Cationic liposome-DNA complexes (lipoplexes) constitute a potentially viable alternative to viral vectors for the delivery of therapeutic genes. This review will focus on various parameters governing lipoplex biological activity, from their mode of formation to in vivo behaviour. Particular emphasis is given to the mechanism of interaction of lipoplexes with cells, in an attempt to dissect the different barriers that need to be surpassed for efficient gene expression to occur. Aspects related to new trends in the formulation of lipid-based gene delivery systems aiming at overcoming some of their limitations will be covered. Finally, examples illustrating the potential of cationic liposomes in clinical applications will be provided.

Antibody-directed liposomes (immunoliposomes) are frequently used for targeted drug delivery. However, delivery of large biotherapeutic molecules (i.e. peptides, proteins, or nucleic acids) with immunoliposomes is often hampered by an inefficient cytosolic release of entrapped macromolecules after target cell binding and subsequent endocytosis of immunoliposomes. To enhance cytosolic drug delivery from immunoliposomes present inside endosomes, a pH-dependent fusogenic peptide (diINF-7) resembling the NH 2 -terminal domain of influenza virus hemagglutinin HA-2 subunit was used. Functional characterization of this dimeric peptide showed its ability to induce fusion between liposome membranes and leakage of liposome-entrapped compounds when exposed to low pH. In a second series of experiments, diINF-7 peptides were encapsulated in immunoliposomes to enhance the endosomal escape of diphtheria toxin A chain (DTA), which inhibits protein synthesis when delivered into the cytosol of target cells. Immunoliposomes targeted to the internalizing epidermal growth factor receptor on the surface of ovarian carcinoma cells (OVCAR-3) and containing encapsulated DTA did not show any cytotoxicity toward OVCAR-3 cells. Cytotoxicity was only observed when diINF-7 peptides and DTA were co-encapsulated in the immunoliposomes. Thus, diINF-7 peptides entrapped inside liposomes can greatly enhance cytosolic delivery of liposomal macromolecules by pH-dependent destabilization of endosomal membranes after cellular uptake of liposomes.To exert an optimal therapeutic effect, an administered drug must safely reach not only its target cell but also the appropriate location within that cell. Many biotherapeutic agents (e.g. peptides, proteins, and nucleic acids) act at sites in the cytosol or nucleus that are difficult to reach due to poor membrane permeation characteristics. Therefore, these biotherapeutic agents rely on drug carriers that allow cytosolic delivery of these agents into diseased cells.One attractive strategy to obtain cytosolic delivery is the development of fusion-competent antibody-directed liposomes (immunoliposomes) that, after receptor binding, can fuse with the plasma-membrane of target cells or, after endocytosis of liposomes, with the endosomal membranes. This membrane fusion process should result in cytosolic access of liposomeentrapped drug molecules. Such fusion-competent liposomes can be obtained by disassembling enveloped viruses, isolating their fusion-promoting components, and reassembling these into vesicles to obtain so-called virosomes (1-4). Another, more refined approach involves the use of synthetic peptides designed to resemble the putative fusion peptide of viruses (5-8).It is hypothesized that destabilization of endosomal membranes is less damaging than plasma membrane destabilization. The latter may result in cytotoxicity through eradication of the electrochemical potential generated by the asymmetric distribution of ions across the plasma membrane. Therefore, drug delivery into the cytosol of target ce...

The use of an efficient carrier for nucleic acid-based medicines is considered to be a determinant factor for the successful application of gene therapy. The drawbacks associated with the use of viral vectors, namely those related with safety problems, have prompted investigators to develop alternative methods for gene delivery, cationic lipid-based systems being the most representative. Despite extensive research in the last decade on the use of cationic liposomes as gene transfer vectors and the development of elegant strategies to enhance their biological activity, these systems are still far from being viable alternatives to the use of viral vectors in gene therapy. In this review considerations are made regarding the structure-activity relationships of cationic liposome/DNA complexes and the key formulation parameters influencing the features of lipoplexes are presented and discussed in terms of their effect on biological activity. Particular emphasis is given to the interaction of the lipoplexes with serum components as well as to novel strategies developed to circumvent difficulties that may emerge upon iv administration of the complexes. Finally, since the ability of the lipoplexes to be stored while preserving their transfection activity is a crucial issue for the repeated use of such carriers, approaches reported on the improvement of their physical stability are also reviewed.

The successful application of gene therapy depends highly on understanding the properties of gene carriers and their correlation with the ability to mediate transfection. An important parameter that has been described to improve transfection mediated by cationic liposomes involves association of ligands to cationic liposome-DNA complexes (lipoplexes). In this study, ternary complexes composed of 1,2-dioleoyl-3-(trimethylammonium) propane:cholesterol, plasmid DNA and transferrin (Tf, selected as a paradigm of a ligand) were prepared under various conditions, namely, in medium with different ionic strengths (HEPES-buffered saline [HBS] or dextrose), at different lipid/DNA (+/-) charge ratios and using different modes for component addition. We investigated the effect of these formulation parameters on transfection (in the absence and presence of serum), size of the complexes, degree of DNA protection and extent of their association with cells (in terms of both lipid and DNA). Our results show that all the tested parameters influenced to some extent the size of the complexes and their capacity to protect the carried genetic material, as well as the levels of cell association and transfection. The best transfection profile was observed for ternary complexes (Tf-complexes) prepared in high ionic strength solution (HBS), at charge ratios close to neutrality and according to the following order of component addition: cationic liposomes-Tf-DNA. Interestingly, in contrast to what was found for dextrose-Tfcomplexes, transfection mediated by HBS-Tf-complexes in the presence of serum was highly enhanced.

PurposeThe aim of this study is to examine the impact of abusive supervision on employees' emotional and work-related outcomes, using a theoretical framework that integrates affective events theory (AET) and self-determination theory (SDT). The research sought to explore the effects of abusive supervision on subordinates' positive and negative affect, and the subsequent impact on customer orientation and life satisfaction.Design/methodology/approachThe data for this study were collected cross-sectionally through a structured questionnaire completed by employees who have experienced abusive supervision in their current or previous jobs. Structural equation modeling (SEM) was used to analyze the data.FindingsThe results indicated that when subordinates experienced abusive supervision, they reported lower levels of positive affect and higher levels of negative affect. These emotions, in turn, impacted their customer orientation and life satisfaction. Positive affect was found to positively influence both customer orientation and life satisfaction, while negative affect had a negative effect on life satisfaction. Surprisingly, customer orientation was positively impacted by negative affect.Originality/valueTherefore, the findings of this study suggest that positive and negative affects mediate the relationship between abusive supervision and life satisfaction, but not with customer orientation. This study advances prior research by linking the impact of an abusive supervisor to employees' customer behavior and life satisfaction, using positive and negative affects as mediators, and building upon the theories of AET and SDT.