Combined antiangiogenic gene therapy with sflt and endostatin is highly effective in a rat model of hepatocellular carcinoma

The multiplicity of regulators acting in the process of tumor angiogenesis offers diverse molecular targets for an antiangiogenic therapy. On the other hand, therapeutical attempts to antagonize one of these players could be equalized by upregulation of another proangiogenic factor leading to some kind of resistance to antiangiogenic therapy. A combination therapy addressing two or more angiogenic targets could therefore be useful. Growth of Hepatocellular carcinoma, an uncurable disease in most cases so far, is supposed to be a suitable target for antiangiogenic therapy.

We tested the effects of expression of endostatin, a direct inhibitor of angiogenesis, and sflt, a soluble receptor for VEGF, as an indirect inhibitor of angiogenesis alone and in combination in our syngeneous rat-hepatoma model. The Morris-hepatoma cell-line 3924 (MH) showed a high expression of VEGF in the supernatant, but no endogeneous sflt-expression, as measured by ELISA. MH-cells were stably transduced using retroviral vector pLXSN-Green-Endostatin encoding rat endostatin (leading to cell-line MHES). For expression of sflt, adenoviral vector Adsflt was used. Cell-lines MH and MHES were transduced at MOI 50 either with Adsflt or a control-vector containing no transgene (Ad0). Next, 6*10E6 cells were injected subcutaneously in ACI rats (n=6 per group). At day 11 animals were sacrificed and tumors explanted. A multivariance analysis was performed on the tumor weights of the four groups. Compared to MH/Ad0 cells, expressing no antiangiogenic factors at all, tumor weight was reduced in the MHES/Ad0-group by factor 4.1 (KI 2.6 to 6.6), in the MH/Adsflt-group by factor 18.7 (KI 11.7 to 29.8) and in the MHES/sflt combination therapy group by factor 77 (KI 40 to 149). No significant interaction between the two factors ES and sflt was found.

In conclusion combined expression of sflt and endostatin most effectively suppresses outgrowth of hepatocellular carcinoma under in vivo conditions.

Supported by the German Research Council DFG GR 1838–1/2

Key words

Antiangiogenese - Gentherapie - HCC - Hepatozelluläres Karzinom - Tumor-Antiangiogenese