Plasmid DNA gene therapy: studies with the human interleukin-2 gene in tumor cells in vitro and in the murine B16 melanoma model in vivo.
The plasmid DNA vector pVCL-1102 containing the coding sequence for the human IL-2 gene was evaluated for expression in tumor cells in vitro and in vivo. In vitro transfection of murine B16 tumor cells with pVCL-1102 resulted in the expression of 36,000 IU (5.7 micrograms) of biologically active IL-2/10(6) cells/48 h. In vitro transfection of human tumor lines and primary cultures from human biopsies with pVCL-1102 resulted in the expression of 1,289 to 9345 IU of IL-2/10(6) cells/48 h and 30 to 794 IU of IL-2/10(6) cells/48 h, respectively. In vivo, direct intratumor injection of pVCL-1102 resulted in retention of intact plasmid DNA in the tumor tissue and IL-2 secretion by cell cultures derived from the injected tumors. Formulation of pVCL-1102 with the cationic lipid DMRIE/DOPE inhibited DNA degradation and enhanced in vivo transfection efficiency over plasmid DNA alone. Antitumor activity of the pVCL-1102/DMRIE/DOPE complex was evaluated in a B16 melanoma model in mice. An IL-2-specific effect could not be demonstrated in a subcutaneous model because the intratumor injection of plasmid DNA lacking the IL-2 coding sequence also resulted in a significant reduction in tumor volume. However, an IL-2-specific effect was observed when B16 cells were transfected in vitro prior to implantation into the mouse. Transient transfection of B16 cells with pVCL-1102 rendered the cells less tumorigenic in vivo and produced a significant reduction in tumor volume. These data demonstrate that a plasmid DNA expression vector can be used to deliver the IL-2 gene to tumor cells in vitro and in vivo, resulting in the expression of significant levels of IL-2 protein. These data also illustrate the need for the use of appropriate controls when evaluating the in vivo biological activity of plasmid DNA in murine tumor models.
Parker
S E
SE
Vical, Inc., San Diego, CA 92121, USA.
Khatibi
S
S
Margalith
M
M
Anderson
D
D
Yankauckas
M
M
Gromkowski
S H
SH
Latimer
T
T
Lew
D
D
Marquet
M
M
Manthorpe
M
M
Hobart
P
P
Hersh
E
E
Stopeck
A T
AT
Norman
J
J
D000818:Animals; D015139:Blotting, Southern; D002460:Cell Line; D004247:DNA / Q000032:analysis; D017931:DNA Primers; D004337:Drug Carriers; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D006801:Humans; D007376:Interleukin-2 / Q000096:biosynthesis / Q000235:genetics*; D007700:Kinetics; D008081:Liposomes; D008546:Melanoma, Experimental / Q000276:immunology* / Q000628:therapy*; D051379:Mice; D008810:Mice, Inbred C57BL; D010957:Plasmids; D016133:Polymerase Chain Reaction; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D004337:Drug Carriers; D007376:Interleukin-2; D008081:Liposomes; D004247:DNA