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Tumor-Targeted rVSV-F Vectors for Treatment of Metastatic Breast Cancer
Background: Oncolytic virotherapy is an exciting novel strategy to treat advanced breast cancer. Vesicular Stomatitis Virus (VSV) is a non-segmented negative-strand RNA virus with inherent selectivity for replication in tumor cells due to their attenuation of anti-viral responses. We have previously demonstrated tumoricidal effects of a fusogenic VSV vector (rVSV-F) in mice bearing multi-focal lesions of 4T1 breast carcinoma in their lungs, which led to statistically significant survival benefits. This vector, however, has no selectivity for tumor cells at the infection level and the overall survival advantage was also modest, as its maximum tolerated dose is limited by neuropathology in some of the treated animals. Objective/Hypothesis: We hypothesize that the therapeutic index of oncolytic rVSV-F can be substantially enhanced by intravenous administration of rVSV-F vectors with altered cellular tropism, such that their tumor cell tropism is elevated while neuronal tropism is diminished. This can be accomplished by pseudo-typing rVSV-F with the glycoprotein gene of Ebola virus (EVG), which uses the folate receptor as a co-receptor that is expressed highly in most cancer cell types but at exceptionally low levels in neurons, which are completely refractory to infection by lentivirus vectors pseudo-typed with EVG. To ensure the safety of EVG pseudo-typed rVSV-F vectors, the mucin-like domain of EVG responsible for endothelial cell tropism and vascular toxicity will be deleted, which does not diminish the folate receptor binding efficiency of the correspondingly pseudo-typed lentivirus vectors. Specific Aims: Pseudo-typed rVSV-F vectors will be rescued in folate receptor positive cells using reverse genetics techniques, and their oncolytic potency in 4T1 murine and various human breast cancer cell lines in vitro will be determined. The MTD of the pseudo-typed rVSV-F vectors will be determined in mice by dose-controlled studies. To test treatment efficacy multi-focal lung lesions of an exceptionally malignant breast cancer model generated by tail vein infusion of murine 4T1 breast cancer cells into immune-competent and syngeneic Balb/c mice will be used, which shares many characteristics with naturally occurring human breast cancers. Pseudo-typed rVSV-F vectors will be injected intravenously into tumor-bearing mice at their respective MTDs, using survival as the initial endpoint. In addition, subsets of tumor-bearing mice in each of the treatment groups will be sacrificed at various time points for the collection of tumor tissues and major organs to determine tumor response, vector bio-distribution, systemic and organ toxicities. Impact: The successful development and validation of these tumor-targeted rVSV-F vectors in the treatment of metastatic breast cancer in mice would constitute a major step towards the future development of oncolytic VSV virotherapy for the effective and safe treatment of metastatic breast cancer in humans.
Despite recent clinical advances in treatment modalities an overwhelming majority of patients diagnosed with metastatic breast cancer will die of the disease, and new therapeutic options based on novel principles are urgently needed. In the recent years, a new scientific principle of using live viruses that can infect and propagate only in malignant, but not normal, cells has emerged as a powerful novel strategy to treat cancer, which is called "Oncolytic Virotherapy". Amongst these "oncolytic viruses", Vesicular Stomatitis Virus (VSV) is a particularly attractive agent because it replicates robustly and rapidly in most animal and human cancer cells in culture, which are completely killed within 1-2 days even if only one virus particle was used to infect one thousand cancer cells. It does not replicate well in normal cells and is not a human pathogen, as it causes only a mild respiratory infection in cattle that is cleared within a few days. We have previously shown that an engineered form of the virus, when administered into the hepatic artery, was effective against multi-focal Hepatocellular Carcinoma (HCC) in the livers of rats and a proposal to conduct a Phase I clinical translational trial in HCC patients will be conducted with sponsorship from the National Cancer Institute. We have also shown that the engineered oncolytic VSV can be used to treat multi-focal lung metastases of 4T1, which is an exceptionally aggressive type of malignant breast cancer model in mice and resembles closely advanced breast cancer in humans. Due to the limitation of virus dose that could be safely administered however, not all breast cancer cells in the multi-focal lung lesions were killed and all treated mice eventually succumbed to tumor relapse. We reasoned that the effectiveness of treatment could be substantially enhanced if the treatment dose can be elevated without side effects. Because VSV can recognize a surface receptor that is present in most mammalian cell types, it can infect a wide range of cells. In the proposed study, we will substitute the surface protein of VSV with one that binds specifically to cancer cells, thereby enhancing its selective binding and entry into cancer cells where they can propagate and exert their killing effects. The effectiveness of the novel oncolytic virus to treat multi-focal lesions of breast cancer in the lungs will then be tested in the murine 4T1 breast cancer model in mice. While enhanced efficacy in tumor killing and prolonged survival might be obtained in tumor-bearing mice, substantially enhancement in safety is also expected, owing to increased tumor-selectivity of the engineered oncolytic virus. These studies will form the scientific basis to develop clinical translational trials for patients with metastatic breast cancer in the future, which can potentially lead to the development of a novel and effective therapeutic agent for this deleterious disease.