Biomedical Engineering Reference
In-Depth Information
systems currently employed for gene therapy, including lipid and polymer-based
vectors, nanomaterials (e.g., magnetic nanoparticles, quantum dots, gold/silica
nanostructures, carbon nanotubes, calcium phosphate nanoparticles, and layered
double hydroxides/clays) vehicles, as well as multifunctional nano-systems among
them. Particular emphasis has been laid on synthetic polymers and the related deli-
very systems. Selective clinical trials of gene therapy using nonviral vectors were
briefly reviewed as well.
2
Gene Delivery Barriers
To achieve successful transgene expression, a series of extracellular and intracel-
lular transport barriers such as DNA protection, internalization, intracellular traf-
ficking and nuclear transport, need to be overcome by delivery vectors. Viral vectors
have already showed their great success in addressing each challenge. Synthetic
vectors, however, lack one or several of the necessary functions. Understanding the
important barriers encountered by delivery vectors is a prerequisite to design and
discover more efficient carriers for gene therapy.
2.1
Gene Packaging
Whereas the naked DNA can be introduced into cells via electrotransfer (Andre and
Mir 2004 ), gene gun inoculation (Fynan et al. 1993a ), direct injection into target
tissue (Wolff et al. 1990 ), or hydrodynamic injection (Zhang et al. 1999 ), their
inherent disadvantages largely limit the translation of these procedures to the clinical
practice. Packaging therapeutic genes can protect them from degradation by nucle-
olytic enzymes, and condense their bulky structures to facilitate cellular
internalization.
Complexation of DNA with most of the synthetic cationic vectors is based on
electrostatic forces between negative phosphates along DNA and positive charges
displayed on vector materials. This electrostatically mediated self-assembly con-
denses DNA into small, compact structures. The process of condensation is consid-
ered to be entropically driven (Bloomfield 1997 ). The size and morphology of
resulting particles are dependent on the type and structure of cationic materials
used, and preparation conditions such as DNA concentration, type and pH of buffer
and N/P ratio. In the case of cationic lipids, most of the different techniques of
preparing liopsomes have been applied, such as hydration of a lipid film, dehydra-
tion-rehydration, ethanolic injection, reverse-phase evaporation, or the detergent
dialysis technique (de Ilarduya et al. 2010 ). The obtained liposomes are then mixed
with DNA to form lipoplexes. In this step, the component concentration, tempera-
ture, and kinetics of mixing should be carefully selected to control the formation of
lipoplexes and their physicochemical characters that may influence the final
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