TrkB/BDNF signaling promotes EMT mediated invasiveness and is a potential therapeutic target for gallbladder cancer

Abstract

Background: Tropomyosin-related kinase B (TrkB)/ brain derived neurotrophic factor (BDNF) signaling has been shown to be activated and be involved with inducing malignant phenotype in several cancers. However, the contribution of its signaling to one of refractory malignancies, gallbladder cancer (GBC), still remains unclear. This study assessed the biological function of TrkB/BDNF signaling in GBC and investigated whether it could be a new therapeutic target in GBC. Methods: 1) Clinical experiment: 69 patients with GBC who underwent curative surgical resection were enrolled in this study. Correlation between TrkB expression and clinicopathological findings was analyzed immunohistochemically. 2) In vitro experiment: TrkB/BDNF signaling was inhibited using k252a, siRNA and shRNA, and was activated by recombinant BDNF (rBDNF). Then, whether TrkB/BDNF signaling contributes to the invasiveness was estimated by Matrigel invasion assay using TrkB expressing GBC cell lines (TGBC, GBd15, NOZ, TYGBK - 1, TYGBK - 2). Epithelial Mesenchymal Transition (EMT) related protein levels were analyzed by western blotting. 3) In vivo experiment: In Xenograft mice model, tumorigenesis and invasiveness of TrkB shRNA transfected cells (NOZ, TYGBK-1) was analyzed. Results: 1) Clinical results; TrkB expression was detected in 63 (91.3%) GBC specimens. TrkB expression detected in invasive front significantly correlated with T factor (p = 0.0391) and clinical staging (p = 0.0391). Overall survival in patients with high TrkB expression was significantly lower than those with low TrkB expression (p = 0.0363). 2) In vitro results; Addition of rBDNF significantly increased invasiveness and K252a treatment significantly decreased invasiveness of GBC cells. TrkB and BDNF siRNA transfection significantly inhibited the invasiveness of GBC cells. SiRNA transfection increased the expression of E-cadherin and decreased the expressions of vimentin, slug, snail, and twist. 3) In vivo results; Tumors from mice injected with TrkB shRNA transfected cells significantly reduced the tumorigenicity and invasiveness. Conclusions: TrkB/BDNF signaling enhances invasiveness of GBC, and it could be a potential therapeutic target.