Gene Delivery Using Physical Methods - Challenges in Delivery of Therapeutic Genomics and Proteomics - page 92

In another study, the efficiency of intrarenal injection of DNA followed by in vivo EP,
using FITC-labeled oligodeoxynucleotides (FITC-ODN) and plasmid DNA express-
ing -galactosidase or luciferase was studied. FITC-ODN or expression vectors were
injected into the left renal artery and thereafter the left kidney was electroporated
between a pair of tweezer-type electrodes. FITC-ODN was transferred into all glom-
eruli, and transfected cells were identified as mesangial cells. Four days after transfec-
tion of the pCAGGS-LacZ gene, -galactosidase expression was observed in 75% of
glomeruli [63] .
EP as a delivery method in cardiac tissue has also shown encouraging results,
with the embryonic chick heart showing uptake of propidium iodide (PI) or GFP and
luciferase DNA following electrical shock, which suggests further application of this
method for therapeutic genes [64] . The EP of murine factor VIII cDNA in gastrocne-
mius skeletal muscle of FVIII null mice in vivo was carried out for phenotypic cor-
rection with minimal tissue injury and significant DNA expression. The results were
confirmed by reduced partial thromboplastin time after EP, and all the hemophilic
mice survived hemostatic challenge as compared to 100% death in untreated mice
[65] . In another investigation, efficient gene delivery with mRNA using EP in human
hematopoietic cells showed superior results to those achieved with lipofection. The
mRNA EP of the tumor antigen showed markedly improved transfection efficiency
(89% cells), when compared with plasmid cDNA EP (40% cells), in the dendritic
cells for tumor vaccines and induced a strikingly lower cell toxicity (15% death rate
with mRNA versus 51% with plasmid DNA), hence presenting significant functional
antigenic peptides to cytotoxic T cells, proving their application in future tumor vac-
cines [66] .
Apart from the exciting advances made in transgene expression with EP in various
tissues, several major drawbacks have been observed to restrict the in vivo applications
and clinical utility of the method. The limited effective range of approximately 1 cm
between the electrodes makes it difficult to transfect cells in a large area of tissues
[67] . Also, the surgical procedure required for EP, implanting electrodes deep into the
Internal organs, has been a huge drawback. Tissue damage was observed because of
high voltage applied during EP, which causes thermal heating of cells, Ca 2 influx
induced protease activation, and instability of genomic DNA during high voltage [67] .
However, some of these concerns may be resolved by optimizing the EP conditions. In
conclusion, even though, EP has been known for approximately 25 years and proven
to be a significant contribution to effective gene delivery, further work is still required
to minimize the method toxicity without compromising gene expression level in order
for research to proceed with significant in vivo and clinical trials.
3.4 Hydrodynamic Gene Therapy
Efficient and safe methods for delivering genetic materials into cells must be devel-
oped before the clinical potential of gene therapy can be fully realized. Recently,
hydrodynamic gene delivery using a rapid injection of a relatively large volume of
DNA solution has opened up a new avenue for gene therapy studies in vivo [26] .