Advertisement

Journal of Cancer Research and Clinical Oncology

, Volume 144, Issue 7, pp 1329–1337 | Cite as

Role of the NRP-1-mediated VEGFR2-independent pathway on radiation sensitivity of non-small cell lung cancer cells

  • Chenxi Hu
  • Panrong Zhu
  • Youyou Xia
  • Kaiyuan Hui
  • Mei Wang
  • Xiaodong Jiang
Original Article – Cancer Research

Abstract

Purpose

To determine if inhibiting neuropilin-1 (NRP-1) affects the radiosensitivity of NSCLC cells through a vascular endothelial growth factor receptor 2 (VEGFR2)-independent pathway, and to assess the underlying mechanisms.

Methods

The expression of VEGFR2, NRP-1, related signaling molecules, abelson murine leukemia viral oncogene homolog 1 (ABL-1), and RAD51 were determined by RT-PCR and Western blotting, respectively. Radiosensitivity was assessed using the colony-forming assay, and the cell apoptosis were analyzed by flow cytometry.

Results

We selected two cell lines with high expression levels of VEGFR2, including Calu-1 cells that have high NRP-1 expression, and H358 cells that have low NRP-1 expression. Upon inhibition of p-VEGFR2 by apatinib in Calu-1 cells, the expression of NRP-1 protein and other related proteins in the pathway was still high. Upon NRP-1 siRNA treatment, the expression of both NRP-1 and RAD51 decreased (p < 0.01; p < 0.05). Upon ABL-1 siRNA treatment, the expression of NRP-1 was increased and the expression of RAD51 was unchanged. Calu-1 cells treated with NRP-1 siRNA exhibited significantly higher apoptosis and radiation sensitivity in radiation therapy compared to Calu-1 cells treated with apatinib alone (p < 0.01; p < 0.01). The apoptosis and radiation sensitivity in H358 cells with NRP-1 overexpression was similar to the control group regardless of VEGFR2 inhibition.

Conclusions

We demonstrated that when VEGFR2 was inhibited, NRP-1 appeared to regulate RAD51 expression through the VEGFR2-independent ABL-1 pathway, consequently regulating radiation sensitivity. In addition, the combined inhibition of VEGFR2 and NRP-1 appears to sensitize cancer cells to radiation.

Keywords

NRP-1 VEGFR2 Non-small cell lung cancer Radiosensitivity 

Notes

Acknowledgements

This study was supported by the National Natural Science Foundation of China (no. 81472792), the Natural Science Foundation of Jiangsu Province, China (no. BK20151279), the Youth Medical Talent Project of Lianyungang (QNRC2016499), the Health Family Planning Technology Project of Lianyungang (QN1602), and the Youth Talents Found of Lianyungang First People’s Hospital (QN150101, QN140202).

Compliance with ethical standards

Conflict of interest

We have read and understood the Journal of Cancer Research and Clinical Oncology’s policy on disclosing conflicts of interest and declare that we have none.

Ethical statements

The use of the tissue microarray was approved by the Medical ethics committee.

