Studies of direct intratumoral gene transfer using cationic lipid-complexed plasmid DNA

Abstract

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% (Ϯ 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. Cancer Gene Therapy (2000) 7, 853-860

Electrotransfer of gene encoding endostatin into normal and neoplastic mouse tissues: Inhibition of primary tumor growth and metastatic spread

“…This method of transfer of a foreign gene is simple and the plasmid DNA may be transferred repeatedly into any type of tissue. The efficiency of transfer, although lower than that with viral vectors, is nonetheless higher than that offered by nonviral lipoplexes whose efficiency of transfer into tumors does not exceed 1.7%, 30 whereas electroporation can yield 3 -8% of transfected tumor cells 31 and up to 10% of muscle cells 32 ( in our hands, however, transfection efficiency of muscle cells was 3% ). In order to successfully adapt electroporation to gene transfer into tissues and to exclude large interexperimental variability, several parameters must be optimized for a given organ or tissue including voltage, number and duration of impulses, distance between electrodes, and so forth ( Table 1).…”

Suicide gene and cytokines combined nonviral gene therapy for spontaneous canine melanoma

“…GCV (5 mg mg À1 DNA) was then added and lipoplexes were injected as described below. 9 Patients Dogs included in this study had confirmed histopathological diagnosis of melanoma. The tumors were staged by a veterinary oncologist according to the WHO staging system of stage II (tumors 2-4 cm diameter, negative nodes), stage III (tumor 44 cm and/or positive lymph nodes) or stage IV (distant metastatic disease).…”

“…In vivo intratumor injections of lipid-complexed plasmid DNA 9 encoding the herpes simplex thymidine kinase suicide gene (SG) should sensitize transfected cells to ganciclovir. 10,11 Due to the bystander effect, this SG-triggered cytotoxic effect allows substantial tumor regression by transfection of only a minority of tumor cells.…”

Cancer Immunogene Therapy: A Review

“…The earliest clinical trial results were, not surprisingly, based on the first strategies examined in preclinical models. For example, direct intratumoral HLA-B7 (MHC class I) gene transfer via liposome/plasmid DNA complex injection has been studied relatively extensively in phase I clinical trials in metastatic melanoma, colonic adenocarcinoma, and renal cell carcinoma patients [11,45,55,69]. All of these trials demonstrated safety and most included at least some patients that appeared to have clinical responses to treatment.…”