The Inhibitory Effect Of A Novel Oncolytic Vaccinia Virus OVV-ING4 Combined With Gemcitabine Against Pancreatic Cancer Cells

Objective With no effective therapy accessible for most patients with pancreatic cancer,pancreatic cancer still remains to be one of the most difficult treated malignancies.So the essential affairs are investigating the pathogenesis of pancreatic cancer and exploring more effective anti-tumor courses.Presently,oncolytic virus mediated-gene therapy has displayed ubiquitous anti-tumor potential.Oncolytic virus is able to selectively replicate in cancer cells and motive cell lysis,without impact on normal cells.Recombinant vaccinia viruses are powerful vectors for cancer gene therapy.ING4 has been reported to be overexpressed in most types of cancer cells.Thus,targeted ING4 gene therapy may be a very effective way to kill cancers.In order to improve the therapeutic effect of pancreatic cancer,in this study,we verified the biological characters of the innovative oncolytic vaccinia virus o VV-ING4carrying the inhibitor of growth family member 4 gene that had constructed successfully,and investigated its anti-tumor efficacy alone or in combination with gemcitabine against human pancreatic cancer cells in vitro and in vivo.Methods ING4 expression was detected via quantitative real-time polymerase chainreaction(q PCR)and Western blot.TCID50(50%tissue culture infectious dose)assay was used to determine the virus titer.Fluorescent inverted microscope was applied to observe the infectious ability of the recombinant vaccinia virus.The cytotoxicity of o VV-ING4 or gemcitabine was measured using MTS cell proliferation assay.Flow cytometry and Western blot were adopted to detect the cell cycle arrest and apoptosis.In addition,combined inhibitory efficiency of o VV-ING4 and gemcitabine was estimated using Chou-Talalay combination index(CI)analysis in vitro and a Blab/c mice model in vivo.Results Results showed that the expression of ING4 was lower in pancreatic cancer cells compared with that in pancreatic normal cells.The titer of o VV-ING4 was1.61×10~8 pfu/m L.Importantly,o VV-ING4 can selectively replicate,significantly increased ING4 expression,showed greater cytotoxic efficiency,and induced cell apoptosis and G2 phase arrest in pancreatic cancer cells.The EC50(concentration for50%of maximal effect)value for PANC-1,BXPC-3 and SW1990 cells was 20.04MOI,34.60 MOI and 9.48 MOI,respectively.The IC50(half maximal inhibitory concentration)value of gemcitabine for PANC-1 and SW1990 cells was 37.70μmol/L and 27.83μmol/L,respectively.Additionally,the combination of o VV-ING4 and gemcitabine synergistically effected in vitro and in vivo.At all the fractions detected,the CI value in SW1990 was 0.903-0.633,log10(CI)<0,and in PANC-1 was0.538-0.439,log10(CI)<0.Conclusion In brief,our data suggest o VV-ING4 has the ability to selectively replicate in cancer cells,higher expression of target gene,powerful tumor-specific killing effect with minimal toxicity to normal cells,and the ability to induce tumor cell apoptosis and to cause G2 phase arrest.Moreover,combination of o VV-ING4 and gemcitabine displays synergistic killing effect.Taken together,o VV-ING4 is a powerful pancreatic cancer therapeutic approach alone or in combination with gemcitabine.We hope to provide an updated,higher efficacy,and lower toxicity method of biological treatment for pancreatic cancer,and further expand the strategy of viral-gene-chemotherapy for the clinical treatment of pancreatic cancer.