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Targeting Redundant ROBO1 and SDF-1 Pathways Prevents Adult Hemangioblast Derived-EPC and CEC Activity Effectively Blocking Tumor Neovascularization

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Abstract

Neovascularization is a key therapeutic target for cancer treatment. However, anti-angiogenic therapies have shown modest success, as tumors develop rapid resistance to treatment owing to activation of redundant pathways that aid vascularization. We hypothesized that simultaneously targeting different pathways of neovascularization will circumvent the current issue of drug resistance and offer enhanced therapeutic benefits. To test this hypothesis, we made use of two distinct models of tumor-neovascularization, which exhibit equally dense microvasculature but show disparate sensitivity to anti-SDF-1 treatment. Lewis lung carcinoma (LLC) is primarily a vasculogenic-tumor that is associated with HSC functioning as a hemangioblast to generate circulating Endothelial Progenitor Cells contributing to formation of new blood vessels, and responds to anti-SDF-1 treatment. B16F0 melanoma is an angiogenic-tumor that derives new blood vessels from existing vasculature and is resistant to anti-SDF-1 therapy. In this study, we observed increased expression of the angiogenic-factor, Robo1 predominantly expressed on the blood vessels of B16F0 tumor. Blockade of Robo1 by the decoy receptor, RoboN, resulted in reduced microvascular-density and tumor-growth. However, this was associated with mobilization of BM-cells into the B16F0 tumor, thus switching the mode of neovascularization from angiogenic to vasculogenic. The use of a combinatorial treatment of RoboN and the monoclonal anti-SDF-1 antibody effectively attenuated tumor-growth and inhibited both angiogenic and BM-derived microvessels.

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References

  1. Hanahan, D., & Weinberg, R. A. (2011). Hallmarks of cancer: The next generation. Cell, 144(5), 646–674. https://doi.org/10.1016/j.cell.2011.02.013

    Article  CAS  PubMed  Google Scholar 

  2. Jayson, G. C., Kerbel, R., Ellis, L. M., & Harris, A. L. (2016). Antiangiogenic therapy in oncology: Current status and future directions. Lancet (London, England), 388(10043), 518–529. https://doi.org/10.1016/S0140-6736(15)01088-0

    Article  CAS  PubMed  Google Scholar 

  3. De Palma, M., Biziato, D., & Petrova, T. V. (2017). Microenvironmental regulation of tumour angiogenesis. Nature Reviews Cancer, 17(8), 457–474. https://doi.org/10.1038/nrc.2017.51

    Article  CAS  PubMed  Google Scholar 

  4. Casanovas, O., Hicklin, D. J., Bergers, G., & Hanahan, D. (2005). Drug resistance by evasion of antiangiogenic targeting of VEGF signaling in late-stage pancreatic islet tumors. Cancer Cell, 8(4), 299–309. https://doi.org/10.1016/j.ccr.2005.09.005

    Article  CAS  PubMed  Google Scholar 

  5. Fischer, C., Jonckx, B., Mazzone, M., Zacchigna, S., Loges, S., Pattarini, L., Chorianopoulos, E., Liesenborghs, L., Koch, M., De Mol, M., Autiero, M., Wyns, S., Plaisance, S., Moons, L., van Rooijen, N., Giacca, M., Stassen, J. M., Dewerchin, M., Collen, D., & Carmeliet, P. (2007). Anti-PlGF inhibits growth of VEGF(R)-inhibitor-resistant tumors without affecting healthy vessels. Cell, 131(3), 463–475. https://doi.org/10.1016/j.cell.2007.08.038

    Article  CAS  PubMed  Google Scholar 

  6. Velazquez, O. C. (2007). Angiogenesis and vasculogenesis: Inducing the growth of new blood vessels and wound healing by stimulation of bone marrow-derived progenitor cell mobilization and homing. Journal of Vascular Surgery, 45 Suppl A(Suppl A), A39–A47. https://doi.org/10.1016/j.jvs.2007.02.068

    Article  PubMed  Google Scholar 

  7. Basile, D. P., & Yoder, M. C. (2014). Circulating and tissue resident endothelial progenitor cells. Journal of Cellular Physiology, 229(1), 10–16. https://doi.org/10.1002/jcp.24423

