Abstract
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.
Highlights
A 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
In the transformed HCC cells, HepG2 and Huh7, the luciferase activity was highest with AFPEn– Pr + 25 – luc followed by NFκBEn–Pr + 25 – luc and lastly by AFPPr + 25 – luc, indicating the relative activity of the AFPEn–Pr + 25 in the transformed cells is significantly higher than Simian virus 40 (SV40) promoter (Figure 1B and C)
Such approaches might be utilized for other therapeutic modalities that are based on specific gene transcription e.g. Gene dependent enzyme prodrug therapy (GDEPT). Short hairpin RNA (shRNA) induced suppression of c-Myc expression by transcriptional gene silencing (TGS) is a possible gene therapy modality that could be utilized in such a delivery system
Summary
A 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. 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. Sendai F-virosome mediated delivery overcomes this limitation since the entrapped cargo is directly delivered into the cytoplasm, evading the endosomal pathway [16,17] This could enhance transgene expression and its longevity for therapeutic purposes
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