Cationic cholesterol derivative efficiently delivers the genes: in silico and in vitro studies

Abstract

The aims of the research work were to synthesize ethyl(cholesteryl carbamoyl)-L-arginate (ECCA), an arginine-conjugated cholesterol derivative, and to evaluate its application as a gene delivery vector. The interactions of ECCA with DNA duplex were studied using molecular dynamics (MD) simulations. It was found that the guanidine group of ECCA could interact with the phosphate group of DNA through ionic interactions as well as hydrogen bonds. The structure of DNA was stable throughout the simulation time. Liposomes were formulated using ECCA and soya phosphatidylcholine (SPC) by a thin-film hydration method. They had the particle size of ~ 150 nm and the zeta potential of + 51 mV. To ensure the efficient binding of DNA to the liposomes, the ratio of DNA to ECCA was optimized using gel retardation assay. Further, serum stability, haemolysis and cytotoxicity studies were carried out to determine the stability and safety of the lipoplexes. Circular dichroism spectroscopy was used to determine the interaction of DNA and cationic liposomes. Cellular uptake pathway was determined by studying the uptake of coumarin-loaded lipoplexes at 4 °C and in the presence of uptake inhibitors, i.e. genistein, chlorpromazine and methyl-β-cyclodextrin. Transfection studies were carried out to evaluate the transfection efficacy of the ECCA-loaded lipoplexes. The binding of DNA and lipoplexes was found to be stable in the presence of serum, and no degradation of DNA was observed. The lipoplexes showed low haemolysis and cytotoxicity. The uptake of coumarin-loaded liposomes was decreased up to ~ 20% in the presence of clathrin- and caveola-mediated uptake inhibitors, indicating a role of both the pathways in the uptake of the inhibitors. Satisfactory transfection efficiency was obtained compared to Lipofectamine. Thus, cationic cholesterol derivative is a useful tool for gene delivery vector.

Barriers and Strategies of Cationic Liposomes for Cancer Gene Therapy

“…Cationic liposomes (CLs) are near-spherical vesicles with a positive charge and are dominantly composed of cationic lipids that effectively condense nucleic acids 1 , 2 , 3 , 4 such as plasmid DNA (pDNA), messenger RNA (mRNA), or small interfering RNA (siRNA) via electrostatic interactions 5 to form complexes called “lipoplexes.” Based on the fact that genes have been revealed to be involved in the development of cancer, 6 , 7 CLs have been extensively studied in cancer gene therapy and have shown potential clinical success in different tumors ( Table 1 ). In general, the selection of CLs as gene carriers is attributed to their permanent positive charge, which promotes their stronger interaction with negatively charged substances such as nucleic acid, cell membrane, and endosomal membrane.…”

A Non-Viral Plasmid DNA Delivery System Consisting on a Lysine-Derived Cationic Lipid Mixed with a Fusogenic Lipid

“…It has been already reported that gene delivery systems formed by amino acid-based cationic lipids are internalized by habitual endocytic uptake pathways. Thus, the incorporation of amino acid moieties to the head groups of cationic lipids does not seem to change the cellular uptake mechanism, but it can improve the intracellular delivery [33,[42][43][44][45].…”

Perspective Chapter: Liposome Mediated Delivery of Immunotherapeutics for Cancer

“…With their positive charge, cationic liposomes, can be used to easily encapsulate plasmid DNA (pDNA), messenger RNA (mRNA), or small interfering RNA (siRNA) via electrostatic interactions [131]. An important example is the T7 peptide modified core-shell nanoparticles (named as T7-LPC/siRNA NPs).…”