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.

  • Research Article
  • Published:

Ablation of microvessels in vivo upon dimerization of iCaspase-9

Gene Therapy volume 9, pages 444–451 (2002)Cite this article

Abstract

Anti-angiogenic therapies based on targeted disruption of the tumor microvascular network have been proposed for cancer treatment. Inhibitors of the endothelial cell pro-survival pathway mediated by VEGF were shown to activate caspases and cause microvascular regression, but the efficacy of this strategy can be hindered by the engagement of redundant survival pathways. Alternatively, if direct activation of an apical pro-apoptotic caspase is sufficient to disrupt microvessels in vivo, such a strategy could potentially override upstream endothelial cell survival inputs and disrupt tumor neovascular networks. Here, we fused caspase-9 to a mutated FKBP12 domain to express an inducible caspase-9 molecule (iCaspase-9) that can be activated by a cell-permeable dimerizer drug, and transduced this construct into primary endothelial cells. We found that drug-induced dimerization of iCaspase-9 is sufficient to activate endogenous caspase-3 and trigger apoptosis even when endothelial cells are treated with the pro-survival factors VEGF or bFGF. A single intraperitoneal injection of the dimerizer drug induced apoptosis of endothelial cells expressing iCaspase-9 and elimination of human microvessels engineered in immunodeficient mice. These results demonstrate that the activation of iCaspase-9 disrupts microvessels in vivo, and suggest a novel anti-angiogenic strategy based on the expression and controlled activation of an inducible death gene in neovascular endothelial cells.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6

Similar content being viewed by others

References

  1. Nör J.E., Polverini P.J. . Role of endothelial cell survival and death signals in angiogenesis Angiogenesis 1999 3: 101–101

    Article  Google Scholar 

  2. Gerber H.B., Dixit V., Ferrara N. . Vascular endothelial growth factor induces expression of the antiapoptotic proteins Bcl-2 and A1 in vascular endothelial cells J Biol Chem 1998 273: 13313–13313

    Article  CAS  Google Scholar 

  3. Nör J.E., Christensen J., Mooney D.J., Polverini P.J. . Vascular endothelial growth factor (VEGF)-mediated angiogenesis is associated with enhanced endothelial cell survival and induction of Bcl-2 expression Am J Pathol 1999 154: 375–375

    Article  Google Scholar 

  4. Gerber H.P. et al. Vascular endothelial growth factor regulates endothelial cell survival through the phosphatidylinositol 3’-kinase/Akt signal transduction pathway. Requirement for Flk-1/KDR activation J Biol Chem 1998 273: 30336–30336

    Article  CAS  Google Scholar 

  5. Papapetropoulos A. et al. Angiopoietin-1 inhibits endothelial cell apoptosis via the Akt/survivin pathway J Biol Chem 2000 275: 9102–9102

    Article  CAS  Google Scholar 

  6. Araki S., Simada Y., Kaji K., Hayashi H. . Role of protein kinase C in the inhibition by fibroblast growth factor of apoptosis in serum-depleted endothelial cells Biochem Biophys Res Commun 1990 172: 1081–1081

    Article  CAS  Google Scholar 

  7. Stromblad S., Cheresh D.A. . Integrins, angiogenesis and vascular cell survival Chem Biol 1996 3: 88–88

    Article  Google Scholar 

  8. Benjamin L.E., Keshet E. . Conditional switching of vascular endothelial growth factor expression in tumors: induction of endothelial cell shedding and regression of hemangioblastoma-like vessels by VEGF withdrawal Proc Natl Acad Sci USA 1997 94: 8761–8761

    Article  CAS  Google Scholar 

  9. Nör J.E. et al. Up-regulation of Bcl-2 in microvascular endothelial cells enhances intratumoral angiogenesis and accelerates tumor growth Cancer Res 2001 61: 2183–2183

    PubMed  Google Scholar 

  10. Shaheen R.M. et al. Tyrosine kinase inhibition of multiple angiogenic growth factor receptors improves survival in mice bearing colon cancer liver metastases by inhibition of endothelial cell survival mechanisms Cancer Res 2001 61: 1464–1464

    CAS  PubMed  Google Scholar 

  11. Detmar M. et al. Overexpression of vascular permeability factor/vascular endothelial growth factor and its receptors in psoriasis J Exp Med 1994 180: 1141–1141

    Article  CAS  Google Scholar 

  12. Nickoloff B.J. et al. Aberrant production of interleukin-8 and thrombospondin-1 by psoriatic keratinocytes mediates angiogenesis Am J Pathol 1994 144: 820–820

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Alon T. et al. Vascular endothelial growth factor acts as a survival factor for newly formed retinal vessels and has implications for retinopathy of prematurity Nat Med 1995 1: 1024–1024

    Article  CAS  Google Scholar 

  14. Bousvaros A. et al. Elevated serum vascular endothelial growth factor in children and young adults with Crohn's disease Dig Dis Sci 1999 44: 424–424

    Article  CAS  Google Scholar 

  15. Kerbel R.S. . Tumor angiogenesis: past, present, and the near future Carcinogenesis 2000 21: 505–505

    Article  CAS  Google Scholar 

  16. Browder T. et al. Anti-angiogenic scheduling of chemotherapy improves efficacy against experimental drug-resistant cancer Cancer Res 2000 60: 1878–1878

    CAS  Google Scholar 

  17. Klement G. et al. Continuous low-dose therapy with vinblastine and VEGF receptor-2 antibody induces sustained tumor regression without overt toxicity J Clin Invest 2000 105: R15–R15

