Purpose: The oncolytic effects of a systemically delivered, replicating, double-deleted vaccinia virus has been previously shown for the treatment of many cancers, including colon, ovarian, and others.The purpose of this study was to investigate the oncolytic potential of double-deleted vaccinia virus alone or in combination with rapamycin or cyclophosphamide to treat malignant gliomas in vitro and in vivo. Experimental Design: Rat (RG2, F98, C6) and human (A172, U87MG, U118) glioma cell lines were cultured in vitro and treated with live or UV-inactivated vaccinia virus. Viral gene [enhanced green fluorescent protein (EGFP)] expression by fluorescence-activated cell sorting, relative cell viability by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), and assays for cytopathic effects were examined. S.c. murine tumor xenografts (U87MG, U118, C6) and i.c. (RG2, F98) tumor models in immunocompetent rats were treated with systemic administration of EGFP-expressing vaccinia virus (vvDD-EGFP), alone or in combination with rapamycin or cyclophosphamide, or controls. Tumor size, viral biodistribution, and animal survival were assessed. Lastly, the oncolytic effects of vvDD-EGFP on human malignant glioma explants were evaluated. Results: vvDD-EGFP was able to infect and kill glioma cells in vitro. A single systemic dose of vvDD-EGFP significantly inhibited the growth of xenografts in athymic mice. Systemic delivery of vvDD-EGFP alone was able to target solitary and multifocal i.c. tumors and prolong survival of immunocompetent rats, whereas combination therapy with rapamycin or cyclophosphamide enhanced viral replication and further prolonged survival. Finally, vvDD-EGFP was able to infect and kill ex vivo primary human malignant gliomas. Conclusions: These results suggest that vvDD-EGFP is a promising novel agent for human malignant glioma therapy, and in combination with immunosuppressive agents, may lead to prolonged survival from this disease.

Oncolytic myxoma virus (MYXV) is being developed as a novel virotherapeutic against human brain cancer and has promising activity against human brain tumor models in immunocompromised hosts. Because an intact immune system could reduce its efficacy, the purpose of this study was to evaluate the oncolytic potential of MYXV in immunocompetent racine glioma models. Here, we report that MYXV infects and kills all racine cell glioma lines and that its effects are enhanced by rapamycin. Intratumoral administration of MYXV with rapamycin improved viral replication in the tumor and significantly prolonged host survival. Similarly, coadministration via a method of convection-enhanced delivery (CED) enhanced viral replication and efficacy in vivo. Mechanisms by which rapamycin improved MYXV oncolysis included an inhibition of type I IFN production in vitro and a reduction of intratumoral infiltration of CD68 + microglia/macrophages and CD163 + macrophages in vivo. Our findings define a method to improve MYXV efficacy against gliomas by rapamycin coadministration, which acts to promote immune responses engaged by viral delivery. CancerRes; 70(2); 598-608. ©2010 AACR.

We have shown previously the oncolytic potential of myxoma virus in a murine xenograft model of human glioma. Here, we show that myxoma virus used alone or in combination with rapamycin is effective and safe when used in experimental models of medulloblastoma in vitro and in vivo. Nine of 10 medulloblastoma cell lines tested were susceptible to lethal myxoma virus infection, and pretreatment of cells with rapamycin increased the extent of in vitro oncolysis. Intratumoral injection of live myxoma virus when compared with control inactivated virus prolonged survival in D341 and Daoy orthotopic human medulloblastoma xenograft mouse models [D341 median survival: 21 versus 12.5 days; P = 0.0008; Daoy median survival: not reached (three of five mice apparently ''cured'' after 223 days) versus 75 days; P = 0.0021]. Rapamycin increased the extent of viral oncolysis, ''curing'' most Daoy tumor-bearing mice and reducing or eliminating spinal cord and ventricle metastases. Rapamycin enhanced tumor-specific myxoma virus replication in vivo and prolonged survival of D341 tumor-bearing mice (median survival of mice treated with live virus (LV) and rapamycin, versus LV alone, versus rapamycin alone, versus inactivated virus: 25 days versus 19, 13, and 11 days, respectively; P < 0.0001). Rapamycin increased the levels of constitutively activated Akt in Daoy and D341 cells, which may explain its ability to enhance myxoma virus oncolysis. These observations suggest that myxoma virus may be an effective oncolytic agent against medulloblastoma and that combination therapy with signaling inhibitors that modulate activity of the phosphatidylinositol 3-kinase/Akt pathway will further enhance the oncolytic potential of myxoma virus.