Molecular Effects of Stromal-Selective Targeting by uPAR-Retargeted Oncolytic Virus in Breast Cancer

Abstract

The tumor microenvironment (TME) is a relevant target for novel biological therapies. MV-m-uPA and MV-h-uPA are fully retargeted, species-specific, oncolytic measles viruses (MVs) directed against murine or human urokinase receptor (PLAUR/uPAR), expressed in tumor and stromal cells. The effects of stromal selective targeting by uPAR retargeted MVs were investigated. In vitro infection, virus-induced GFP expression and cytotoxicity by MV-h-uPA and MV-m-uPA were demonstrated in human and murine cancer cells and cancer associated fibroblasts (CAFs) in a species-specific manner. In a murine fibroblast/human breast cancer 3D co-culture model, selective fibroblast targeting by MV-m-uPA inhibited breast cancer cell growth. Systemic administration of murine specific MV-m-uPA in mice bearing human MDA-MB 231 xenografts was associated with a significant delay in tumor progression and improved survival compared to controls. Experiments comparing tumor (MV-h-uPA) vs. stromal (MV-m-uPA) vs. combined virus targeting showed that tumor and stromal targeting was associated with improved tumor control over the other groups. Correlative studies confirmed in vivo viral targeting of tumor stroma by MV-m-uPA, increased apoptosis, and virus induced differential regulation of murine stromal genes associated with inflammatory, angiogenesis and survival pathways, as well as indirect regulation of human cancer pathways, indicating viral induced modulation of tumor-stromal interactions. These data demonstrate the feasibility of stromal selective targeting by an oncolytic MV, virus-induced modulation of tumor-stromal pathways, and subsequent tumor growth delay. These findings further validate the critical role of stromal uPAR in cancer progression and the potential of oncolytic viruses as anti-stromal agents.

In vivo antitumor activity by dual stromal and tumor-targeted oncolytic measles viruses

“…Although both MV-GFP and MV-CD46-muPA induced inhibition of tumor cell proliferation and enhanced in vivo apoptosis, the magnitude of the proapoptotic effect was significantly higher in tumors treated by the dual targeted virus. The antistromal effects by MV-CD46-muPA further validate a previous report by our group, showing that single targeted, stromal specific MV-muPA (selectively targeting mouse uPAR) induces significant modulation of murine stromal pathways, such as angiogenesis, inflammatory networks, as well as indirect modulation of human cancer cell pathways [27]. In this report, we further characterized the in vivo MV oncolytic mechanisms by functional RPPA of tumors treated by the tumor vs. tumor and stromal targeted MV vectors.…”

“…4). We have previously reported the in vivo effects of stromal targeted only (MV-muPA) MV vectors compared with tumor only targeted vectors in breast cancer models, showing measurable, but modest effects of stromal only in vivo targeting [27]. The improved in vivo effects were associated with increased tumor viral protein (MV-N) expression, as well as increased tumor cell apoptosis.…”

“…We demonstrated that systemic 10 3 10 4 10 5 10 6 10 3 10 4 10 5 10 6 10 3 10 4 10 5 10 6 10 3 10 4 10 5 10 6 10 3 10 4 10 5 10 6 10 3 10 4 10 5 10 6 10 2 10 3 10 4 10 5 10 2 10 3 10 4 10 5 10 2 10 3 10 4 10 5 10 2 10 3 10 4 10 5 10 2 10 3 10 4 10 5 10 2 10 3 10 4 10 5 10 2 10 3 10 4 10 5 administration of species specific, human (MV-huPA) or murine (MV-muPA) uPAR retargeted MVs is safe, successfully targets tumor tissues over non-cancer tissues, and is associated with significant tumor delaying effects in primary or metastatic, xenograft, and syngeneic cancer models, respectively [18,26]. Taking advantage of the species specificity of MV-muPA, we recently reported that (murine) stromal-selective MV-muPA was associated with direct stromal targeting in a human breast cancer xenograft (where human cancer cells are not permissive to the murine targeted virus), leading to modulation of murine stromal gene expression, indirect effects on tumor cell gene expression, and measurable in vivo tumor growth delay by MV-muPA [27]. However, the in vivo antitumor effects of targeting tumor-stromal cells alone were modest compared to tumor selective targeting, suggesting that combined tumor and stromal targeting may further enhance antitumor efficacy.…”

Three-dimensional tumor cell cultures employed in virotherapy research

“…Further approaches use collagen-derived matrices or synthetically engineered polyethylene glycol (PEG)-based hydrogels ( Table 3 ). 9 , 47 , 48 Collagen is a well-known structural protein of the connective tissue, and can be selectively lysed by collagenase in cell culture to release cells from the collagen matrix. Hydrogels provide a three-dimensional scaffold that encapsulates the cells and makes nutrient exchange possible ( Figure 1B ).…”

“…Supported by a mouse model, the authors concluded that stroma-selective targeting per se constituted an efficient treatment option, even though targeting both tumor and tumor stroma led to the strongest antitumor effect. 47 …”