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Zeng, Y., Yao, X., Liu, X., He, X., Li, L., Liu, X., Yan, Z., Wu, J., & Fu, B. M. (2019). Anti-angiogenesis triggers exosomes release from endothelial cells to promote tumor vasculogenesis. Journal of Extracellular Vesicles, 8(1), 1629865. https://doi.org/10.1080/20013078.2019.1629865

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Madlambayan, G. J., Butler, J. M., Hosaka, K., Jorgensen, M., Fu, D., Guthrie, S. M., Shenoy, A. K., Brank, A., Russell, K. J., Otero, J., Siemann, D. W., Scott, E. W., & Cogle, C. R. (2009). Bone marrow stem and progenitor cell contribution to neovasculogenesis is dependent on model system with SDF-1 as a permissive trigger. Blood, 114(19), 4310–4319. https://doi.org/10.1182/blood-2009-03-211342

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Purhonen, S., Palm, J., Rossi, D., Kaskenpää, N., Rajantie, I., Ylä-Herttuala, S., Alitalo, K., Weissman, I. L., & Salven, P. (2008). Bone marrow-derived circulating endothelial precursors do not contribute to vascular endothelium and are not needed for tumor growth. Proceedings of the National Academy of Sciences of the United States of America, 105(18), 6620–6625. https://doi.org/10.1073/pnas.0710516105

    Article  PubMed  PubMed Central  Google Scholar 

  11. Chalasani, S. H., Sabelko, K. A., Sunshine, M. J., Littman, D. R., & Raper, J. A. (2003). A chemokine, SDF-1, reduces the effectiveness of multiple axonal repellents and is required for normal axon pathfinding. The Journal of neuroscience : the official journal of the Society for Neuroscience, 23(4), 1360–1371. https://doi.org/10.1523/JNEUROSCI.23-04-01360.2003

    Article  CAS  PubMed  Google Scholar 

  12. Rama, N., Dubrac, A., Mathivet, T., Ní Chárthaigh, R. A., Genet, G., Cristofaro, B., Pibouin-Fragner, L., Ma, L., Eichmann, A., & Chédotal, A. (2015). Slit2 signaling through Robo1 and Robo2 is required for retinal neovascularization. Nature Medicine, 21(5), 483–491. https://doi.org/10.1038/nm.3849

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Wang, B., Xiao, Y., Ding, B. B., Zhang, N., Yuan, X. B., Gui, L., Qian, K. X., Duan, S., Chen, Z., Rao, Y., & Geng, J. G. (2003). Induction of tumor angiogenesis by Slit-Robo signaling and inhibition of cancer growth by blocking Robo activity. Cancer Cell, 4(1), 19–29. https://doi.org/10.1016/s1535-6108(03)00164-8

    Article  PubMed  Google Scholar 

  14. Wang, L. J., Zhao, Y., Han, B., Ma, Y. G., Zhang, J., Yang, D. M., Mao, J. W., Tang, F. T., Li, W. D., Yang, Y., Wang, R., & Geng, J. G. (2008). Targeting Slit-roundabout signaling inhibits tumor angiogenesis in chemical-induced squamous cell carcinogenesis. Cancer Science, 99(3), 510–517. https://doi.org/10.1111/j.1349-7006.2007.00721.x

    Article  CAS  PubMed  Google Scholar 

  15. Butler, J. M., Guthrie, S. M., Koc, M., Afzal, A., Caballero, S., Brooks, H. L., Mames, R. N., Segal, M. S., Grant, M. B., & Scott, E. W. (2005). SDF-1 is both necessary and sufficient to promote proliferative retinopathy. The Journal of Clinical Investigation, 115(1), 86–93. https://doi.org/10.1172/JCI22869

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Pi, L., Xia, H., Liu, J., Shenoy, A. K., Hauswirth, W. W., & Scott, E. W. (2011). Role of connective tissue growth factor in the retinal vasculature during development and ischemia. Investigative Ophthalmology & Visual Science, 52(12), 8701–8710. https://doi.org/10.1167/iovs.11-7870

