Background: Growth of solid tumours beyond 1mm3 requires the recruitment of new blood vessels from the host (angiogenesis). Multiple studies have correlated neovascularisation with aggressive tumour behaviour, and factors driving the process such as VEGF-A, have been identified. However, this process is not well understood, and new molecular targets are required to improve upon existing first generation anti-angiogenic therapies.
Aim: Use a novel mutagenesis strategy to identify previously unrecognised modulators of tumour angiogenesis and compare this to normal physiological angiogenic processes.
Methods: Sleeping Beauty (SB) is a random insertional mutagenesis system for modifying cellular behaviour. Mutation of the endothelial genome was achieved by using the Tie-2 promoter/enhancer in a CRE transgenic mouse model to activate SB. IHC for SB confirmed the endothelial-specific nature of this activation. Allografts of B16F10 malignant melanoma cells were implanted in 8-10 week-old mice. Mice were sacrificed 10 or 14 days post implantation and tissues formalin fixed, paraffin embedded and analysed histologically. Vascular density was assessed with CD31 and LYVE-1 immunohistochemistry.
Results: No significant difference in growth or organ weight was observed between CRE+ (n=44) and CRE- (n=23) mice indicating that endothelial mutagenesis had no effect on physiological angiogenesis. Approximately 45% of CRE+ mice developed tumours ranging in size from large to small. In many of these mice an irregular relationship between vascular density and tumour size was observed indicating that the SB-mediated mutagenesis altered angiogenic potential. This perturbation of neovascularisation was also observed in lymphatic vessels indicating that lymph-angiogenic potential was also altered.
Conclusion: Changes to the endothelial genome via SB insertional mutagenesis: (1) alters the angiogenic response, which directly translated into altered tumour growth; (2) perturbs the known correlation between tumour size and vascularity. Future work will identify the mutations responsible for these changes and reveal the identity of novel tumour-specific regulators of neovascularisation.