Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Blockade of Dll4 inhibits tumour growth by promoting non-productive angiogenesis

Abstract

Tumour growth requires accompanying expansion of the host vasculature, with tumour progression often correlated with vascular density. Vascular endothelial growth factor (VEGF) is the best-characterized inducer of tumour angiogenesis. We report that VEGF dynamically regulates tumour endothelial expression of Delta-like ligand 4 (Dll4), which was previously shown to be absolutely required for normal embryonic vascular development. To define Dll4 function in tumour angiogenesis, we manipulated this pathway in murine tumour models using several approaches. Here we show that blockade resulted in markedly increased tumour vascularity, associated with enhanced angiogenic sprouting and branching. Paradoxically, this increased vascularity was non-productive—as shown by poor perfusion and increased hypoxia, and most importantly, by decreased tumour growth—even for tumours resistant to anti-VEGF therapy. Thus, VEGF-induced Dll4 acts as a negative regulator of tumour angiogenesis; its blockade results in a striking uncoupling of tumour growth from vessel density, presenting a novel therapeutic approach even for tumours resistant to anti-VEGF therapies.

This is a preview of subscription content

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Dll4 is expressed in tumour vessels, and its expression is dependent on VEGF signalling.
Figure 2: Blockade of Dll4/Notch signalling results in smaller C6 tumours with increased vessel density.
Figure 3: Despite an increase in blood vessel density, tumours overexpressing Dll4–Fc have increased hypoxia and poor vascular perfusion.
Figure 4: Systemic delivery of Dll4–Fc using adenovirus results in smaller C6 tumours and increased vessel density, similar to effects of local tumour overexpression.
Figure 5: Systemic delivery of Dll4–Fc or blocking Dll4 antibodies to mice bearing tumours that are resistant to blockade of VEGF results in decreased tumour growth and dramatic changes in tumour vessels.

References

  1. 1

    Folkman, J. The role of angiogenesis in tumor growth. Semin. Cancer Biol. 3, 65–71 (1992)

    CAS  PubMed  Google Scholar 

  2. 2

    Ferrara, N. Vascular endothelial growth factor as a target for anticancer therapy. Oncologist 9, (Suppl. 1)2–10 (2004)

    CAS  Article  Google Scholar 

  3. 3

    Rudge, J. S. et al. VEGF trap as a novel antiangiogenic treatment currently in clinical trials for cancer and eye diseases, and VelociGene-based discovery of the next generation of angiogenesis targets. Cold Spring Harb. Symp. Quant. Biol. 70, 411–418 (2005)

    CAS  Article  Google Scholar 

  4. 4

    Holash, J. et al. VEGF-Trap: a VEGF blocker with potent antitumor effects. Proc. Natl Acad. Sci. USA 99, 11393–11398 (2002)

    CAS  Article  ADS  Google Scholar 

  5. 5

    Hurwitz, H. et al. Bevacizumab plus irinotecan, fluorouracil, and leucovorin for metastatic colorectal cancer. N. Engl. J. Med. 350, 2335–2342 (2004)

    CAS  Article  Google Scholar 

  6. 6

    Laskin, J. J. & Sandler, A. B. First-line treatment for advanced non-small-cell lung cancer. Oncology 19, 1671–6; discussion 1678–80. (2005)

    PubMed  Google Scholar 

  7. 7

    Casanovas, O., Hicklin, D. J., Bergers, G. & Hanahan, D. Drug resistance by evasion of antiangiogenic targeting of VEGF signaling in late-stage pancreatic islet tumors. Cancer Cell 8, 299–309 (2005)

    CAS  Article  Google Scholar 

  8. 8

    Jain, R. K., Duda, D. G., Clark, J. W. & Loeffler, J. S. Lessons from phase III clinical trials on anti-VEGF therapy for cancer. Nature Clin. Pract. Oncol. 3, 24–40 (2006)

    CAS  Article  Google Scholar 

  9. 9

    Kerbel, R. S. et al. Possible mechanisms of acquired resistance to anti-angiogenic drugs: implications for the use of combination therapy approaches. Cancer Metastasis Rev. 20, 79–86 (2001)

    CAS  Article  Google Scholar 

  10. 10

    Yancopoulos, G. D. et al. Vascular-specific growth factors and blood vessel formation. Nature 407, 242–248 (2000)

    CAS  Article  Google Scholar 

  11. 11

    Jain, R. K. Normalization of tumor vasculature: an emerging concept in antiangiogenic therapy. Science 307, 58–62 (2005)

    CAS  Article  ADS  Google Scholar 

  12. 12

    Carmeliet, P. Angiogenesis in life, disease and medicine. Nature 438, 932–936 (2005)

    CAS  Article  ADS  Google Scholar 

  13. 13

    Shawber, C. J. & Kitajewski, J. Notch function in the vasculature: insights from zebrafish, mouse and man. Bioessays 26, 225–234 (2004)

    CAS  Article  Google Scholar 

  14. 14

    Artavanis-Tsakonas, S., Rand, M. D. & Lake, R. J. Notch signaling: cell fate control and signal integration in development. Science 284, 770–776 (1999)

    CAS  Article  ADS  Google Scholar 

  15. 15

    Gridley, T. Notch signaling during vascular development. Proc. Natl Acad. Sci. USA 98, 5377–5378 (2001)

    CAS  Article  ADS  Google Scholar 

  16. 16

    Duarte, A. et al. Dosage-sensitive requirement for mouse Dll4 in artery development. Genes Dev. 18, 2474–2478 (2004)

    CAS  Article  Google Scholar 

  17. 17

    Gale, N. W. et al. Haploinsufficiency of delta-like 4 ligand results in embryonic lethality due to major defects in arterial and vascular development. Proc. Natl Acad. Sci. USA 101, 15949–15954 (2004)

