo studies.
KEY WORDS confocal microscopy . cornea . flow cytometry . gemini surfactant . gene expression . neurotrophic factors . non-viral gene delivery
ABBREVIATIONS CMC CLSM DOPE DMAP ESI-MS GP GPL GFP 1 H NMR LPX NPX TE TFA TIS DMEM FBS
Critical micelle concentration Confocal laser scanning microscope 1,2-Dioleoyl-sn-glycero-3-phosphoethanolamine 4-(Dimethylamino)pyridine Electro-spray ionization mass spectrometry gemini:plasmid gemini:plasmid:lipid Green fluorescent protein Proton- nuclear magnetic resonance spectrosc
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