Expression of thymidine kinase driven by an endothelial-specific promoter inhibits tumor growth of Lewis lung carcinoma cells in transgenic mice

Abstract

The possibility of inhibiting tumor growth by limiting angiogenesis has raised considerable interest. In this study, we examined the feasibility of inhibiting tumor growth by targeting a suicide gene in the endothelium. Toxicity must be directed solely to angiogenic cells. Therefore, we used the herpes simplex virus-thymidine kinase (TK) gene, in combination with the prodrug ganciclovir (GCV), which affects replicative cells. To test this strategy, we produced transgenic mice carrying the TK gene driven by the vascular endothelial (VE)-cadherin promoter. Lewis lung carcinoma cells were injected subcutaneously to establish tumors and to test the effect of GCV on tumor growth. In two independent transgenic lines, GCV treatment (75 mg/kg/day) resulted in a 66-71% reduction of tumor volume at day 20 postimplantation compared to wild-type mice (650 and 550 versus 1930 mm 3 , Po0.02 and 0.01, respectively), whereas no significant difference was observed when vehicle alone was injected. Tumor growth inhibition was accompanied by a marked reduction in tumor vascular density (151 versus 276 vessels/mm 2 , Po0.05) and an increase in tumor cell death, suggesting that tumor growth inhibition was caused by a reduction in tumor angiogenesis. Our data support the potential utility of endothelial targeting of suicide genes in cancer therapy.

The human VE-cadherin promoter is subjected to organ-specific regulation and is activated in tumour angiogenesis

“…Interestingly, b-galactosidase activity was particularly high in angiogenic situations, namely in ovaries, Matrigel plugs impregnated with bFGF, tumours and during development. Similarly, the mouse promoter showed strong activity in tumour endothelium (Dancer et al, 2003). The fact that we observed angiogenic responsiveness with the hVE- …”

“…Lewis lung carcinoma cells were implanted as described (Dancer et al, 2003). Mice were killed when tumour volume reached 500 mm 3 .…”

“…This region contains Ets and Sp1 binding sites essential for its activity (Gory et al, 1998). Recently, we used this promoter to drive herpes virus-thymidine kinase gene expression in the endo thelium of transgenic mice (Dancer et al, 2003). Thymidine kinase immunohistological data revealed that transgene expression was high in tumour vasculature but weak in normal tissues, except in the endocardium.…”

VE-Cadherin

“…7 A roughly 2.5-kb region of the promotor was characterized to direct endothelial expression of reporter genes in vivo, although expression was not found in all endothelial cells. 8,9,10 Stronger endothelial expression was observed, when another 4 kb from the 5Ј half of the first intron were added to the regulatory sequences. 11 The first evidence for the function of VE-cadherin in vivo was established with antibodies.…”

Gene therapy targeting to tumor endothelium

“…Adenoviral expression of HSV-TK under the control of VE-cadherin promoter in vivo inhibited tumor growth and tumor vascular density. 59 Another adenovirus expressing HSV-TK under the control of an optimized hypoxia responsive promoter (OBHRE) derived from phosphoglycerate kinase 1 promoter is well tolerated in liver after intravenous administration contrary to administration of Ad.CMV-TK/GCV. 60 In addition, an adenoviral vector simultaneously expressing both cytosine deaminase (CD) and HSV-TK genes under the transcriptional control of KDR promoter showed that this combined CD/TK therapy is more effective at killing HUVEC than with either treatment alone.…”