Hepatocellular carcinoma (HCC) develops predominantly in the inflammatory environment of a cirrhotic liver caused by hepatitis, toxin exposure, or chronic liver disease. A targeted therapeutic approach is required to enable cancer killing without causing toxicity and liver failure. Poly(beta-amino-ester) (PBAE) nanoparticles (NPs) were used to deliver a completely CpG-free plasmid harboring mutant herpes simplex virus type 1 sr39 thymidine kinase (sr39) DNA to human HCC cells. Transfection with sr39 enables cancer cell killing with the prodrug ganciclovir and accumulation of 9-(4-18F-fluoro-3-hydroxymethylbutyl)guanine (18F-FHBG) for in vivo imaging. Targeting was achieved using a CpG-free human alpha fetoprotein (AFP) promoter (CpGf-AFP-sr39). Expression was restricted to AFP-producing HCC cells, enabling selective transfection of orthotopic HCC xenografts. CpGf-AFP-sr39 NP treatment resulted in 62% reduced tumor size, and therapeutic gene expression was detectable by positron emission tomography (PET). This systemic nanomedicine achieved tumor-specific delivery, therapy, and imaging, representing a promising platform for targeted treatment of HCC.

Dendritic cell (DC)-based antigen-targeted immunotherapy may offer effective adjuvant therapy for hepatocellular carcinoma (HCC), in which cytotoxic T lymphocytes (CTLs) are key. However, in a number of cases, the activity of CTLs is completely inhibited due to the downregulated expression of major human leukocyte antigen (HLA) class I molecules by HCC cells. The aim of the present study was to overcome this issue. Hep3B cells were transduced by HCC‑specific recombinant adeno‑associated virus (rAAV) carrying human α‑fetoprotein promoter (AFPp) and the interferon‑γ (IFN‑γ) gene (rAAV/AFPp‑IFN‑γ). rAAV carrying the cytomegalovirus promoter (CMVp) and human α‑fetoprotein (AFP) gene (rAAV/CMVp‑AFP) was used to transduce professional antigen‑presenting DCs for the purpose of stimulating a CTL response. It was observed that transduction of DCs with rAAV/CMVp‑AFP resulted in: (i) AFP and interleukin‑12 expression; (ii) high expression levels of cluster of differentiation (CD)80, CD83, CD86, CD40, HLA‑death receptor and CD1a; (iii) T cell populations with marked IFN‑γ expression; (iv) a high percentage of CD69+/CD8+ T cells; and (v) the activity of CTLs against HLA‑A2‑expressing Hep3B cells. The transduction of Hep3B cells with rAAV/AFPp‑IFN‑γ resulted in: (i) IFN‑γ expression; (ii) upregulated expression of HLA‑A2; and (iii) an improved CTL response against HLA‑A2‑deficient Hep3B cells. rAAV/CMVp‑AFP‑transduced DCs elicited an AFP‑specific and HLA‑class I‑restricted CTL response against Hep3B cells. In conclusion, it was shown that the transduction of Hep3B with rAAV/AFPp-IFN-γ upregulated the expression of HLA-A2 and improved the sensitivity to CTL response.

Hepatocellular carcinoma (HCC) is one of the most common and difficult to treat types of cancer worldwide. Antigen‑targeted immunotherapy has the potential to be a novel and effective adjuvant for use in HCC. In the present study, recombinant adeno‑associated virus carrying the α‑fetoprotein gene (rAAV/AFP) and cancer cell lysates were used to pulse antigen‑presenting dendritic cells (DCs) in order to stimulate a cytotoxic T lymphocyte (CTL) response against HCC. rAAV/AFP‑pulsed and cancer cell lysate‑pulsed DCs resulted in a mature DC phenotype with high expression of major histocompatibility complex (MHC) class I, MHC class II, CD80, CD83 and CD86 molecules. However, rAAV/AFP‑pulsed DCs exhibited superiority over cancer cell lysate‑pulsed DCs in terms of stimulating proliferation of T cells, activating T cells to secret interferon‑γ (IFN‑γ) and inducing an AFP‑specific MHC class I‑restricted CTL response. The current data suggest that pulsing of DCs using rAAV/AFP is more effective than the cancer cell lysate‑pulsing technique, and that this technique may be used for the development of immunotherapy in AFP‑positive HCC.