Development of improved vectors for DNA-based immunization and other gene therapy applications.
Optimizing gene expression and delivery are necessary steps in the production of vectors for DNA-based immunization as well as for other gene therapy applications. A mouse muscle/reporter gene assay system was used to systematically improve a plasmid DNA vector. The optimized vector VR1255 contained: (1) CMV promoter and enhancer; (2) CMV IE Intron A; (3) kanamycin resistance gene; (4) deleted SV40 origin of replication; (5) optimized lux coding region; and (6) a minimal synthetic terminator from the rabbit beta globin gene, mRBG. The vector VR1255 expressed 137 times greater than an earlier prototype RSV-based vector. For plasmid vector delivery into nonmuscle tissues, a recently synthesized cationic lipid, GAP-DLRIE, was found to greatly enhance the uptake and expression of plasmid DNA by 100-fold when instilled into the mouse lung. The time-course of CAT expression with GAP-DLRIE indicated that peak expression occurs 2-5 days after intranasal administration and expression diminished to about one-third the peak value by day 21. This cationic lipid may be useful for immunization by pulmonary and perhaps other nonmuscle routes.
Norman
J A
JA
Vical Inc. San Diego, CA 92121, USA.
Hobart
P
P
Manthorpe
M
M
Felgner
P
P
Wheeler
C
C
D000818:Animals; D003587:Cytomegalovirus / Q000235:genetics; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D010957:Plasmids / Q000235:genetics; D011817:Rabbits; D014611:Vaccination / Q000379:methods*; D019444:Vaccines, DNA / Q000008:administration & dosage; D014765:Viral Vaccines / Q000008:administration & dosage
D019444:Vaccines, DNA; D014765:Viral Vaccines