Novel Near-Infrared Fluorescence-Guided Surgery With Vesicular Stomatitis Virus for Complete Surgical Resection of Osteosarcomas in Mice

Attempts have been made to visualize tumor cells intraoperatively with fluorescence guidance. However, the clear demarcation and complete tumor resection have always been a challenging task. To address this, we have developed a novel fluorescence bioimaging system with vesicular stomatitis virus (VSV) incorporating Katushka, near-infrared fluorescent protein. VSV is tumor-specific owing to the deficiency of antiviral interferon signaling pathways in tumor cells. We aimed to evaluate the tumorspecificity of the recombinant VSV-Katushka (rVSV-K) in osteosarcoma cells and to assess the feasibility of complete tumor resectionby the rVSV-K fluorescence guidance. In in vitro experiments, mouse and human osteosarcoma cell lines and normal humanmesenchymal stem cells were infected with rVSV-K and observed by fluorescence microscopy. Near-infrared fluorescence was observedonly in osteosarcoma cells, even at a low-concentration of virus infections. In in vivo experiments, mouse osteosarcoma (LM8) cells were transplanted subcutaneously into the back of immune-competent mice to produce an osteosarcoma, which was then injected with rVSV-K. The areas emitting fluorescence were resected using a bioimaging system. The distance between the surgical and tumor margins ofthe fluorescence-guided resection with rVSV-K group was significantly larger than that of the non-guided resection groups. The local recurrence rate was significantly lower in the fluorescence-guided resection with rVSV-K group than in the non-guided resection groups. The distant metastasis rate and average survival rate were not significantly different between all groups. These results suggest