References

  1. Alpay K, Farshchian M, Tuomela J, Sandholm J, Aittokallio K, Siljamäki E, Kallio M, Kähäri VM, Hietanen S (2014) Inhibition of c-Abl kinase activity renders cancer cells highly sensitive to mitoxantrone. PLoS One 9(8):e105526CrossRefPubMedPubMedCentralGoogle Scholar
  2. Brieger J, Schroeder P, Gosepath J, Mann WJ (2005) VEGF-subtype specific protection of SCC and HUVECs from radiation induced cell death. Int J Mol Med 15(1):145–151PubMedGoogle Scholar
  3. Cao S, Yaqoob U, Das A, Shergill U, Jagavelu K, Huebert RC, Routray C, Abdelmoneim S, Vasdev M, Leof E, Charlton M, Watts RJ, Mukhopadhyay D, Shah VH (2010) Neuropilin-1 promotes cirrhosis of the rodent and human liver by enhancing PDGF/TGF-beta signaling in hepatic stellate cells. J Clin Invest 120(7):2379–2394CrossRefPubMedPubMedCentralGoogle Scholar
  4. Chatterjee S, Heukamp LC, Siobal M, Schöttle J, Wieczorek C, Peifer M, Frasca D, Koker M, König K, Meder L, Rauh D, Buettner R, Wolf J, Brekken RA, Neumaier B, Christofori G, Thomas RK, Ullrich RT (2013) Tumor VEGF:VEGFR2 autocrine feed-forward loop triggers angiogenesis in lung cancer. J Clin Invest 123(4):1732–1740CrossRefPubMedPubMedCentralGoogle Scholar
  5. Ding MH, Liu L, Hu CX, Liu Y, Qiao Y, Jiang XD (2014) Expression of VEGFR2 and NRP-1 in non-small cell lung cancer and their clinical significance. Chin J Cancer Res 26(6):669–677PubMedPubMedCentralGoogle Scholar
  6. Dong JC, Gao H, Zuo SY, Zhang HQ, Zhao G, Sun SL, Han HL, Jin LL, Shao LH, Wei W, Jin SZ (2015) Neuropilin 1 expression correlates with the Radio-resistance of human non-small-cell lung cancer cells. J Cell Mol Med 19(9):2286–2295CrossRefPubMedPubMedCentralGoogle Scholar
  7. Ellis PM, Al-Saleh K (2012) Multitargeted anti-angiogenic agents and NSCLC: clinical update and future directions. Crit Rev Oncol Hematol 84(1):47–58CrossRefPubMedGoogle Scholar
  8. Evans IM, Yamaji M, Britton G, Pellet-Many C, Lockie C, Zachary IC, Frankel P (2011) Neuropilin-1 signaling through p130Cas tyrosine phosphorylation is essential for growth factor-dependent migration of glioma and endothelial cells. Mol Cell Biol 31(6):1174–1185CrossRefPubMedPubMedCentralGoogle Scholar
  9. Fantin A, Vieira JM, Plein A, Denti L, Fruttiger M, Pollard JW, Ruhrberg C (2013) NRP1 acts cell autonomously in endothelium to promote tip cell function during sprouting angiogenesis. Blood 121(12):2352–2362CrossRefPubMedPubMedCentralGoogle Scholar
  10. Gao X, Zhao Y, Stemmer-Rachamimov AO, Liu H, Huang P, Chin S, Selig MK, Plotkin SR, Jain RK, Xu L (2015) Anti-VEGF treatment improves neurological function and augments radiation response in NF2 schwannoma model. Proc Natl Acad Sci USA 112(47):14676–14681CrossRefPubMedPubMedCentralGoogle Scholar
  11. Grandclement C, Borg C (2011) Neuropilins: a new target for cancer therapy. Cancers (Basel) 3(2):1899–1928CrossRefGoogle Scholar
  12. Hainsworth JD, Waterhouse DM, Penley WC, Shipley DL, Thompson DS, Webb CD, Anthony Greco F (2013) Sorafenib and everolimus in advanced clear cell renal carcinoma: a phase I/II trial of the SCRI Oncology Research Consortium. Cancer Invest 31(5):323–329CrossRefPubMedGoogle Scholar
  13. Hamerlik P, Lathia JD, Rasmussen R, Wu Q, Bartkova J, Lee M, Moudry P, Bartek J Jr, Fischer W, Lukas J, Rich JN, Bartek J (2012) Autocrine VEGF-VEGFR2-Neuropilin-1 signaling promotes glioma stem-like cell viability and tumor growth. J Exp Med 209(3):507–520CrossRefPubMedPubMedCentralGoogle Scholar
  14. Jiang XD, Dai P, Wu J, Song DA, Yu JM (2012) Effect of recombinant human endostatin on radiosensitivity in patients with non-small-cell lung cancer. Int J Radiat Oncol Biol Phys 83(4):1272–1277CrossRefPubMedGoogle Scholar