    Article  CAS  Google Scholar 

  18. MacCorkle R.A., Freeman K.W., Spencer D.M. . Synthetic activation of caspases: artificial death switches Proc Natl Acad Sci USA 1998 95: 3655–3655

    Article  CAS  Google Scholar 

  19. Clackson T. . Redesigning small molecule–protein interfaces Curr Opin Struct Biol 1998 8: 451–451

    Article  CAS  Google Scholar 

  20. Hu Y., Ding L., Spencer D.M., Nuñez G. . WD-40 repeat region regulates Apaf-1 self-association and procaspase-9 activation J Biol Chem 1998 273: 33489–33489

    Article  CAS  Google Scholar 

  21. Fan L. et al. Improved artificial death switches based on caspases and FADD Hum Gene Ther 1999 10: 2273–2273

    Article  CAS  Google Scholar 

  22. Jin L. et al. In vivo selection using a cell-growth switch Nat Genet 2000 26: 64–64

    Article  CAS  Google Scholar 

  23. Li P. et al. Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade Cell 1997 91: 479–479

    Article  CAS  Google Scholar 

  24. Nuñez G., Benedict M.A., Hu Y., Inohara N. . Caspases: the proteases of the apoptotic pathway Oncogene 1998 17: 3237–3237

    Article  Google Scholar 

  25. Nör J.E. et al. Engineering and characterization of functional human microvessels in immunodeficient mice Lab Invest 2001 81: 453–453

    Article  Google Scholar 

  26. Folkman J. . Tumor angiogenesis: therapeutic implications N Engl J Med 1971 285: 1182–1182

    Article  CAS  Google Scholar 

  27. Folkman J. . Angiogenesis in cancer, vascular, rheumatoid and other disease Nat Med 1995 1: 27–27

    Article  CAS  Google Scholar 

  28. O'Reilly M.S. et al. Endostatin: an endogenous inhibitor of angiogenesis and tumor growth Cell 1997 88: 277–277

    Article  CAS  Google Scholar 

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

    CAS  PubMed  Google Scholar 

  30. Korhonen J. et al. Endothelial-specific gene expression directed by the Tie gene promoter in vivo Blood 1995 86: 1828–1828

    CAS  Google Scholar 

  31. Gory S. et al. Vascular endothelial-cadherin promoter directs endothelial-specific expression in transgenic mice Blood 1999 93: 184–184

    CAS  PubMed  Google Scholar 

  32. St. Croix B. et al. Genes expressed in human tumor endothelium Science 2000 289: 1197–1197

    Article  CAS  Google Scholar 

  33. Reynolds P.N. et al. A targetable, injectable adenoviral vector for selective gene delivery to pulmonary endothelium in vivo Mol Ther 2000 2: 562–562

    Article  CAS  Google Scholar 

  34. Arap W., Pasqualini R., Ruoslahti E. . Cancer treatment by targeted drug delivery to tumor vasculature in a mouse model Science 1998 279: 377–377

    Article  CAS  Google Scholar 

  35. Trepel M., Grifman M., Weitzman M.D., Pasqualini R. . Molecular adaptors for vascular-targeted adenoviral gene delivery Hum Gene Ther 2000 11: 1971–1971

    Article  CAS  Google Scholar 

  36. Nicklin S.A. et al. Selective targeting of gene transfer to vascular endothelial cells by use of peptides isolated by phage display Circulation 2000 102: 231–231

    Article  CAS  Google Scholar 

  37. Nör J.E. et al. Thrombospondin-1 induces endothelial cell apoptosis and inhibits angiogenesis by activating the caspase death pathway J Vasc Res 2000 37: 209–209

    Article  Google Scholar 

  38. Nicholson D.W. et al. Identification and inhibition of the ICE/CED-3 protease necessary for mammalian apoptosis Nature 1995 376: 37–37

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank Y Lazebnik for the gift of monoclonal antibodies, AD Miller for the gift of retroviral vector, and ARIAD Pharmaceuticals for the dimerizing agent AP20187. We thank J Westman and C Strayhorn for preparation of histological sections; M Kukuruga for help with flow cytometry; and T Ross and P Lucas for critical review of this manuscript. This research was supported by grants CA70057 (to GN) and CA77266 (to DMS) from the National Institutes of Health; and the University of Michigan Comprehensive Cancer Center's Institutional Grant from the American Cancer Society and start-up funds from the University of Michigan School of Dentistry (to JEN).

Author information

Authors and Affiliations

  1. Department of Cariology, Restorative Sciences and Endodontics, University of Michigan School of Dentistry, Ann Arbor, MI, USA

    JE Nör & W Song

  2. Comprehensive Cancer Center, University of Michigan School of Medicine, Ann Arbor, MI, USA

    JE Nör & G Núñez

  3. Department of Pathology, University of Michigan School of Medicine, Ann Arbor, MI, USA

    Y Hu & G Núñez

  4. Department of Immunology, Baylor College of Medicine, Houston, TX, USA

    DM Spencer

Authors
  1. JE Nör
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Y Hu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. W Song
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. DM Spencer
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. G Núñez
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Nör, J., Hu, Y., Song, W. et al. Ablation of microvessels in vivo upon dimerization of iCaspase-9. Gene Ther 9, 444–451 (2002). https://doi.org/10.1038/sj.gt.3301671

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.gt.3301671

Keywords

This article is cited by

Search

Advanced search

Quick links