    Article  CAS  Google Scholar 

  17. Pi, L., Shenoy, A. K., Liu, J., Kim, S., Nelson, N., Xia, H., Hauswirth, W. W., Petersen, B. E., Schultz, G. S., & Scott, E. W. (2012). CCN2/CTGF regulates neovessel formation via targeting structurally conserved cystine knot motifs in multiple angiogenic regulators. FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology, 26(8), 3365–3379. https://doi.org/10.1096/fj.11-200154

    Article  CAS  PubMed  Google Scholar 

  18. Wu, J. Y., Feng, L., Park, H. T., Havlioglu, N., Wen, L., Tang, H., Bacon, K. B., Jiang, Z., Xc, Z., & Rao, Y. (2001). The neuronal repellent Slit inhibits leukocyte chemotaxis induced by chemotactic factors. Nature, 410(6831), 948–952. https://doi.org/10.1038/35073616

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Prasad, A., Fernandis, A. Z., Rao, Y., & Ganju, R. K. (2004). Slit protein-mediated inhibition of CXCR4-induced chemotactic and chemoinvasive signaling pathways in breast cancer cells. The Journal of Biological Chemistry, 279(10), 9115–9124. https://doi.org/10.1074/jbc.M308083200

    Article  CAS  PubMed  Google Scholar 

  20. Blockus, H., & Chédotal, A. (2016). Slit-Robo signaling. Development (Cambridge, England), 143(17), 3037–3044. https://doi.org/10.1242/dev.132829

    Article  CAS  PubMed  Google Scholar 

  21. Marlow, R., Strickland, P., Lee, J. S., Wu, X., Pebenito, M., Binnewies, M., Le, E. K., Moran, A., Macias, H., Cardiff, R. D., Sukumar, S., & Hinck, L. (2008). SLITs suppress tumor growth in vivo by silencing Sdf1/Cxcr4 within breast epithelium. Cancer Research, 68(19), 7819–7827. https://doi.org/10.1158/0008-5472.CAN-08-1357

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Legg, J. A., Herbert, J. M., Clissold, P., & Bicknell, R. (2008). Slits and roundabouts in cancer, tumour angiogenesis and endothelial cell migration. Angiogenesis, 11(1), 13–21. https://doi.org/10.1007/s10456-008-9100-x

    Article  PubMed  Google Scholar 

  23. Gara, R. K., Kumari, S., Ganju, A., Yallapu, M. M., Jaggi, M., & Chauhan, S. C. (2015). Slit/Robo pathway: A promising therapeutic target for cancer. Drug Discovery Today, 20(1), 156–164. https://doi.org/10.1016/j.drudis.2014.09.008

    Article  CAS  PubMed  Google Scholar 

  24. Dallol, A., Morton, D., Maher, E. R., & Latif, F. (2003). SLIT2 axon guidance molecule is frequently inactivated in colorectal cancer and suppresses growth of colorectal carcinoma cells. Cancer Research, 63(5), 1054–1058.

    CAS  PubMed  Google Scholar 

  25. Tavora, B., Mederer, T., Wessel, K. J., Ruffing, S., Sadjadi, M., Missmahl, M., Ostendorf, B. N., Liu, X., Kim, J. Y., Olsen, O., Welm, A. L., Goodarzi, H., & Tavazoie, S. F. (2020). Tumoural activation of TLR3-SLIT2 axis in endothelium drives metastasis. Nature, 586(7828), 299–304. https://doi.org/10.1038/s41586-020-2774-y

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Dallol, A., Dickinson, RE, Latif, F. (2005) Epigenetic disruption of the SLIT-ROBO interactions in human cancer. In M. Esteller (Ed.) Cancer Metastasis - Biology and Treatment. DNA Methylation, Epigenetics and Metastasis. Springer, p. 191–214.

