A gene delivery approach for antimicrobials: expression of defensins.
BACKGROUND
Peptide antibiotics as new therapeutic agents are becoming a popular option to investigate due to their broad bacterial target selectivity and limited resistance problems. Although attractive, these new drug candidates have several limitations including low potency and delivery issues which face all peptides/proteins.


METHODS
In this study, we designed a plasmid expression system for human beta defensin 3. This sequence was cloned from a human epithelial lung cell into a CMV driven expression cassette. This expression plasmid was then evaluated for its ability to produce human-beta defensin 3 with the use of the non-viral transfection agent, polyethylenimine (PEI).


RESULTS
The results indicate the expression cassette was transcriptionally active in HEK 293 cells, as measured by RT-PCR and that a beta defensin peptide was produced by the cells as confirmed by Western blot. The biological activity of the peptide was confirmed against both gram negative E. coli and gram positive Bacillus species using in vitro screening.


CONCLUSIONS
Both the cultured media as well as the transfected cell lysate demonstrated biological activity demonstrating the peptide is also secreted.
Zhang
Chun
C
Department of Pharmaceutics, University of Florida, Gainesville, FL 32610, USA.
Yadava
Preeti
P
Sun
Jing
J
Hughes
Jeffrey A
JA
D000900:Anti-Bacterial Agents / Q000096:biosynthesis* / Q000494:pharmacology*; D015153:Blotting, Western; D002460:Cell Line; D003470:Culture Media; D023082:Defensins / Q000096:biosynthesis* / Q000494:pharmacology*; D016503:Drug Delivery Systems; D004591:Electrophoresis, Polyacrylamide Gel; D004847:Epithelial Cells / Q000378:metabolism; D004926:Escherichia coli / Q000187:drug effects; D005079:Excipients; D015316:Genetic Therapy; D005822:Genetic Vectors; D006801:Humans; D011094:Polyethyleneimine; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D023083:beta-Defensins / Q000096:biosynthesis / Q000235:genetics
D000900:Anti-Bacterial Agents; D003470:Culture Media; C422950:DEFB103A protein, human; D023082:Defensins; D005079:Excipients; D023083:beta-Defensins; D011094:Polyethyleneimine