Significant Antitumor Effects Obtained by Autologous Tumor Cell Vaccine Engineered to Secrete Interleukin (IL)-12 and IL-18 by Means of the EBV/Lipoplex

Abstract

The EBV/lipoplex is a nonviral gene delivery system composed of a cationic lipid and Epstein-Barr virus (EBV)-based plasmid vector that carries the EBV oriP and EBV nuclear antigen 1 (EBNA1) gene. Because the EBNA1 supports retention, nuclear localization, and transcriptional upregulation of the oriP-bearing plasmid, cells transfected with the EBV/lipoplex express the transgene at a very high level. We hypothesized that tumor cells genetically manipulated with the EBV/lipoplex may be used as a tumor vaccine without drug selection, strongly contributing to immunotherapy of patients with malignancies. The cytokines interleukin (IL)-12 and IL-18 exert a variety of immune-regulatory functions including interferon (IFN)-gamma production and cytotoxic T lymphocyte (CTL) and natural killer (NK) activation. Here, we investigated the possible therapeutic effects of an autologous tumor cell vaccine in the B16 melanoma model. The vaccine was engineered to secrete IL-12 and IL-18 by means of the EBV/lipoplex. B16 cells were subcutaneously implanted into syngenic mice followed by repetitive immunization with irradiated B16 cells that had been transfected 3 days earlier by TFL2-3, a novel cationic lipid, with EBV-plasmid vectors encoding IL-12 and/or IL-18 genes (B16/mIL-12, B16/mIL-18, and B16/mIL-12+mIL-18). The mice vaccinated with B16/mIL-12 underwent strong tumor suppression accompanied by a high IFN-gamma production. Both CTL and NK activities were significantly elevated in these mice. When the tumor cell vaccine was prepared by means of conventional (non-EBV) plasmid vectors combined with the same cationic lipid, the therapeutic outcome was not as good, suggesting the superiority of the EBV-based plasmid in engineering these types of tumor vaccines. Vaccination with B16/mIL-18 was not effective in suppressing tumors, whereas B16/mIL-12+mIL-18 showed comparable antitumor therapeutic validity as B16/mIL-12 did. When IFN-gamma mutant (IFN-gamma(-/-) mice were treated, B16/mIL-12 vaccine did not show any therapeutic activity, suggesting the necessity of IFN-gamma in the anti-melanoma immune responses. In contrast, the antitumor effect was not affected by NK depletion in mice that received repetitive injections with anti-asialo GM1 antibody. Furthermore, vaccination with B16/mIL-12 significantly suppressed pulmonary metastases in mice that had been intravenously injected with parental B16. Our results suggest that the EBV/lipoplex is quite useful in generating an autologous tumor cell vaccine and that IL-12 is an important component of the vaccine.

Nonviral genetic transfer of Fas ligand induced significant growth suppression and apoptotic tumor cell death in prostate cancer in vivo

“…18,19 Compared with conventional polyplex and lipoplex systems, EBV-based plasmid vectors conjugated with cationic polymers and cationic lipids (EBV/polyplex and EBV/lipoplex, respectively) achieve quite successful genetic delivery, resulting in strong and persistent transgene expression in various tumor cells both in vitro [6][7][8][9][10] and in vivo. 8,10,11 Fas ligand (FasL) is a type 2 transmembrane glycoprotein belonging to the tumor necrosis factor superfamily. 20 FasL leads to trimerization of Fas, the cell surface apoptotic signal molecule, and triggers sequential activation of cysteine proteases, subsequently inducing apoptosis in Fas-positive cells.…”

Complete tumor prevention by engineered tumor cell vaccines employing nonviral vectors

“…1,15,[35][36][37] In contrast, little work has been reported thus far in the case of cell vaccines engineered by nonviral vectors to secrete cytokines. 38 Our results of vaccination with cells producing GM-CSF offer expectations regarding the promising results obtained by applying nonviral gene therapy, which aims to afford safer therapy despite comparatively lower efficacy. The present study was designed to establish the antitumor efficacy after vaccination of two different cytokines, GM-CSF and IL-4, in genetically modified tumor cells, as cell clones or freshly transfected cells.…”

Clinically feasible approaches to potentiating cancer cell-based immunotherapies

“…31 Vaccination with tumor cells genetically modified for IL-12 production (a key cytokine boosting Th1 immunity) has been shown to cause a significant tumor suppression in mice paralleled with high IFN-g production and activation of CTL and NK cells. 12,39 Normally, tumor cells are devoid of molecules that could co-stimulate TAA-specific T cells. Therefore, transduction of tumor cell vaccines with co-stimulatory molecules such as B7-1 represents another possible approach to potentiate antitumor immune responses.…”