Adeno-associated virus-mediated expression of kallistatin suppresses local and remote hepatocellular carcinomas

Background The current treatments for hepatocellular carcinoma (HCC) are poor, particularly for metastatic HCC. Intraportal transfusion of adeno‐associated virus (AAV) leads to long‐term and persistent transgenic expression in livers. Kallistatin, a novel angiogenesis inhibitor, exhibits anti‐tumor activity. The aim of the study was to investigate whether intraportal injection of AAV‐kallistatin could suppress local and metastatic HCC in mice. Methods An AAV vector encoding kallistatin was constructed, and its transduction efficiency by intraportal transfusion in livers was examined by RT‐PCR, immunohistochemical and Western blotting analysis. The anti‐tumor activity was tested in three HCC models including hepatic and subcutaneous human Hep3B HCC tumors in BALB/c athymic (nu/nu) mice, and subcutaneous mouse BNL HCC tumors in BALB/c mice. Tumor cell proliferation in situ was examined by anti‐Ki‐67 staining, and apoptosis by TUNEL. Results Gene transfection by rAAV‐kallistatin inhibited proliferation of human umbilical vein endothelial cells and HCC cells in vitro. Intraportal injection of rAAV‐kallistatin resulted in persistent and specific expression of kallistatin in livers detected by RT‐PCR and immunohistochemical analysis, and kallistatin protein in circulation detected by Western blotting analysis. Intraportal injection of rAAV‐kallistatin significantly suppressed angiogenesis and growth of hepatic Hep3B tumors. The kallistatin released by hepatocytes into the circulation suppressed remote Hep3B and BNL tumors established subcutaneously. The rAAV‐kallistatin gene therapy significantly inhibited tumor cell proliferation and induced apoptosis. Conclusions Intraportal injection of rAAV‐kallistatin suppressed hepatic and subcutaneous HCC tumors, relying on its anti‐angiogenic and anti‐proliferative activities. Copyright © 2008 John Wiley & Sons, Ltd.