Diversity of PEGylation methods of liposomes and their influence on RNA delivery.

Gene therapy is a promising approach for personalized medicine, but its application in humans requires development of efficient and safe vehicles. PEGylated liposomes are some of the most suitable delivery systems for nucleic acids because of their stability under physiological conditions and prolonged circulation time, compared to conventional and other types of "stealth" liposomes. In vitro/in vivo activity of PEGylated liposomes is highly dependent on PEG motif abundance. The process of "stealth" coverage formation is a very important parameter for efficient transfection assays and further fate determination of the PEG layer after tissue penetration. In this review, we discuss the latest methods of PEGylated liposome preparation.

Effect of lipopeptide structure on gene delivery system properties: Evaluation in 2D and 3D in vitro models.

Development of efficient biodegradable, environmentally responsive, biocompatible and non-toxic delivery system is needed for efficient gene delivery. As well known, properties of the vehicle are determined by the structure of carrier components. The aim of the current study was to estimate in vitro transfection efficacy of aliphatic di-, tri- and tetrapeptide-based cationic lipoplexes loaded with siRNA in function of a number of cationic groups using 2D (monolayer culture) and 3D (multicellular tumor spheroids) in vitro models. Physicochemical properties and cytotoxicity of the liposomes were found to be dependent upon a number of amino acid derivatives in an amphiphilic polar head. Uptake of liposomes loaded with nucleic acid (lipoplexes) and their localization in HEK293T cells was studied by confocal microscopy. The liposomes based on lipotripeptides had the highest transfection efficiency which was 20-fold higher than those fabricated from lipotetrapeptides.

[Lactose-based glycoconjugates with variable spacers for design of liver-targeted liposomes]. / Glikokon"iugaty na osnove laktozy so speiserami razlichnoi dliny dlia sozdaniia transportnykh sistem k kletkam pecheni.

Asialoglycoprotein receptors are highly abundant on the hepatocyte surface and have specific binding sites for blood serum glycoproteins. Such discovery resulted in development of liver-targeted drug delivery systems because modification of the liposomal surface by carbohydrate derivatives results in an increase of endocytosis, which facilitates selective uptake of such systems by hepatocytes. In this study we have synthesized novel lactose derivatives containing a palmitic hydrophobic domain. They were used for modification of the liposome surface. Transfection activity of modified liposomes was analyzed on the HepG2 cell line (hepatocytes) and showed an increase in the transfection efficiency as compared to the non-modified liposomes. At the same time transfection activities of modified and non-modified liposomes were similar in the case of a non-hepatocyte cell line (293T). The novel lactose-based glycoconjugates may be a promising tool for developing efficient vectors for delivery of nucleic acids to hepatocytes.

[Physicochemical properties of lipopeptide-based liposomes and their complexes with siRNA]. / Fiziko-khimicheskie svoistva kompleksov molekul miRNK i liposom na osnove lipopeptidov.

siRNA/cationic liposome complexes are efficient systems for transmembrane delivery. The aim of this study was to prepare a novel complex consisted of lipotripeptide OrnOrnGlu(C16H33)2 and siRNA molecule and examined their physicochemical properties. Electron microscopy study has shown that the siRNA/liposome complex (m/m 1/10) tends to form sandwich-like structures that may protect nucleic acid from nuclease degradation. Photon correlation spectroscopy data indicate that the particle size increased after siRNA adding, but did not exceed 300 nm in diameter, while z-potential of lipoplexes decreased from 22 mV to 14 mV, compared to the empty liposomes thus indicating positive charge neutralization by negatively charged siRNA. These data allow to hypothesize that such size and total positive charge could provide efficient cellular uptake by endocytosis. That may have good prospects for gene silencing therapy.

Synthesis and evaluation of novel lipopeptide as a vehicle for efficient gene delivery and gene silencing.

Nucleic acid-based therapeutics have recently emerged as a new class of next generation agents for treatment and prevention of viral infection, cancer, and genetic disorders, but their wide use is limited by their relatively weak delivery into target cells. Usage of synthetic cationic amphiphiles with peptide hydrophilic domain as agents for non-viral gene delivery is an attractive approach. We developed the schemes for the synthesis of aliphatic peptides with different length of the hydrocarbon chains in hydrophobic domains and different amino acids in polar head. For the obtained derivatives we determined transfection efficiency, critical vesicle concentration, particle size, ζ-potential and aggregates stability. We have found that the transfection efficiency is increased if the ornithine is a part of polar head in an amphiphile. The most promising amphiphile for liposomal formation OrnOrnGlu(C16H33)2 was examined more carefully. It has been shown that the lipopeptide possesses low toxicity (in vitro and in vivo) and high transfection efficiency with pDNA and siRNA in different cell lines. In addition, the production of liposomes based on this lipopeptide is simple, quick and cheap. Thus OrnOrnGlu(C16H33)2 is a promising vehicle for gene delivery and gene silencing.