    CAS  Article  ADS  Google Scholar 

  18. 18

    Krebs, L. T. et al. Haploinsufficient lethality and formation of arteriovenous malformations in Notch pathway mutants. Genes Dev. 18, 2469–2473 (2004)

    CAS  Article  Google Scholar 

  19. 19

    Mailhos, C. et al. Delta4, an endothelial specific notch ligand expressed at sites of physiological and tumor angiogenesis. Differentiation 69, 135–144 (2001)

    CAS  Article  Google Scholar 

  20. 20

    Patel, N. S. et al. Up-regulation of delta-like 4 ligand in human tumor vasculature and the role of basal expression in endothelial cell function. Cancer Res. 65, 8690–8697 (2005)

    CAS  Article  Google Scholar 

  21. 21

    Hicks, C. et al. A secreted Delta1–Fc fusion protein functions both as an activator and inhibitor of Notch1 signaling. J. Neurosci. Res. 68, 655–667 (2002)

    CAS  Article  Google Scholar 

  22. 22

    Taylor, K. L., Henderson, A. M. & Hughes, C. C. Notch activation during endothelial cell network formation in vitro targets the basic HLH transcription factor HESR-1 and downregulates VEGFR-2/KDR expression. Microvasc. Res. 64, 372–383 (2002)

    CAS  Article  Google Scholar 

  23. 23

    Iso, T., Kedes, L. & Hamamori, Y. HES and HERP families: multiple effectors of the Notch signaling pathway. J. Cell. Physiol. 194, 237–255 (2003)

    CAS  Article  Google Scholar 

  24. 24

    Shawber, C. J., Das, I., Francisco, E. & Kitajewski, J. Notch signaling in primary endothelial cells. Ann. NY Acad. Sci. 995, 162–170 (2003)

    CAS  Article  ADS  Google Scholar 

  25. 25

    Karsan, A. The role of notch in modeling and maintaining the vasculature. Can. J. Physiol. Pharmacol. 83, 14–23 (2005)

    CAS  Article  ADS  Google Scholar 

  26. 26

    Lamar, E. et al. Nrarp is a novel intracellular component of the Notch signaling pathway. Genes Dev. 15, 1885–1899 (2001)

    CAS  Article  Google Scholar 

  27. 27

    Krebs, L. T., Deftos, M. L., Bevan, M. J. & Gridley, T. The Nrarp gene encodes an ankyrin-repeat protein that is transcriptionally regulated by the Notch signaling pathway. Dev. Biol. 238, 110–119 (2001)

    CAS  Article  Google Scholar 

  28. 28

    Krneta, J. et al. Dissociation of angiogenesis and tumorigenesis in follistatin- and activin-expressing tumors. Cancer Res. 66, 5686–5695 (2006)

    CAS  Article  Google Scholar 

  29. 29

    Lee, C. G. et al. Anti-vascular endothelial growth factor treatment augments tumor radiation response under normoxic or hypoxic conditions. Cancer Res. 60, 5565–5570 (2000)

    CAS  PubMed  Google Scholar 

  30. 30

    Jain, R. K. Tumor angiogenesis and accessibility: role of vascular endothelial growth factor. Semin. Oncol. 29, 3–9 (2002)

    CAS  Article  Google Scholar 

  31. 31

    Jain, R. K. Antiangiogenic therapy for cancer: current and emerging concepts. Oncology. 19, 7–16 (2005)

    PubMed  Google Scholar 

  32. 32

    Valenzuela, D. M. et al. High-throughput engineering of the mouse genome coupled with high-resolution expression analysis. Nature Biotechnol. 21, 652–659 (2003)

    CAS  Article  Google Scholar 

  33. 33

    Thurston, G., Baluk, P., Hirata, A. & McDonald, D. M. Permeability-related changes revealed at endothelial cell borders in inflamed venules by lectin binding. Am. J. Physiol. 271, H2547–H2562 (1996)

    CAS  PubMed  Google Scholar 

  34. 34

    Liu, Z. J. et al. Inhibition of endothelial cell proliferation by Notch1 signaling is mediated by repressing MAPK and PI3K/Akt pathways and requires MAML1. FASEB J. 20, 1009–1011 (2006)

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We acknowledge the following Regeneron colleagues: Y. Wei for gene expression analysis, A. Adler, A. Rafique, B. Li, H. Huang, E. Pasnikowski, J. McClain, E. Burova, D. Hylton, P. Burfeind and J. Griffiths for technical assistance, S. Staton for assistance with graphics, and S. Wiegand, I. Lobov, T. Daly, S. Davis, E. Ioffe, J. Holash and J. Rudge for scientific input.

Author Contributions I. N.-T. directed and helped perform tumour experiments, generation of tumour lines, immunohistochemical staining, and data analysis. C.D. directed, helped perform, and analysed in vitro experiments. N.J.P. helped develop protein reagents and biochemical assays. S.C. performed and helped analyse tumour experiments and construction of tumour cell lines. P.B. performed and helped analyse immunohistochemical studies. N.W.G. helped perform and analyse experiments with gene-targeted mice. H.C.L. helped perform and analyse gene expression studies. G.D.Y. helped analyse and interpret results. G.T. helped design experiments, analyse data and interpret results.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Gavin Thurston.

Ethics declarations

Competing interests

All authors are employees of Regeneron Pharmaceuticals.

Supplementary information

Supplementary information

This file contains Supplementary Methods and Supplementary Figures 1-9 with legends.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Noguera-Troise, I., Daly, C., Papadopoulos, N. et al. Blockade of Dll4 inhibits tumour growth by promoting non-productive angiogenesis. Nature 444, 1032–1037 (2006). https://doi.org/10.1038/nature05355

Download citation

Further reading

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Quick links