ISSN:
1573-904X
Keywords:
cationic lipid
;
DNA
;
in vivo
;
gene transfer
;
alkaline phosphatase
Source:
Springer Online Journal Archives 1860-2000
Topics:
Chemistry and Pharmacology
Notes:
Abstract Purpose. A stable cationic lipid/DNA complex has been developed for in vivo gene transfer. The formulation capitalizes on a previously described procedure to obtain stable lipid/DNA complexes for in vitro gene transfer (1). Methods. Conditions for DNA/lipid complex formation were modified to yield a DNA concentration of 1 mg/ml. Heat stable alkaline phosphatase (AP) under a CMV promoter was used as a reporter gene. Results. The resulting complex was completely insensitive to serum inactivation. Tail vein injection of a 80 μg DNA into Balb C mice yielded significant levels of reporter enzyme activity in the lung, heart, spleen, muscle, and liver. Less AP activity was observed in the kidney. No AP activity was observed in blood, bone marrow or brain. A titration of the lipid (DOSPA) to DNA-nucleotide ratio showed the optimal molar ratio for in vivo gene transfer to be 1/1. Using this ratio in a dose response study showed approximately 80 μg of DNA/mouse yielded the highest level of gene expression. Using this dose at a 1/1 lipid to DNA nucleotide ratio, the time course for alkaline phosphatase activity was determined. Maximal AP activity was observed 24 hours after injection for all tissues. By day 5, the activity dropped approximately 10 fold for all tissues. By day 7, residual activity was detected in the lung, heart, and muscle. Histology of the lung showed both interstitial and endothelial cells to be transfected. In all other tissues, however, endothelial cells were the only transfected cell type. Conclusions. These results demonstrate that reformulation of an existing cationic lipid can result in the formation of a stable lipid/DNA complex, which is able to reproducibly transfect lung, heart, spleen, and liver upon intravenous administration.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1023/A:1012146305040
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