Cancer gene therapy by IL-12 gene delivery using liposomal bubbles and tumoral ultrasound exposure.
ABSTRACT Interleukin-12 (IL-12) gene therapy is expected to be effective against cancers because it primes the immune system for cancer cells. In this therapy, it is important to induce IL-12 gene expression in the tumor tissue. Sonoporation is an attractive technique for developing non-invasive and non-viral gene delivery systems, but simple sonoporation using only ultrasound is not an effective cancer gene therapy because of the low efficiency of gene delivery. We addressed this problem by combining ultrasound and novel ultrasound-sensitive liposomes (Bubble liposomes) which contain the ultrasound imaging gas perfluoropropane. Our previous work showed that this is an effective gene delivery system, and that Bubble liposome collapse (cavitation) is induced by ultrasound exposure. In this study, we assessed the utility of this system in cancer gene therapy using IL-12 corded plasmid DNA. The combination of Bubble liposomes and ultrasound dramatically suppressed tumor growth. This therapeutic effect was T-cell dependent, requiring mainly CD8(+) T lymphocytes in the effector phase, as confirmed by a mouse in vivo depletion assay. In addition, migration of CD8(+) T cells was observed in the mice, indicating that the combination of Bubble liposomes and ultrasound is a good non-viral vector system in IL-12 cancer gene therapy.
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ABSTRACT: Interleukin-12 (IL-12) has long been considered to be effective in triggering an anticancer immune response, however, the dosage has been limited by potential systemic immunotoxicity. Since focused ultrasound (FUS) has been confirmed to temporally and locally open the blood-brain barrier (BBB), the purpose of this study was to elucidate the possibility of combining FUS-induced BBB opening with IL-12 delivery to enhance the anticancer immunological response for glioma treatment. FUS energy combined with microbubble administration was delivered transcranially to open BBB, and C-6 glioma rats were used in this study. The efficacy in inducing BBB opening and the corresponding immunological response were primarily evaluated in normal animals. The anticancer immune-triggering chemokine, IL-12, was intraperitoneally administered during the treatment phase to evaluate the effect of immunological response on tumor progression. Glioma animals were sub-grouped to evaluate the effect of the immune response in suppressing glioma when IL-12 was combined with FUS-induced BBB opening. We performed flow cytometry to verify consequent immune cell population changes of peripheral/ tissue lymphocytes as well as macrophages from the animals. Brain sections of sacrificed animals were also used for histological and immunohistochemical analysis. IL-12 level among experimental groups were measured via ELISA analysis. We also analyzed survival and followed tumor progression in vivo via T2-weighted magnetic resonance imaging. FUS-induced BBB opening had no obvious effect on the T lymphocytes population in normal animals, either in the brain or systemically. Yet, it triggered mild changes in the tumor-infiltrating lymphocyte (TIL) population, particularly in numbers of CD3 + CD8+ cytotoxic T lymphocytes (CTLs) in the tumor region. IL-12 administration triggered a profound increase in all TIL populations, including CD3 + CD4+ T helper cells (Th), CTL, and CD4 + CD25+ regulatory T cells (Treg), but combined FUS-BBB opening with IL-12 administration produced the most significant IL-12 increase, CTL increase and CTL/Treg ratio increase, thus contributing to the most significant suppression of tumor progression and increased animal survival. This study provides evidence that FUS-BBB opening can enhance immune-modulating agent delivery to the brain, which improve the anticancer immune response in brain tumor treatment.Journal of Translational Medicine 12/2015; 13(1):451. DOI:10.1186/s12967-015-0451-y · 3.99 Impact Factor
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ABSTRACT: Cancer immunotherapy attempts to stimulate the immune system to reject and destroy tumors and is one of the cancer treatment strategies. Recently, interluekin36 (IL36) has been used as immunotherapeutic agents in cancer gene therapy. Present study investigated that the IL36 gene therapy effects on the regression of tumor masses in mouse model. Aim of this study is determination of the gene therapy effects by IL36 in the regression of tumor masses in mouse model. To study the therapeutic efficacy of this cytokine, WEHI-164 tumor cells were transected with mIL36 plasmids. ELISA test was used to check cytokine production by transected cells. To establish fibro sarcoma mouse model, Tumoral transfected cells were injected subcutaneously to inoculate tumor in BALB/C mice. Tumor volumes were measured by caliper. Mice were sacrificed and tumors were extracted. The expression of IL36 and IFN-γ was studied with Real-time PCR and immunoblotting. The expression of Ki-67 (a tumor proliferation marker) in tumor masses was studied by immunohistochemistry staining. In this study we had 2 groups which are treated with IL-36 and Untreated with IL-36 as a blank. The group treated with IL36 indicated decrease of tumor mass volume (p<0.001). The results of western blotting and real-time PCR showed the IL36 expression increased in the group treated with IL36 (with relative expression of 1.9). Immunohistochemistry staining indicated that the Ki-67expression has been reduced in the group interfered with IL36. IL36 gene therapy has therapeutic effects on the regression of tumor masses in fibro sarcoma mouse model.Iranian Journal of Cancer Prevention 09/2014; 7(4):197-203.
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ABSTRACT: MMP-9 participates in tumor growth, invasion, metastasis and vascularization. Thus, inhibition of MMP-9 may be involved in the process of tumorigenesis. We have investigated the effect of RNAi-mediated MMP-9 silencing on inhibiting invasion and migration of mouse melanoma cell B16. A specific and optimized siRNA vector was used to target MMP-9 mRNA synthesis in B16 cells. Changes of invasion and migration capability of B16 cell were examined after transfection at different time, and a footpad tumor model was performed to measure the effect of MMP-9 siRNA on melanoma tumorigenesis in vivo. In vitro, down-regulation of MMP-9 expression significantly inhibited B16 cell invasion and migration. In vivo, intratumoral injection of plasmid DNA expressing MMP-9 siRNA every 3 days for 3 times remarkably inhibited melanoma growth and also suppressed tumor metastasis. The results indicate that RNA mediated targeting of MMP-9 may have promising applications for the treatment of melanoma.Cell Biology International 08/2013; 37(8). DOI:10.1002/cbin.10107 · 1.64 Impact Factor