  27. Latil, A., Chêne, L., Cochant-Priollet, B., Mangin, P., Fournier, G., Berthon, P., & Cussenot, O. (2003). Quantification of expression of netrins, slits and their receptors in human prostate tumors. International Journal of Cancer, 103(3), 306–315. https://doi.org/10.1002/ijc.10821

    Article  CAS  PubMed  Google Scholar 

  28. Narayan, G., Goparaju, C., Arias-Pulido, H., Kaufmann, A. M., Schneider, A., Dürst, M., Mansukhani, M., Pothuri, B., & Murty, V. V. (2006). Promoter hypermethylation-mediated inactivation of multiple Slit-Robo pathway genes in cervical cancer progression. Molecular Cancer, 5, 16. https://doi.org/10.1186/1476-4598-5-16

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Lyden, D., Hattori, K., Dias, S., Costa, C., Blaikie, P., Butros, L., Chadburn, A., Heissig, B., Marks, W., Witte, L., Wu, Y., Hicklin, D., Zhu, Z., Hackett, N. R., Crystal, R. G., Moore, M. A., Hajjar, K. A., Manova, K., Benezra, R., & Rafii, S. (2001). Impaired recruitment of bone-marrow-derived endothelial and hematopoietic precursor cells blocks tumor angiogenesis and growth. Nature Medicine, 7(11), 1194–1201. https://doi.org/10.1038/nm1101-1194

    Article  CAS  PubMed  Google Scholar 

  30. Bailey, A. S., Jiang, S., Afentoulis, M., Baumann, C. I., Schroeder, D. A., Olson, S. B., Wong, M. H., & Fleming, W. H. (2004). Transplanted adult hematopoietic stems cells differentiate into functional endothelial cells. Blood, 103(1), 13–19. https://doi.org/10.1182/blood-2003-05-1684

    Article  CAS  PubMed  Google Scholar 

  31. Méndez-Ferrer, S., Bonnet, D., Steensma, D. P., Hasserjian, R. P., Ghobrial, I. M., Gribben, J. G., Andreeff, M., & Krause, D. S. (2020). Bone marrow niches in haematological malignancies. Nature Reviews Cancer, 20(5), 285–298. https://doi.org/10.1038/s41568-020-0245-2

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Naito, H., Iba, T., & Takakura, N. (2020). Mechanisms of new blood-vessel formation and proliferative heterogeneity of endothelial cells. International Immunology, 32(5), 295–305. https://doi.org/10.1093/intimm/dxaa008

    Article  CAS  PubMed  Google Scholar 

  33. De Palma, M., & Naldini, L. (2006). Role of haematopoietic cells and endothelial progenitors in tumour angiogenesis. Biochimica et Biophysica Acta, 1766(1), 159–166. https://doi.org/10.1016/j.bbcan.2006.06.003

    Article  CAS  PubMed  Google Scholar 

  34. Murdoch, C., Muthana, M., Coffelt, S. B., & Lewis, C. E. (2008). The role of myeloid cells in the promotion of tumour angiogenesis. Nature Reviews Cancer, 8(8), 618–631. https://doi.org/10.1038/nrc2444

    Article  CAS  PubMed  Google Scholar 

  35. Ahn, G. O., & Brown, J. M. (2009). Role of endothelial progenitors and other bone marrow-derived cells in the development of the tumor vasculature. Angiogenesis, 12(2), 159–164. https://doi.org/10.1007/s10456-009-9135-7

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Lugano, R., Ramachandran, M., & Dimberg, A. (2020). Tumor angiogenesis: Causes, consequences, challenges and opportunities. Cellular and Molecular Life Sciences : CMLS, 77(9), 1745–1770. https://doi.org/10.1007/s00018-019-03351-7

    Article  CAS  PubMed  Google Scholar 

  37. Takakura, N., Watanabe, T., Suenobu, S., Yamada, Y., Noda, T., Ito, Y., Satake, M., & Suda, T. (2000). A role for hematopoietic stem cells in promoting angiogenesis. Cell, 102(2), 199–209. https://doi.org/10.1016/s0092-8674(00)00025-8

    Article  CAS  PubMed  Google Scholar 

  38. Fukumura, D., Kloepper, J., Amoozgar, Z., Duda, D. G., & Jain, R. K. (2018). Enhancing cancer immunotherapy using antiangiogenics: Opportunities and challenges. Nature Reviews Clinical Oncology, 15(5), 325–340. https://doi.org/10.1038/nrclinonc.2018.29

