The online version of this article (doi:10.1007/s13277-016-5433-z) contains supplementary material, which is available to authorized users.
Brain-derived neurotrophic factor (BDNF) and its tyrosine kinase receptor TrkB have been reported to be associated with poor prognosis in neuroblastoma (NB) patients. Our previous studies indicated that BDNF activation of TrkB induces chemo-resistance through activation of phosphoinositide-3-kinase (PI3K)/Akt pathway. In this study, we investigated the role of BDNF/TrkB on metastasis in NB. A tetracycline-regulated TrkB-expressing NB cell line (TB3) was used. Scratch wound healing assay, Boyden chamber migration, and invasion assays were performed to study the migration and invasion of TB3 cells. A tumor xenograft model using SCID-Beige mice was utilized to detect the metastasis of NB tumors in vivo. Inhibitors of PI3K, MAPK, Akt, and mTOR were used. Western blotting was performed to study the expressions of P-Akt, P-Erk, and P-mTOR. Our results showed that in TrkB-expressing NB cells, BDNF treatment significantly increased gap closing (P < 0.01) in scratch wound healing assay, also significantly enhanced the numbers of migrating cells (P < 0.01) and invading cells (P < 0.01) in the Boyden chamber migration and invasion assays. In vivo, NB distant metastases were significantly increased in mice with TrkB-expressing xenograft tumors compared to those with non-TrkB-expressing tumors (P < 0.05). Pre-treatment with any of the inhibitors for PI3K (LY294002), MAPK (PD98059), Akt (perifosine), or mTOR (rapamycin) blocked the BDNF/TrkB-induced increases of cell migration and invasion in TB3 cells, and also blocked the BDNF/TrkB-induced expressions of P-Akt, P-Erk, and P-mTOR. These data indicated that BDNF/TrkB increased metastasis in NB via PI3K/Akt/mTOR and MAPK pathways, and BDNF/TrkB and the downstream targets may be potential targets for the treatment of NB metastasis.
Suppl. Fig. 1 The effect of BDNF/TrkB on the expressions of N-cadherin, E-cadherin and Slug. TrkB-expressing TB3 cells were treated with BDNF (100 ng/ml) for 15 min, 1 h, 2 h, then harvested. Western blotting was performed to detect the expressions of N-cadherin, E-cadherin, and Slug (1:1000 dilution, Cell Signaling Tech.). GAPDH (1:10,000 dilution, Kangchen bio-tech) was used as the loading control. (PDF 80.7 kb)13277_2016_5433_MOESM1_ESM.pdf
Suppl. Fig. 2 The effect of PI3K, MAPK, Akt, and mTOR inhibitors on the BDNF/TrkB-induced changes of N-cadherin, E-cadherin, and Slug. TrkB-expressing TB3 cells were pre-treated with each of the inhibitors for 1 h (LY294002, 10 μM; PD98059, 10 μM; perifosine, 5 μM; rapamycin, 100 nM) followed by BDNF(100 ng/ml, 1 h) treatment. Cells were harvested and Western blotting was performed to detect the expressions of N-cadherin, E-cadherin, and Slug (1:1000 dilution, Cell Signaling Tech.), GAPDH (1:10,000 dilution, Kangchen bio-tech) was used as the loading control. (PDF 111 kb)13277_2016_5433_MOESM2_ESM.pdf
Jaboin J, Kim CJ, Kaplan DR, Thiele CJ. Brain-derived neurotrophic factor activation of TrkB protects neuroblastoma cells from chemotherapy-induced apoptosis via phosphatidylinositol 3'-kinase pathway. Cancer Res. 2002;62(22):6756–63. PubMed
Ho R, Eggert A, Hishiki T, Minturn JE, Ikegaki N, Foster P, et al. Resistance to chemotherapy mediated by TrkB in neuroblastomas. Cancer Res. 2002;62(22):6462–6. PubMed
Matsumoto K, Wada RK, Yamashiro JM, Kaplan DR, Thiele CJ. Expression of brain-derived neurotrophic factor and p145TrkB affects survival, differentiation, and invasiveness of human neuroblastoma cells. Cancer Res. 1995;55(8):1798–806. PubMed
Yang ZF, Ho DW, Lam CT, Luk JM, Lum CT, Yu WC, et al. Identification of brain-derived neurotrophic factor as a novel functional protein in hepatocellular carcinoma. Cancer Res. 2005;65(1):219–25. PubMed
Tanaka K, Okugawa Y, Toiyama Y, Inoue Y, Saigusa S, Kawamura M, et al. Brain-derived neurotrophic factor (BDNF)-induced tropomyosin-related kinase B (Trk B) signaling is a potential therapeutic target for peritoneal carcinomatosis arising from colorectal cancer. PLoS One. 2014;9(5):e96410. CrossRefPubMedPubMedCentral
- PI3K and MAPK pathways mediate the BDNF/TrkB-increased metastasis in neuroblastoma
Carol J. Thiele
- Springer Netherlands