Abstracts
1998 Digestive Disease Week

#3636

ATTENUATED HSV-1 KILLS HUMAN PANCREATIC CANCER CELLS IN VITRO. J.H. Lee1, H.J. Federoff2, J.L. Peacock1, and L.O. Schoeniger1. Departments of Surgery1 and Neurology2, University of Rochester Medical Center, Rochester, NY.

The objective of this study was to determine the efficacy of G207 in killing human pancreatic cancer cells in vitro. G207 is a multi-mutated strain of HSV-1 (Herpes Simplex Virus Type 1); unlike other vectors previously used in cytotoxic gene therapies, this virus is capable of efficient growth in many dividing cells, including certain tumor cells, but not in non-dividing cells. Background: Pancreas cancer is common and lethal, thus requiring formulation of additional management strategies. G207 is an attenuated and replication restricted mutant of HSV-1 that has E. coli lacZ reporter gene inserted in its genome. This virus has previously been shown, in animal models, to be cytopathic only in human malignant meningioma and malignant glioma cell lines. Methods and Results: In a tissue culture setting, two human pancreatic cell lines, AsPC-1 and BxPC-3, were infected with G207 at MOI's (Multiplicity Of Infection) of 5X10-5 to 5X10-1. After 24 hrs of incubation at 37°C with 5% CO2, the viral gene expression was tested using a histochemical X-gal assay to detect _-galactosidase expression. The results show a viral dose-dependent infectivity and gene expression. In addition, the above cell lines were infected with G207 at a MOI of 0.02, incubated at 34°C with 5% CO2 for 72 hrs, then the supernatants were collected and titered on a permissive cell (Vero) line for 72 hrs. The results show a viral dose-dependent plaque formation, with viral titers of 2X103 pfu/ml and 6X105 pfu/ml from the supernatants of AsPC-1 and BxPC-3 respectively. Lastly the above cell lines were infected with G207 at MOI's of 0.001 to 10. The MTS (Owen's reagent) assays were done to assess the proportion of viable cells at 24 - 96 hrs post-infection. The results show a MOI dependent decrease in absorbance at 490 nm, thus indicating a viral dose-dependent cytotoxicity of G207 to above tested cell lines. More specifically, for the MOI's of 1 and 10, the MTS assay shows significant cell killing (> 80% for AsPC-1, and > 90% for BxPC-3, compared to controls -- p <0.001) by 72 hrs post-infection, and was confirmed by direct microscopic evaluation. Conclusion: The results indicate that G207 is an effective vector for gene therapy in human pancreatic cancer cell lines in vitro; it replicates in, and is cytotoxic to the above listed human pancreatic cell lines, AsPC-1 and BxPC-3. These data, combined with previous works on non-human primates suggesting that G207 is non-pathogenic in normal tissue, indicate that replication restricted viruses, particularly G207, are potentially important new reagents for the study of pancreatic cancer, and its treatment.

Copyright 1996 - 1998, SSAT, Inc. Revised 29 June 1998.