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. 2007 Sep;98(9):1381-7.
doi: 10.1111/j.1349-7006.2007.00542.x. Epub 2007 Jul 11.

Gene transfer of endostatin enhances the efficacy of doxorubicin to suppress human hepatocellular carcinomas in mice

Fengjun Liu  1 , Gang Tan, Jie Li, Xuesong Dong, Geoffrey W Krissansen, Xueying Sun
Affiliations

Gene transfer of endostatin enhances the efficacy of doxorubicin to suppress human hepatocellular carcinomas in mice

Fengjun Liu et al. Cancer Sci. 2007 Sep.

Abstract

Hepatocellular carcinoma (HCC) is one of the most common cancer-related causes of death, and is chemoresistant to anticancer drugs. Anti-angiogenic therapy has been shown to enhance the efficacy of chemotherapy to treat solid tumors. The aim of the present study was to determine whether endostatin, a potent antiangiogenic agent, could enhance the efficacy of doxorubicin to combat HCC. An endostatin expression plasmid was constructed and its expression in vitro and in vivo was detected after gene transfer. Recombinant endostatin inhibited angiogenesis in the chorioallantoic membrane assay, and showed synergistic effects with doxorubicin in inhibiting the in vitro proliferation of endothelial cells, but not that of tumor cells. Both endostatin gene therapy and doxorubicin suppressed the growth of subcutaneous human HepG2 tumors established in BALB/c nude mice, and tumor angiogenesis. Combination therapy with endostatin gene therapy and doxorubicin showed a stronger effect in suppressing tumor growth, and tumor angiogenesis, than the respective monotherapies. Gene transfer of endostatin down-regulated the expression of both hypoxia-inducible factor-1alpha and vascular endothelial growth factor (VEGF), whereas doxorubicin only down-regulated VEGF expression. Endostatin and doxorubicin synergized to down-regulate VEGF expression. Endostatin and doxorubicin combination therapy warrants investigation as a therapeutic strategy to combat HCC.

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Figures

Figure 1
Figure 1
Recombinant endostatin inhibits angiogenesis in the chorioallantoic membrane (CAM) of chicken eggs. (a) Western blot analysis of recombinant endostatin expression in COS‐1 cells transfected with the End‐pcDNA3.1 plasmid. Cell lysates (lanes 1 and 3) and concentrated conditioned media (lanes 2 and 4) of COS‐1 cells transfected with End‐pcDNA3.1 (lanes 1 and 2) and empty pcDNA3.1 plasmid (lanes 3 and 4) were western blotted with anti‐endostatin (upper panel) and anti‐tubulin (lower panel) antibodies. (b) Illustrated are representative photographs of CAM treated with conditioned media from cultures of COS‐1 cells transfected with pcDNA3.1 and End‐pcDNA3.1 plasmids. (c) The numbers of branch‐points of the blood vessels were counted. A significant difference between the End‐pcDNA3.1 and pcDNA3.1 groups is denoted by ‘*’.
Figure 2
Figure 2
Proliferation of human umbilical vein endothelial cells (HUVEC) and HepG2 cells in vitro. (a,b) HUVEC and HepG2 cells were incubated in RPMI medium containing different concentrations of doxorubicin. (c,d) HUVEC and HepG2 cells were incubated in the presence or absence of doxorubicin in RPMI medium containing different volumes of conditioned media from COS‐1 cells transfected with End‐pcDNA3.1 plasmid. Conditioned media from COS‐1 cells transfected with empty vector pcDNA3.1 served as control. The proliferation of HUVEC cells was assessed, and the proliferation index was calculated. A significant difference between End‐pcDNA3.1 and End‐pcDNA3.1 + doxorubicin treatment groups is denoted by ‘*’, and between pcDNA3.1 and End‐pcDNA3.1 treatment groups by ‘†’.
Figure 3
Figure 3
Intense expression of endostatin in situ after intratumoral gene transfer. Illustrated are representative tumor sections prepared 2 days following intratumoral gene transfer of (a) pcDNA3.1 and (b) End‐pcDNA3.1 plasmids. The sections were stained with an anti‐endostatin antibody. (c) Homogenates of tumors 2 (lane 2), 7 (lane 3) or 14 (lane 4) days following intratumoral injection of the End‐pcDNA3.1, or pcDNA3.1 (lane 1) plasmids were western blotted with either anti‐endostatin (upper panel) or ‐tubulin (lower panel) antibodies.
Figure 4
Figure 4
Endostatin gene therapy synergizes with doxorubicin to suppress tumor growth. HepG2 hepatomas were established. When the tumors reached approximately 100 mm3 (indicated by a vertical arrow), they received pcDNA3.1, pcDNA3.1 + doxorubicin, End‐pcDNA3.1, or End‐pcDNA3.1 + doxorubicin treatments. Untreated tumors served as controls. The sizes (mm3) of tumors were monitored and recorded. A significant difference in tumor volumes from control is denoted by ‘*’, and a highly significant difference by ‘†’. ‘‡’ Indicates significant difference from End‐pcDNA3.1 or doxorubicin treatments.
Figure 5
Figure 5
Endostatin gene transfer synergizes with doxorubicin to inhibit tumor angiogenesis. Illustrated are representative tumor sections prepared 2 weeks after treatment from mice receiving (a) pcDNA3.1 (control), (b) End‐pcDNA3.1, (c) doxorubicin, or (d) doxorubicin + End‐pcDNA3.1 treatment. (e) Tumor microvessels in sections were stained with the anti‐CD31 antibody and counted in blindly chosen random fields to record microvessel density. A significant difference in microvessel density between tumors treated with doxorubicin, or End‐pcDNA3.1 versus control is denoted by ‘*’, and a highly significant difference between tumors treated with combinational therapy with doxorubicin + End‐pcDNA3.1 and control by ‘**’. (f) Homogenates of tumors from mice treated with pcDNA3.1 (lane 1), End‐pcDNA3.1 (lane 2), doxorubicin (lane 3), or doxorubicin + End‐pcDNA3.1 (lane 4) were western blotted with anti‐ hypoxia‐inducible factor (HIF)‐1α (upper panel), ‐vascular endothelial growth factor (VEGF; middle panel) and ‐tubulin (lower panel) antibodies. (g) Western blot analysis of HIF‐1α, VEGF and endostatin expression in HepG2 cells in vitro. HepG2 cells were transfected with End‐pcDNA3.1, followed by CoCl2 treatment. Cell lysates were western blotted with anti‐HIF‐1α, VEGF, endostatin and tubulin antibodies.
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References

    1. Befeler AS, Di Bisceglie AM. Hepatocellular carcinoma: diagnosis and treatment. Gastroenterology 2002; 122: 1609–19. - PubMed
    1. Schafer DF, Sorrell MF. Hepatocellular carcinoma. Lancet 1999; 353: 1253–7. - PubMed
    1. Parkin DM, Bray F, Ferlay J, Pisani P. Global cancer statistics, 2002. CA Cancer J Clin 2005; 55: 74–108. - PubMed
    1. El‐Assal ON, Yamanoi A, Soda Y, Yamaguchi M, Yu L, Nagasue N. Proposal of invasiveness score to predict recurrence and survival after curative hepatic resection for hepatocellular carcinoma. Surgery 1997; 122: 571–7. - PubMed
    1. Zhu AX. Systemic therapy of advanced hepatocellular carcinoma: how hopeful should we be? Oncologist 2006; 11: 790–800. - PubMed

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