Biomedical Engineering Reference
In-Depth Information
(both blood cells and vascular endothelial cells) and serum proteins. Also, following i.v. injec-
tion, such DNA complexes in practice tend to accumulate in the lung and liver. Targeting of DNA
complexes to specifi c cell types also poses a considerable (largely unmet) technical challenge.
Approaches, such as the incorporation of antibodies directed against specifi c cell surface antigens
may provide a future avenue of achieving such cell-selective targeting. However, it is currently
believed that ionic interactions constitute a predominant binding force between the positively
charged lipoplexes/polyplexes and the negatively charged eukaryotic cell surface. Such electro-
static interactions may even override more biospecifi c interactions characteristic of antibody- or
receptor-based systems. Currently, probably the most effective means of delivering such vectors to
target tissue/cells is to inject them into/beside the target area.
However targeted to the appropriate cell surface, if it is to be clinically effective, the therapeutic
plasmid must enter the cell and reach the nucleus intact. Cellular entry is generally achieved via
endocytosis (Figure 14.10). A proportion of endocytosed plasmid DNA escapes from the endo-
some by entering the cytoplasm, thereby escaping liposomal destruction (Figure 14.10). The mo-
lecular mechanism by which escape is accomplished is, at best, only partially understood. Anionic
lipid constituents of lipoplexes, for example, may fuse directly with the endosomal membranes,
facilitating direct expulsion of at least a portion of the plasmid DNA into the cytoplasm. Generally,
the DNA is released in free form (i.e. uncomplexed to any lipid).
Some attempts have been made to rationally increase the effi ciency of endosomal escape. One
such avenue entails the incorporation of selected hydrophobic (viral) peptides into the gene deliv-
ery systems. Many viruses naturally enter animal cells via receptor-mediated endocytosis. These
viruses have evolved effi cient means of endosomal escape, usually relying upon membrane-
disrupting peptides derived from the viral coat proteins.
Figure 14.10
Over view of cellular entr y of (non-viral) gene deliver y systems, with subsequent plasmid relocation
to the nucleus. The delivery systems (e.g. lipoplexes and polyplexes) initially enter the cell via endocytosis (the
invagination of a small section of plasma membrane to form small membrane-bound vesicles termed endosomes).
Endosomes subsequently fuse with golgi-derived vesicles, forming lysosomes. Golgi-derived hydrolytic lysosomal
enzymes then degrade the lysosomal contents. A proportion of the plasmid DNA must escape lysosomal destruction
via entry into the cytoplasm. Some plasmids subsequently enter the nucleus. Refer to text for further details
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