Cocktail gene therapy for hepatocellular carcinoma and liver cirrhosis
Date Issued
2011
Date
2011
Author(s)
Huang, Kai-Wen
Abstract
Background & Aims: Cytokine and antiangiogenic gene therapies have proven effective in many implanted hepatocellular carcinoma(HCC) models with small tumor burdens. However, these models may not realistically reflect human HCCs, which are typically large, multifocal and often noted on a background of hepatitis or liver cirrhosis. So, to develop anti-HCC therapeutic strategies, an animal model bearing multifocal liver tumors accompanied with viral hepatitis or cirrhosis is a preferred experimental setting. This study utilized woodchucks to investigate the efficacy of a cocktail-type gene therapy strategy. The animals bore spontaneously-occurring HCCs induced by woodchuck hepatitis virus infection, and therefore closely resembled human HCCs. In the second part, we tried to develop a tumor-bearing rat model with liver cirrhosis. By adjusting the duration of administration of diethylnitrosamine (DEN), the animals could be induced to develop HCC alone, or HCC and liver cirrhosis simultaneously.
Methods: Adenoviruses encoding antiangiogenic factors [pigment epithelium-derived factor and endostatin] or cytokines [granulocyte macrophage colony-stimulating factor and interleukin ] were delivered, either separately or in combination, into woodchuck livers bearing HCCs via the intrahepatic artery. Tumor sizes were monitored by sequential abdominal ultrasound examination and CT. To develop HCC-bearing cirrhotic animal model, we modified the feeding protocol by adjusting the DEN intake amount weekly based on BW of the animals. The results show that animals fed with DEN for 6 consecutive weeks develop multifocal HCC with normal liver parenchyma; whereas animals fed with DEN for 9 consecutive weeks develop multifocal HCC complicated with moderate cirrhosis. The tumor burdens between these two groups are comparable, determined by the ratio of liver weight/BW, the estimated volume of all visible tumor nodules, and the density of glutathione S-transferase placental form-positive foci. On the other hand, liver cirrhosis occurs slightly and transiently in the former group, but severely and irreversibly in the latter group, shown by immunohistochemical staining of α-smooth muscle actin, the bile flow rate, and the liver hydroxyproline content, then adenoviruses encoding interferon-α(IFN-α) were delivered into rats in the latter group for treatment of liver cirrhosis and HCC simultaneously.
Results: Our results demonstrate that monotherapy with antiangiogenic genes or cytokine genes had significant therapeutic effects. Furthermore, cocktail-type therapies containing both types of genes displayed synergistic antitumor effects under the same viral doses, particularly for large tumor loads. Effector cell analyses based on flow cytometry demonstrate that the cytokine-based gene therapies resulted in increased CD3+ T-cell infiltration of the HCCs. Examination of microvessel density also showed that tumor vasculature was significantly reduced by the antiangiogenesis-based gene therapies. No evidence of adenovirus-induced hepatic toxicity was found in these animals. As for the research about treatment of liver cirrhosis, Our results demonstrated that targeting of IFN-α expression to the liver significantly reduced liver tumor volume and ameliorated liver cirrhosis. Mechanistic studies revealed that IFN-α gene therapy induced immunomodulatory, antiproliferative, and proapoptotic activities that were effective in the control of tumor growth, and reduced the expressions of transforming growth factor-β (TGF-β) and tissue inhibitor of metalloproteinase-1 (TIMP-1), leading to amelioration of liver cirrhosis.
Conclusions: The combination of immunotherapy and antiangiogenic gene therapy exhibited the most significant antitumor effect compared to either therapy alone. This indicates a strong synergism induced by the combination of two different therapeutic strategies. Using the clinically relevant model, future clinical application as a neoadjuvant therapy would be encouraged. Except the woodchuck model, the HCC-bearing rat model with cirrhosis is an ideal model to study therapeutic strategies that can treat both diseases simultaneously, and our results suggest that IFN-α gene therapy is a promising strategy to treat HCC patients who have concomitant liver cirrhosis.
Subjects
antiangiogensis
cirrhosis
gene therapy
hepatocellular carcinoma
immunotherapy
interferon-α
woodchuck
SDGs
Type
thesis
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