Viral-mediated oncolysis is the most critical factor in the late-phase of the tumor regression process upon vaccinia virus infection.

BACKGROUND: In principle, the elimination of malignancies by oncolytic virotherapy could proceed by different mechanisms--e.g. tumor cell specific oncolysis, destruction of the tumor vasculature or an anti-tumoral immunological response. In this study, we analyzed the contribution of these factors to elucidate the responsible mechanism for regression of human breast tumor xenografts upon colonization with an attenuated vaccinia virus (VACV). METHODS: Breast tumor xenografts were analyzed 6 weeks post VACV infection (p.i.; regression phase) by immunohistochemistry and mouse-specific expression arrays. Viral-mediated oncolysis was determined by tumor growth analysis combined with microscopic studies of intratumoral virus distribution. The tumor vasculature was morphologically characterized by diameter and density measurements and vessel functionality was analyzed by lectin perfusion and extravasation studies. Immunological aspects of viral-mediated tumor regression were studied in either immune-deficient mouse strains (T-, B-, NK-cell-deficient) or upon cyclophosphamide-induced immunosuppression (MHCII+-cell depletion) in nude mice. RESULTS: Late stage VACV-infected breast tumors showed extensive necrosis, which was highly specific to cancer cells. The tumor vasculature in infected tumor areas remained functional and the endothelial cells were not infected. However, viral colonization triggers hyperpermeability and dilatation of the tumor vessels, which resembled the activated endothelium in wounded tissue. Moreover, we demonstrated an increased expression of genes involved in leukocyte-endothelial cell interaction in VACV-infected tumors, which orchestrate perivascular inflammatory cell infiltration. The immunohistochemical analysis of infected tumors displayed intense infiltration of MHCII-positive cells and colocalization of tumor vessels with MHCII+/CD31+ vascular leukocytes. However, GI-101A tumor growth analysis upon VACV-infection in either immunosuppressed nude mice (MHCII+-cell depleted) or in immune-deficient mouse strains (T-, B-, NK-cell-deficient) revealed that neither MHCII-positive immune cells nor T-, B-, or NK cells contributed significantly to VACV-mediated tumor regression. In contrast, tumors of immunosuppressed mice showed enhanced viral spreading and tumor necrosis. CONCLUSIONS: Taken together, these results indicate that VACV-mediated oncolysis is the primary mechanism of tumor shrinkage in the late regression phase. Neither the destruction of the tumor vasculature nor the massive VACV-mediated intratumoral inflammation was a prerequisite for tumor regression. We propose that approaches to enhance viral replication and spread within the tumor microenvironment should improve therapeutical outcome.

Regression of human prostate tumors and metastases in nude mice following treatment with the recombinant oncolytic vaccinia virus GLV-1h68.

Virotherapy using oncolytic vaccinia virus strains is one of the most promising new strategies for cancer therapy. In the current study, we analyzed the therapeutic efficacy of the oncolytic vaccinia virus GLV-1h68 against two human prostate cancer cell lines DU-145 and PC-3 in cell culture and in tumor xenograft models. By viral proliferation assays and cell survival tests, we demonstrated that GLV-1h68 was able to infect, replicate in, and lyse these prostate cancer cells in culture. In DU-145 and PC-3 tumor xenograft models, a single intravenous injection with GLV-1h68 resulted in a significant reduction of primary tumor size. In addition, the GLV-1h68-infection led to strong inflammatory and oncolytic effects resulting in drastic reduction of regional lymph nodes with PC-3 metastases. Our data documented that the GLV-1h68 virus has a great potential for treatment of human prostate carcinoma.

Systemic treatment of xenografts with vaccinia virus GLV-1h68 reveals the immunologic facet of oncolytic therapy.

BACKGROUND: GLV-1h68 is an attenuated recombinant vaccinia virus (VACV) that selectively colonizes established human xenografts inducing their complete regression. RESULTS: Here, we explored xenograft/VACV/host interactions in vivo adopting organism-specific expression arrays and tumor cell/VACV in vitro comparing VACV replication patterns. There were no clear-cut differences in vitro among responding and non-responding tumors, however, tumor rejection was associated in vivo with activation of interferon-stimulated genes (ISGs) and innate immune host's effector functions (IEFs) correlating with VACV colonization of the xenografts. These signatures precisely reproduce those observed in humans during immune-mediated tissue-specific destruction (TSD) that causes tumor or allograft rejection, autoimmunity or clearance of pathogens. We recently defined these common pathways in the "immunologic constant of rejection" hypothesis (ICR). CONCLUSION: This study provides the first prospective validation of a universal mechanism associated with TSD. Thus, xenograft infection by oncolytic VACV, beyond offering a promising therapy of established cancers, may represent a reliable pre-clinical model to test therapeutic strategies aimed at modulating the central pathways leading to TSD; this information may lead to the identification of principles that could refine the treatment of cancer and chronic infection by immune stimulation or autoimmunity and allograft rejection through immune tolerance.

Anti-VEGF single-chain antibody GLAF-1 encoded by oncolytic vaccinia virus significantly enhances antitumor therapy.

We previously reported that the replication-competent vaccinia virus (VACV) GLV-1h68 shows remarkable oncolytic activity and efficacy in different animal models as a single treatment modality and also in combination with chemotherapy [Yu YA, et al. (2009) Mol Cancer Ther 8:141-151]. Here, we report the construction of 3 VACV strains encoding GLAF-1, a previously undescribed engineered single-chain antibody (scAb). This unique scAb is transcribed from 3 vaccinia promoters (synthetic early, early/late, and late) and directed against both human and murine VEGFs. The expression of GLAF-1 was demonstrated in cell cultures. Also, the replication efficiency of all GLAF-1-expressing VACV strains in cell culture was similar to that of the parental GLV-1h68 virus. Successful tumor-specific delivery and continued production of functional scAb derived from individual VACV strains were obtained in tumor xenografts following a single intravenous injection of the virus. The VACV strains expressing the scAb exhibited significantly enhanced therapeutic efficacy in comparison to treatment of human tumor xenografts with the parental virus GLV-1h68. This enhanced efficacy was comparable to the concomitant treatment of tumors with a one-time i.v. injection of GLV-1h68 and multiple i.p. injections of Avastin. Taken together, the VACV-mediated delivery and production of immunotherapeutic anti-VEGF scAb in colonized tumors may open the way for a unique therapy concept: tumor-specific, locally amplified drug therapy in humans.

Resveratrol, a natural diphenol, reduces metastatic growth in an experimental cancer model.

Administration of different doses of the diphenol resveratrol had no effect on the growth of an intramuscularly implanted experimental tumour, the Lewis lung carcinoma. These results do not agree with previous reports where a clear effect of resveratrol was shown on tumour burden in both mice and rats. However, administration of the diphenol had a clear anti-metastatic effect, decreasing both the number and the weight of the lung metastases. Similar effects were observed both at 5 and 25mg/kg body weight per day, resulting in an approximately 40% reduction in the number of metastases. These results suggest that resveratrol could be tentatively given as a preventive agent in cancer patients undergoing radiotherapy or chemotherapy.