Studies of direct intratumoral gene transfer using cationic lipid-complexed plasmid DNA.
Cationic lipid-mediated gene transfer is a safe and effective means of delivering potent immunomodulatory cytokines directly into tumors. This approach avoids undesirable side effects, including systemic toxicities. To investigate key factors affecting intratumoral (i.t.) gene transfer, cationic lipid-DNA complexes were injected into subcutaneous human melanoma tumors in severe combined immunodeficient mice. Animals received i.t. injections of VR1103, a DNA plasmid encoding the gene for human interleukin-2 (IL-2), either alone or complexed with the cationic lipid N-(1-(2,3-dimyristyloxypropyl)-N,N-dimethyl-(2-hydroxyethyl) ammonium bromide/dioleoyl phosphatidylethanolamine (DMRIE/DOPE). Tumors were subcultured and supernatants were tested for IL-2 secretion by enzyme-linked immunosorbent assay. IL-2 secretion was consistently higher when lipid:DNA (L:D) complexes were formulated at high L:D ratios (wt/wt), and IL-2 transgene expression increased in a DNA dose-dependent manner. A comparison of naked plasmid and lipid-complexed DNA revealed that lipid complexes were more effective for i.t. gene transfer. Using an enhanced green fluorescent protein reporter plasmid and flow cytometry, i.t. transfection efficiency was 1.74% (+/- 1.08%). Tumor injection technique, including injection volume and location, had a limited impact on i.t. gene transfer. These results indicate that the formulation and dosage of cationic L:D complexes, but not injection technique, play a key role in determining the level of i.t. transgene expression.
Clark
P R
PR
Arizona Cancer Center, University of Arizona, Tucson 85724, USA.
Stopeck
A T
AT
Ferrari
M
M
Parker
S E
SE
Hersh
E M
EM
D000818:Animals; D004247:DNA / Q000378:metabolism*; D005434:Flow Cytometry; D015316:Genetic Therapy; D020404:Glycerophospholipids / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D006801:Humans; D007167:Immunotherapy; D007376:Interleukin-2 / Q000096:biosynthesis / Q000235:genetics; D050356:Lipid Metabolism; D008055:Lipids; D008164:Luminescent Proteins / Q000378:metabolism; D008545:Melanoma / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D016513:Mice, SCID; D008822:Mice, Transgenic; D010714:Phosphatidylethanolamines; D010957:Plasmids / Q000235:genetics*; D000644:Quaternary Ammonium Compounds / Q000378:metabolism*; D012878:Skin Neoplasms / Q000235:genetics* / Q000378:metabolism; D014162:Transfection / Q000379:methods*; D014407:Tumor Cells, Cultured
C410262:1,2-dioleoyl-glycero-3-phosphatidyl ethanolamine; D020404:Glycerophospholipids; D007376:Interleukin-2; D008055:Lipids; D008164:Luminescent Proteins; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D049452:Green Fluorescent Proteins; C087276:(3-dimyristyloxypropyl)(dimethyl)(hydroxyethyl)ammonium; D004247:DNA