Abstract
Hepatocellular carcinoma (HCC) is a major health problem as evidenced by its increasing incidence and high morbidity and mortality rates. Most patients with HCC have underlying liver disease and dysfunction which limits the current therapeutic options. Treatments that spare the liver and destroy the HCC are needed. Targeting transcriptional differences between HCC and liver cells may provide this therapeutic window. In this study, we examine the potential of the Glypican 3 (GPC3) promoter as a targeting strategy. GPC3 is an oncofetal protein belonging to the proteoglycan family which is normally only expressed during fetal development. However, in HCC, the expression of this protein is reactivated. Here, we show that GPC3 is expressed primarily in HCC and not in normal liver lines. We show that the GPC3 promoter can be used to drive expression of significantly more luciferase and eYFP in HCC cell lines compared to normal liver cells. Further, we show that vectors containing cytosine deaminase (CD) under GPC3 promotor control induced significantly more killing of HCC cell lines after treatment with 5-FC compared to normal liver cell lines. These data suggest that transcriptionally targeted delivery of transgene in HCC cells can be achieved using the GPC3 promoter and this targeting strategy produces limited toxicity to normal liver cells.
Highlights
Oncolytic viral therapy has shown preclinical and clinical promise, but the large amount of virus needed for adequate dosing results in extreme expense
We demonstrate that histone-induced cytotoxicity can be reversed by treatment with the RNA aptamers both in vitro and in vivo in a mouse model of MODS/ARDS
Gp130 alternative splicing can be modulated to generate soluble antagonist isoforms, which can bind to IL-6/sIL-6R
Summary
Oncolytic viral therapy has shown preclinical and clinical promise, but the large amount of virus needed for adequate dosing results in extreme expense. Our lab previously reported that an acute insulin therapy co-administered with AAV vector delivery to skeletal muscle and liver (mouse tissue and human cultured cells) increased transgene product. The promoter of alpha fetoprotein (AFP), an established HCC-specific promoter, was used for comparison The activity of these promoters in a panel of normal liver, HCC and non-HCC cell lines was evaluated by correlating it with the expression levels of corresponding genes using real time rtPCR. Conclusion: We have demonstrated that the promoter for GPC3 is active in the majority of HCC and not in normal liver and can be used to selectively target HCC cells with gene therapy; as with other promoter driven systems, additional targeting strategies may be required if treatment is delivered systemically. Methods: Third generation self-inactivating lentiviral vector constructs were used to co-deliver an anti-CD19 CAR and HSV-sr39TK or EGFRt
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