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Butler, J. M., Kobayashi, H., & Rafii, S. (2010). Instructive role of the vascular niche in promoting tumour growth and tissue repair by angiocrine factors. Nature Reviews Cancer, 10(2), 138–146. https://doi.org/10.1038/nrc2791

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Nolan, D. J., Ciarrocchi, A., Mellick, A. S., Jaggi, J. S., Bambino, K., Gupta, S., Heikamp, E., McDevitt, M. R., Scheinberg, D. A., Benezra, R., & Mittal, V. (2007). Bone marrow-derived endothelial progenitor cells are a major determinant of nascent tumor neovascularization. Genes & Development, 21(12), 1546–1558. https://doi.org/10.1101/gad.436307

    Article  CAS  Google Scholar 

  41. Srivastava, S., Pang, K. M., Iida, M., Nelson, M. S., Liu, J., Nam, A., Wang, J., Mambetsariev, I., Pillai, R., Mohanty, A., McDaniel, N., Behal, A., Kulkarni, P., Wheeler, D. L., & Salgia, R. (2020). Activation of EPHA2-ROBO1 heterodimer by SLIT2 attenuates non-canonical signaling and proliferation in squamous cell carcinomas. iScience, 23(11), 101692. https://doi.org/10.1016/j.isci.2020.101692

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Jin, J., You, H., Yu, B., Deng, Y., Tang, N., Yao, G., Shu, H., Yang, S., & Qin, W. (2009). Epigenetic inactivation of SLIT2 in human hepatocellular carcinomas. Biochemical and Biophysical Research Communications, 379(1), 86–91. https://doi.org/10.1016/j.bbrc.2008.12.022

    Article  CAS  PubMed  Google Scholar 

  43. Kim, H. K., Zhang, H., Li, H., Wu, T. T., Swisher, S., He, D., Wu, L., Xu, J., Elmets, C. A., Athar, M., Xu, X. C., & Xu, H. (2008). Slit2 inhibits growth and metastasis of fibrosarcoma and squamous cell carcinoma. Neoplasia (New York, N.Y.), 10(12), 1411–1420. https://doi.org/10.1593/neo.08804

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

The authors thank Neal Benson and the Flow Cytometry Core (courtesy of UFSCC) at the University of Florida, Marda Jorgenson (immunohistochemistry), and Douglas Smith (imaging) as part of the Cell and Tissue Analysis Core, at the University of Florida. The authors would also like to thank Dr. Mark Krebs for his help with confocal imaging.

Funding

This work was supported by grant numbers NIHR01 CA142808 and HL70738 from NIH (to EWS). The funding sources played no role in the study design, data collection, data management, data analysis, data interpretation, manuscript preparation, manuscript review, or manuscript approval.

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Authors and Affiliations

  1. Program in Stem Cell Biology and Regenerative Medicine, University of Florida College of Medicine, Gainesville, FL, USA

    Anitha K. Shenoy, Liya Pi, Alexander P. Ligocki, Koji Hosaka, Christopher R. Cogle & Edward W. Scott

  2. Department of Molecular Genetics and Microbiology, University of Florida, Gainesville, FL, USA

    Anitha K. Shenoy, Alexander P. Ligocki & Edward W. Scott

  3. Department of Pediatrics, University of Florida, Gainesville, FL, USA

    Liya Pi

  4. Department of Neurosurgery, University of Florida, Gainesville, FL, USA

    Koji Hosaka

  5. Department of Medicine, University of Florida, Gainesville, FL, USA

    Christopher R. Cogle

  6. Program in Stem Cell Biology and Regenerative Medicine, Department of Molecular Genetics and Microbology, University of Florida, PO Box 100232, Gainesville, FL, 32610, USA

    Edward W. Scott

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  1. Anitha K. Shenoy
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Shenoy, A.K., Pi, L., Ligocki, A.P. et al. Targeting Redundant ROBO1 and SDF-1 Pathways Prevents Adult Hemangioblast Derived-EPC and CEC Activity Effectively Blocking Tumor Neovascularization. Stem Cell Rev and Rep 19, 928–941 (2023). https://doi.org/10.1007/s12015-022-10498-7

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