  15. Jiang XD, Ding MH, Qiao Y, Liu Y, Liu L (2014a) Recombinant human endostatin combined with radiotherapy in the treatment of brain metastases of non-small cell lung cancer. Clin Transl Oncol 16(7):630–636CrossRefPubMedGoogle Scholar
  16. Jiang XD, Ding MH, Qiao Y, Liu Y, Liu L (2014b) Study on lung cancer cells expressing VEGFR2 and the impact on the effect of RHES combined with radiotherapy in the treatment of brain metastases. Clin Lung Cancer 15(2):e23-9CrossRefPubMedGoogle Scholar
  17. Ko JC, Ciou SC, Cheng CM, Wang LH, Hong JH, Jheng MY, Ling ST, Lin YW (2008) Involvement of Rad51 in cytotoxicity induced by epidermal growth factor receptor inhibitor (gefitinib, IressaR) and chemotherapeutic agents in human lung cancer cells. Carcinogenesis 29(7):1448–1458CrossRefPubMedGoogle Scholar
  18. Liu Y, Qiao Y, Hu C, Liu L, Zhou L, Liu B, Chen H, Jiang X (2016a) VEGFR2 inhibition by RNA interference affects cell proliferation, migration, invasion, and response to radiation in Calu-1 cells. Clin Transl Oncol 18(2):212–219CrossRefPubMedGoogle Scholar
  19. Liu L, Qiao Y, Hu C, Liu Y, Xia Y, Wang L, Liu B, Chen H, Jiang X (2016b) Endostatin exerts radiosensitizing effect in non-small cell lung cancer cells by inhibiting VEGFR2 expression. Clin Transl Oncol 18(1):18–26CrossRefPubMedGoogle Scholar
  20. Panigrahy D, Adini I, Mamluk R, Levonyak N, Bruns CJ, D’Amore PA, Klagsbrun M, Bielenberg DR (2014) Regulation of soluble neuropilin 1, an endogenous angiogenesis inhibitor, in liver development and regeneration. Pathology 46(5):416–423CrossRefPubMedPubMedCentralGoogle Scholar
  21. Raimondi C (2014) Neuropilin-1 enforces extracellular matrix signalling via ABL1 to promote angiogenesis. Biochem Soc Trans 42(5):1429–1434CrossRefPubMedGoogle Scholar
  22. Raimondi C, Fantin A, Lampropoulou A, Denti L, Chikh A, Ruhrberg C (2014) Imatinib inhibits VEGF-independent angiogenesis by targeting neuropilin 1-dependent ABL1 activation in endothelial cells. J Exp Med 211(6):1167–1183CrossRefPubMedPubMedCentralGoogle Scholar
  23. Raimondi C, Fantin A, Ruhrberg C (2015) Imatinib may be ABL to improve anti-angiogenic therapy. Mol Cell Oncol 2(1):e968034CrossRefPubMedPubMedCentralGoogle Scholar
  24. Satoda M, Takagi S, Ohta K, Hirata T, Fujisawa H (1995) Differential expression of two cell surface proteins, neuropilin and plexin, in Xenopus olfactory axon subclasses. J Neurosci 15(1 Pt 2):942–955CrossRefPubMedGoogle Scholar
  25. Subramanyam S, Ismail M, Bhattacharya I, Spies M (2016) Tyrosine phosphorylation stimulates activity of human RAD51 recombinase through altered nucleoprotein filament dynamics. Proc Natl Acad Sci USA 113(41):E6045–E6054CrossRefPubMedPubMedCentralGoogle Scholar
  26. Wang L, Dutta SK, Kojima T, Xu X, Khosravi-Far R, Ekker SC, Mukhopadhyay D (2007) Neuropilin-1 modulates p53/caspases axis to promote endothelial cell survival. PLoS One 2(11):e1161CrossRefPubMedPubMedCentralGoogle Scholar
  27. Xu Y, Li P, Zhang X, Wang J, Gu D, Wang Y (2013) Prognostic implication of neuropilin-1 upregulation in human nasopharyngeal carcinoma. Diagn Pathol 8:155PubMedPubMedCentralGoogle Scholar
  28. Yeo SG, Kim ES (2013) Efficient approach for determining four-dimensional computed tomography-based internal target volume in stereotactic radiotherapy of lung cancer. Radiat Oncol J 31(4):247–251CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Tumor Laboratory, Department of OncologyThe Affiliated Lianyungang Hospital of Xuzhou Medical UniversityLianyungangChina
  2. 2.Department of OncologyThe Affiliated Lianyungang Hospital of Xuzhou Medical UniversityLianyungangChina

Personalised recommendations