Study On Synthesis And Properties Of L-Arabinose-Based Cationic Liposomes For Gene Delivery

Gene therapy is a promising strategy to down or regulate the expression of defective genes by transporting foreign gene to targeted cells.The key factor of this high-performance therapeutics is required to design high efficient and safely gene delivery systems.Cationic lipids as one of the most promising non-viral gene delivery systems have been intensively investigated for their efficient transfection,non-immunogenic nature,low biotoxicity and good biocompatibility.In this paper,a series of cationic lipids?Ara-DiC12MA,Ara-DiC14MA,Ara-DiC16MA and Ara-DiC18MA?with different hydrophobic chain lengths were constructed using L-arabinose as starting material,by peracetylation,selective1-O-deacetylation,trichloroacetimi-dation,glycosylation,azidation,deacetylation,Staudinger reaction,tertiary amination and quaternization.The structures of these synthesized lipids and some intermediates were characterized by ¹H NMR,¹³C NMR,¹H-¹H COSY,¹H-¹³C HMQC and ESI-MS.The size,poly dispersition index and zeta of liposomes and lipid/pDNA complexs were evaluated by dynamic light scattering.The results showed that the average particle sizes of cationic lipids were111-135 nm,and the average sizes of pDNA-complexes vary from 63 nm to 181 nm.Meanwhile,the zeta potential of four naked liposomes is between 50 and 70 mV,and the zeta potential of lipoplexes were increased with the increasing of mole ratio.The Ara-DiC18MA lipoplexes exhibited a positively charged surface at the N/P ratios of 6:1.The Ara-DiC12MA and Ara-DiC16MA lipoplexes became positively charged at N/P ratios higher than 2:1.Nevertheless,Ara-DiC14MA always maintain positively charged under various N/P ratios?2:1-10:1?.Cationic lipids and lipolexes were well dispersed and had a regular spherical surface by atom force microscopy?AFM?.The affinity with pDNA was measured by gel electrophoresis assay.The DNA was successfully retarded by cationic liposomes at the N/P ratios of 1:1,escept Ara-DiC18MA.Using Lipo2000 as the positive control,the expression efficiency of cationic lipids carrying pDNA in HEK293,HeLa,PC-3,Mat,HepG2,SW480,MCF-7,7721 and A375 cells were estimated.As depicted in result,Ara-DiC16MA liposome possessed the highest transfection activity in all transfected cell lines compared with other three kinds of liposomes.At the same time,Ara-DiC18MA liposome has hardly any transfection efficacy in all cell lines.Additionally,all the cationic lipids showed low gene transfection efficiency in the HeLa and 7721 cells.According to the results of pDNA transfection,HEK293,PC-3 and Mat cells were selected to investigate the efficiency of siRNA delivery in vitro by using luciferase knockdown assay.The results were found that Ara-DiC14MA and Ara-DiC16MA lipids achieved perfectly silencing activity in these three cells.To further explore the correlation between cellular uptake and transfection efficiencies,the cellular uptake of the Ara-DiC16MA liposomes were analyzed qualitatively in HEK293,PC-3,Mat and HeLa cells.The result shows that the different of cellular uptake efficiencies were clearly observed in these four cells,which is consistent with the results of transfection efficiency.The results demonstrated that the higher cellular uptake capability may be one of the important parameters at pDNA expression.Meanwhile,cationic lipids Ara-DiC14MA and Ara-DiC16MA had low cytotoxicity in HEK293,HeLa,PC-3,Mat,HepG2,SW480,MCF-7,7721 and A375 cells.In a few words,cationic lipids Ara-DiC14MA and Ara-DiC16 MA exhibit highe transfection efficiency,silencing activity,cellular uptake capability and low cell cytotoxicy,paving the way for gene therapy with cationic lipids.