Abstract
We previously identified CLEC14A as a tumour endothelial marker. Here we show that CLEC14A is a regulator of sprouting angiogenesis in vitro and in vivo. Using a human umbilical vein endothelial cell spheroid-sprouting assay, we found CLEC14A to be a regulator of sprout initiation. Analysis of endothelial sprouting in aortic ring and in vivo subcutaneous sponge assays from clec14a+/+ and clec14a−/− mice revealed defects in sprouting angiogenesis in CLEC14A-deficient animals. Tumour growth was retarded and vascularity reduced in clec14a−/− mice. Pull-down and co-immunoprecipitation experiments confirmed that MMRN2 binds to the extracellular region of CLEC14A. The CLEC14A-MMRN2 interaction was interrogated using mouse monoclonal antibodies. Monoclonal antibodies were screened for their ability to block this interaction. Clone C4, but not C2, blocked CLEC14A-MMRN2 binding. C4 antibody perturbed tube formation and endothelial sprouting in vitro and in vivo, with a similar phenotype to loss of CLEC14A. Significantly, tumour growth was impaired in C4-treated animals and vascular density was also reduced in the C4-treated group. We conclude that CLEC14A-MMRN2 binding has a role in inducing sprouting angiogenesis during tumour growth, which has the potential to be manipulated in future antiangiogenic therapy design.
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Acknowledgements
We thank Dr Laurens van der Flier, Somantix and Dr Raj Mehta, Cancer Research Technology, for helpful discussions. PN was funded by Cancer Research UK (CRUK, grant number C4719/A6766 to RB). The Portuguese Fundacao para a Ciencia e tecnologia funded AV. XZ was funded by a knowledge transfer partnership (KTP007696) from the Technology Strategy Board. PL is funded by MRC and the University of Birmingham, and KK is funded by the University of Birmingham. The University of Birmingham Transgenic Mouse Facility is part of the MRC Centre for Immune Regulation.
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RB and XZ are named inventors of a patent filed by Cancer Research UK in the United States Patent and trademark Office on 3 September 2009 under No. 61/ 239,584, bearing Attorney Docket No. P0357.70004US00 and entitled ‘Inhibitors’. All the other authors declare no conflict of interest.
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Noy, P., Lodhia, P., Khan, K. et al. Blocking CLEC14A-MMRN2 binding inhibits sprouting angiogenesis and tumour growth. Oncogene 34, 5821–5831 (2015). https://doi.org/10.1038/onc.2015.34
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DOI: https://doi.org/10.1038/onc.2015.34
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