Targeting gene therapy for hepatocarcinoma cells with the E. coli purine nucleoside phosphorylase suicide gene system directed by a chimeric α-fetoprotein promoter

Cai, Xiaokun; Zhou, Junli; Chang, Ying; Sun, Xiaowen; Li, Peiyuan; Lin, Junsheng

doi:10.1016/j.canlet.2008.01.027

Cited by 15 publications

(23 citation statements)

References 13 publications

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“…To date, RNA polymerase III-based promoters, U6 and H1, and the RNA polymerase II promoter, CMV, have been used to drive the expression of shRNAs, which in turn were processed into siRNAs [24,25]. However, these promoters have ubiquitous activity in all types of cells and would induce gene silencing without cell specificity [26]. In the present work, to avoid side-effects of nonspecific interference with targeted genes, an activated-HSC-specific gene silencing method was established, using an artificial intronic microRNA (miRNA) expression system to produce mature miRNAs against the target gene TGF-b1 in an activated-HSC-specific manner.…”

Section: Discussionmentioning

confidence: 99%

Hepatic Stellate Cell-Specific Gene Silencing Induced by an Artificial MicroRNA for Antifibrosis In Vitro

et al. 2009

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Section: Discussionmentioning

confidence: 99%

Hepatic Stellate Cell-Specific Gene Silencing Induced by an Artificial MicroRNA for Antifibrosis In Vitro

et al. 2009

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“…The AFP promoter has been used earlier to drive specific genes, mostly apoptotic or pro-drug metabolizing enzymes in hepatoma cells [55–59]. However, in our study, we have taken the minimal AFP promoter and added upstream enhancer regions from the AFP gene itself and, in another construct, the NFκB response element.…”

Section: Discussionmentioning

confidence: 99%

Combination of hepatocyte specific delivery and transformation dependent expression of shRNA inducing transcriptional gene silencing of c-Myc promoter in hepatocellular carcinoma cells

et al. 2014

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BackgroundA specific targeting modality for hepatocellular carcinoma (HCC) could ideally encompass a liver cell specific delivery system of a transcriptional unit that is active only in neoplastic cells. Sendai virosomes, derived from Sendai viral envelopes, home to hepatocytes based on the liver specific expression of asialoglycoprotein receptors (ASGPRs) which are recognized by the Sendai virosomal fusion (F) proteins. As reported earlier by us and other groups, transcriptional gene silencing (TGS) does not require continuous presence of the effector siRNA/shRNA molecule and is heritable, involving epigenetic modifications, leading to long term transcriptional repression. This could be advantageous over conventional gene therapy approaches, since continuous c-Myc inactivation is required to suppress hepatocarcinoma cells.MethodsExploiting such virosomal delivery, the alpha-fetoprotein (AFP) promoter, in combination with various tumour specific enhancers, was used to drive the expression of shRNA directed against ME1a1 binding site of the proto-oncogene c-Myc P2 promoter, in order to induce TGS in neoplastic liver cells.ResultsThe dual specificity achieved by the Sendai virosomal delivery system and the promoter/enhancer guided expression ensured that the shRNA inducing TGS was active only in liver cells that had undergone malignant transformation. Our results indicate that such a bimodal therapeutic system induced specific activation of apoptosis in hepatocarcinoma cells due to heterochromatization and increased DNA methylation of the CpG islands around the target loci.ConclusionsThe Sendai virosomal delivery system, combined with AFP promoter/enhancer expression machinery, could serve as a generalized mechanism for the expression of genes deleterious to transformed hepatocarcinoma cells. In this system, the epigenetic suppression of c-Myc could have an added advantage for inducing cell death in the targeted cells.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-582) contains supplementary material, which is available to authorized users.

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“…However, more detailed antitumor evidence is required, such as both in vitro and in vivo. Studies on the further deve lopments of GPC3 in other vectors with tumor-specific promoters, such as the AFPpromoter (15) or the telomerase promoter (16)(17) are underway.…”

Section: Disscusionmentioning

confidence: 99%

A new membrane re-anchored protein originating from GPC3 against hepatoma cells HepG2

Yang

et al. 2011

Mol Med Report

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Abstract. The aim of this study was to confirm the localization of recombinant pGPC3+afp-EGFP which expressed a new re-anchored protein named GPC3+afp-EGFP on the cytoplasmic membrane and to investigate its functions against hepatocellular carcinoma (HCC). EGFP expression in transfected HepG2 cells was observed using fluorescence and a confocal microscope. pGPC3+afp-EGFP expression was detected in membranous and soluble proteins extracted from transfected human embryonic kidney 293 cells by Western blot analysis using GPC3 mAb. The proliferation of transfected HepG2 cells with pGPC3+afp-EGFP (experimental group) was detected using SRB assay and compared to those of transfected HepG2 cells with pGPC3 (control group) and non-transfected HepG2 cells (blank group). Quantitative analysis of mRNA expression of the Fas gene was conducted by real-time PCR using the β-actin housekeeping gene as the internal control at variable times. Apoptotic HepG2 cells in the three groups were counted and statistically analyzed by a contingency table Chi-square test using Spss 11.5 software and TUNEL assay. Production of both TNF-α and IFN-γ/ IL2 was detected by ELISPOT after co-cultivation of transfected HepG2 cells with peripheral blood lymphocytes at different time-points in the experimental group. Green fluorescence was mainly found around the transfected HepG2 cell periphery through fluorescence and confocal microscopy. GPC3+afp-EGFP could not be detected in soluble protein but only in membranous protein. Proliferation curves showed that the proliferative quantities of transfected HepG2 cells in the experimental group decreased, whereas the mRNA expression of the Fas gene increased significantly compared to those of the other two groups. The numbers of apoptotic cells in the experimental group were significantly higher compared to those in the other two groups, as shown by statistical analysis. Both TNF-α and IFN-γ/IL2 were induced and were much higher in the experimental groups than in the diverse control groups at variable times. A new re-anchored protein GPC3+afp-EGFP expressed by recombinant pGPC3+afp-EGFP was localized on the cytoplasmic membrane, and had multiple functions against HCC, such as inhibition of transfected HepG2 cell proliferation, promotion of transfected HepG2 apoptosis and induction of antitumor cytokine excretion.

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Targeting gene therapy for hepatocarcinoma cells with the E. coli purine nucleoside phosphorylase suicide gene system directed by a chimeric α-fetoprotein promoter

Cited by 15 publications

References 13 publications

Hepatic Stellate Cell-Specific Gene Silencing Induced by an Artificial MicroRNA for Antifibrosis In Vitro

Hepatic Stellate Cell-Specific Gene Silencing Induced by an Artificial MicroRNA for Antifibrosis In Vitro

Combination of hepatocyte specific delivery and transformation dependent expression of shRNA inducing transcriptional gene silencing of c-Myc promoter in hepatocellular carcinoma cells

A new membrane re-anchored protein originating from GPC3 against hepatoma cells HepG2

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