| To reduce the increase in angiogenesis reported following o ncolytic herpes simplex virus (OV) therapy, we first describe the construction and testing of a novel oncolytic HSV-1 derived virus: 34.5ENVE (viral ICP 34.5 is Expressed by Nestin promotor and Vstat120 Expressing). This virus showed significant glioma specific killing and anti-angiogenic effects in vitro and in vivo. Treatment of mice bearing subcutaneous and intracranial glioma with 34.5ENVE resulted in a significant increase in animal survival, with 100% (subcutaneous) and 75% (intracranial) of mice showing a complete response. Histology and dynamic contrast enhanced magnetic resonance imaging (DCE MRI) revealed reduced microvessel density and increased tumoral necrosis in tumors treated with 34.5ENVE compared to tumors treated with a control virus. Collectively, these results describe the enhanced therapeutic efficacy of a transcriptionally driven OV by way of exploiting its impact on the tumor microenvironment.;Next, we describe the role of Cysteine rich 61 (CCN1) in the tumor microenvironment on OV efficacy. CCN1 is a secreted extracellular matrix (ECM) protein elevated in cancer cells that modulates their adhesion and migration by binding cell surface receptors. We examined a hypothesized role for CCN1 in limiting the efficacy of oncolytic viral therapy for glioma, based on evidence of CCN1 induction that occurs in this setting. Expression is up-regulated in a variety of cancers, including glioma, resulting in a worse prognosis for these patients. As a significant induction of secreted CCN1 shortly following oncolytic viral therapy of glioma cells has been shown, we evaluated its role in the cellular response to viral infection. We found that exogenous CCN1 in glioma ECM orchestrates a cellular antiviral response that reduces viral replication and limits oncolytic virus efficacy. Gene expression profiling and real time PCR analysis revealed a significant induction of type-I interferon responsive genes in response to CCN1. Using function blocking antibodies we discovered this effect was mediated by CCN1 binding the alpha6beta1 integrin receptor, resulting in the rapid secretion of IFNalpha which was essential for this innate antiviral effect. Collectively, these results describe the novel role of a CCN1-integrin interaction in the activation of the type-I antiviral interferon response and ultimate inhibition of OV therapy.;Lastly, we investigate the relationship between CCN1 and macrophage-mediated oncolytic virus clearance in glioma. Using function-neutralizing antibodies, we show that inhibition of endogenously up-regulated CCN1 following OV therapy reduces OV mediated macrophage migration. Interestingly, CCN1's coordinated increase in macrophage migration was found due both to its direct interaction with macrophages, as evidenced by an enhancement in macrophage migration with purified CCN1 protein, as well as to its direct interaction with glioma cells, which results in an increased production of chemokines. CCN1 enhanced the pro-inflammatory activation of macrophages following OV infection, which led to an increase in macrophage-mediated viral clearance in vitro. Though knock-down of the integrin alpha6beta1 on glioma cells did reduce the type I IFN response presented above, it had no effect on CCN1's relationship with macrophages. Examination of the cell surface integrin alphaMbeta2, known to mediate the CCN1-macrophage interaction, revealed it to be the main effector for CCN1's effect. In vivo, use of an anti-CCN1 antibody in mice bearing subcutaneous gliomas treated with oncolytic rHSVQ-IE4/5-Luc revealed enhanced luciferase activity along with reduced macrophage infiltration. Taken together, our findings indicate CCN1 not only has a role in the immediate activation of the type I IFN response, thereby inhibiting virus infection, but also in increasing macrophage infiltration and activation resulting in a macrophage mediated reduction in viral oncolysis. Further, this study warrants investigation of therapeutic strategies to reduce CCN1 following OV therapy, with the intention of creating a more suitable tumor microenvironment for virus therapy. (Abstract shortened by UMI.). |
