Challenges and Opportunities in Gene Delivery - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 47

Although “naked” DNA on direct local delivery by injection at site of action has
shown a transgene expression in certain physiological tissues such as skeletal muscle
[23] , liver, [24-26] , skin [27,28] , or airway instillation into the lungs [29] , many tis-
sues in the body still require an additional delivery system to facilitate transfection.
The barriers to “naked” DNA delivery involve DNA degradation by endogenous
nuclease enzymes in extracellular spaces, poor penetration because of high hydro-
philicity, hepatic first-pass metabolism, and a highly negatively charged phosphate
backbone of DNA, which hinders the passage across negatively charged cellular mem-
branes [25] . Hence, development of suitable delivery system with effective and non-
toxic delivery of the DNA at the desired site of action after local or systemic delivery
is a key challenge and serves as the most significant barrier between the DNA tech-
nology and its therapeutic application.
Systemic in vivo gene delivery can be visualized as a series of intracellular and
extracellular barriers and challenges that successively deplete the mass of DNA that
progresses toward the nucleus of the target site. Hence, there is an urgent need to
target the DNA to the desired target site within the target cells using a competent
vector system. The in vivo gene delivery may be achieved by naked DNA deliv-
ery and by using viral- and nonviral-mediated-gene transfer delivery systems [18] .
Because of their natural ability to infect cells efficiently, viral vectors such as retro-
virus, adenovirus, adeno-associated virus, and herpesvirus, with part of their coding
sequences replaced by that of a therapeutic gene, have been investigated for in vivo
viral-mediated gene delivery [30] . These vectors can be extremely efficient at pro-
ducing expression, with essentially only a single viral particle necessary to induce a
measurable effect. However, clinical applications of these vectors are hampered by
the barriers of viral immunogeneicity, inability to transfect nondividing cells, pos-
sible oncogenecity, and strong toxicity. Because of these significant barriers, the
nonviral gene delivery vectors evolved. The nonviral vectors involve plasmid DNA
delivery complexed to synthetic carrier molecules, such as positively charged lipids,
polymers, peptides, and inorganic materials [15,18,22] . Although these vectors are
comparatively less cytotoxic, with little or no immune response, and show a con-
trolled and reproducible transgene expression, their clinical utility is hampered by
the barrier of low transfection efficiency compared to viral vectors and transient gene
expression, thus causing a need of repeated gene administration [18] .
For transfecting a single cell, about 10 6 plasmid copies must be injected, with or with-
out vector, out of which around 10 2 -10 4 copies reach the nucleus for transgene expres-
sion [31,32] . Such reduced efficiency of gene delivery can be attributed to the barriers
encountered by the DNA and the DNA-vectors complexes between the site of admin-
istration and localization in the cell nucleus. DNA delivery systems presently used have
shown maximum transgene expression at local site (disease) of application [23-29] or
when administered in the vicinity of the site of target cells [8,23,24,26] . Insignificant
transgene expression at target tissue after systemic (i.e., intravenous or even intramuscu-
lar delivery), and the need to deliver these agents locally near target cells in order to elicit
a therapeutic effect, are some of the hurdles attributable to the barriers encountered dur-
ing successful gene delivery.