Anti-tumor Effect of P19Arf and Interferon-beta Gene Transference to Mouse Melanoma and Mouse Lung Carcinoma Cells is Revealed by a Strong Bystander Activity and a Potential Immune Response
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conferenceObject
Data de publicação
2012
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Editora
ELSEVIER SCI LTD
Citação
EUROPEAN JOURNAL OF CANCER, v.48, suppl.5, p.S94-S95, 2012
Resumo
Introduction: Two hallmarks of tumor progression are resistance to cell death and lack of an effective anti-tumor immune response. Loss of p53 function, by genetic mutation or alterations in its pathway, such as p19Arf loss and/or mdm2 over-expression, inhibits one of the primary coordinators of cell death. Interferon-beta (IFN), a stimulator of the immune response with anti-neoplastic functions, is also frequently lost in some tumor types. We propose that p19Arf and IFN gene transfer would be an effective strategy, especially in wild-type p53 tumor cells, since cell death and immune activation would be combined to combat the tumor at primary treatment site and metastasis. Material and Methods: Recombinant adenoviral vectors with RGD-modified fiber (rAdRGD) and a p53-responsive promoter (PG) were constructed containing genes of p19Arf, IFN or the combination of both. Evaluation of in vitro antiproliferative effect of transgenes in B16F10 cells (B16, mouse melanoma, p53 wt) was done by annexin/PI staining and MTT assays. Bystander effect was revealed by cell cycle analysis of populations transduced with different proportions of the viruses. Antitumor effect in vivo was observed by treatment of established LLC1 tumors (mouse lung carcinoma, p53 wt) with intratumoral injection of rAdRGD in C57BL/6 mice. Involvement of immune response was revealed by second tumor challenge at contralateral flank of mice with a developed and treated first tumor. Results and Discussion: Cell death was resulted from the p19Arf and IFN combined transference (74% subG0), yet single gene transfer yielded only half the number of subG0 cells. A similar result was seen by measurement of cell viability with MTT. Evidence on a bystander effect was revealed when approximately 50% subG0 cells were observed, even though only 10% of the cells had been transduced with IFN. In a population of cells transduced with p19Arf, when 10% of them also expressed IFN, the number of subG0 cells increased to 68%, compared to transduction of p19Arf alone, which results in 45% subG0 cells. This indicates that p19Arf can sensitize cells to death by IFN bystander effect. In vivo assays with the LLC1 model have shown that in situ gene therapy of p19Arf and IFN combination was more effective to inhibit tumor progression and increase survival than application of a single gene. These animals were then challenged with the implantation of a second tumor, revealing greater retardation of growth at the secondary tumor site in mice treated with the combined gene therapy at the primary tumor locus as compared to animals that received single gene treatment. Conclusion: The use of p53-responsive vectors to express p19Arf and IFN represents a potential strategy for melanoma and lung carcinoma tumor suppression. We have shown that complementation of the p53/Arf and interferon pathways in the primary tumor may generate a strong bystander effect as well as immune stimulation.