Herpes simplex virus-1 (HSV-1) oncolytic therapy and gene therapy are promising treatment modalities against cancer. NV1066, one such HSV-1 virus, carries a marker gene for enhanced green fluorescent protein (EGFP). The purpose of this study was to determine whether NV1066 is cytotoxic to lung cancer and whether EGFP is a detectable marker of viral infection in vitro and in vivo. We further investigated whether EGFP expression in infected cells can be used to localize the virus and to identify small metastatic tumor foci (<1 mm) in vivo by means of minimally invasive endoscopic systems equipped with fluorescent filters. In A549 human lung cancer cells, in vitro viral replication was determined by plaque assay, cell kill by LDH release assay, and EGFP expression by flow cytometry. In vivo, A549 cells were injected into the pleural cavity of athymic mice. Mice were treated with intrapleural injection of NV1066 or saline and examined for EGFP expression in tumor deposits using a stereomicroscope or a fluorescent thoracoscopic system. NV1066 replicated in, expressed EGFP in infected cells and killed tumor cells in vitro. In vivo, treatment with intrapleural NV1066 decreased pleural disease burden, as measured by chest wall nodule counts and organ weights. EGFP was easily visualized in tumor deposits, including microscopic foci, by fluorescent thoracoscopy. NV1066 has significant oncolytic activity against a human NSCLC cell line and is effective in limiting the progression of metastatic disease in an in vivo orthotopic model. By incorporating fluorescent filters into endoscopic systems, a minimally invasive means for diagnosing small metastatic pleural deposits and localization of viral therapy for thoracic malignancies may be developed using the EGFP marker gene inserted in oncolytic herpes simplex viruses.
Bibliographical noteFunding Information:
We thank Brian Horsburgh, PhD and Medigene, Inc. for constructing and providing us with the NV1066 virus. We thank Mithat Gonen, PhD, of the Department of Epidemiology and Biostatistics, and Katia Manova, PhD, John Waka, BS, and members of the Molecular Cytology Core Facility at Memorial Sloan-Kettering Cancer Center for their assistance with this project. We thank Hai Nguyen, VMD, of the Research Animal Resource Center of Cornell University Medical College for review of pathology specimens. Special thanks to Kan Matsumoto from Olympus America Inc., for design and construction of the fluorescent thoracoscopic system. This study was presented at the sixth annual meeting of the ‘American Society of Gene Therapy’, 2003, in Washington DC. This work was supported in part by training grant T32 CA09501 (BMS), AACR-AstraZeneca Cancer Research and Prevention Foundation Fellowship (PSA), grants RO1 CA 76416 and RO1 CA/DK80982 (YF) from the National Institutes of Health, grant MBC-99366 (YF) from the Am erican Cancer Society, grant BC024118 from the US Army (YF), grant IMG0402501 from the Susan Komen Cancer foundation (YF and PSA) and grant 032047 from Flight Attendant Medical Research Institute (YF and PSA).
Copyright 2008 Elsevier B.V., All rights reserved.
- Green fluorescent protein
- Lung neoplasm
- Minimally invasive
- Targeted therapy