Neurotrophin-3 production promotes human neuroblastoma cell survival by inhibiting TrkC-induced apoptosis
Jimena Bouzas-Rodriguez, … , Servane Tauszig-Delamasure, Patrick Mehlen
Jimena Bouzas-Rodriguez, … , Servane Tauszig-Delamasure, Patrick Mehlen
Published February 15, 2010
Citation Information: J Clin Invest. 2010;120(3):850-858. https://doi.org/10.1172/JCI41013.
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Research Article Oncology

Neurotrophin-3 production promotes human neuroblastoma cell survival by inhibiting TrkC-induced apoptosis

Abstract

Tropomyosin-related kinase receptor C (TrkC) is a neurotrophin receptor with tyrosine kinase activity that was expected to be oncogenic. However, it has several characteristics of a tumor suppressor: its expression in tumors has often been associated with good prognosis; and it was recently demonstrated to be a dependence receptor, transducing different positive signals in the presence of ligand but inducing apoptosis in the absence of ligand. Here we show that the TrkC ligand neurotrophin-3 (NT-3) is upregulated in a large fraction of aggressive human neuroblastomas (NBs) and that it blocks TrkC-induced apoptosis of human NB cell lines, consistent with the idea that TrkC is a dependence receptor. Functionally, both siRNA knockdown of NT-3 expression and incubation with a TrkC-specific blocking antibody triggered apoptosis in human NB cell lines. Importantly, disruption of the NT-3 autocrine loop in malignant human neuroblasts triggered in vitro NB cell death and inhibited tumor growth and metastasis in both a chick and a mouse xenograft model. Thus, we believe that our data suggest that NT-3/TrkC disruption is a putative alternative targeted therapeutic strategy for the treatment of NB.

Authors

Jimena Bouzas-Rodriguez, Jorge Ruben Cabrera, Céline Delloye-Bourgeois, Gabriel Ichim, Jean-Guy Delcros, Marie-Anne Raquin, Raphaël Rousseau, Valérie Combaret, Jean Bénard, Servane Tauszig-Delamasure, Patrick Mehlen

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Figure 3

NT-3/TrkC interference promotes TrkC proapoptotic activity.

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NT-3/TrkC interference promotes TrkC proapoptotic activity. (A) CLB-Ge2 ...
(A) CLB-Ge2 cells were transfected with either empty vector or with a plasmid encoding the dominant-negative TrkC-IC D641N and treated for 24 hours with or without anti-TrkC blocking antibody Cell death was monitored by TUNEL labeling of cells plated on slides. The control panel shows TrkC-IC D641N by Western blots using anti-TrkC antibody (bottom panel). Representative images are shown. Original magnification, ×20. (B) The efficacy of TrkC siRNA was evaluated by Western blot on nonexpressing TrkC 13.S.24 olfactive neuroblasts. Cells were transfected either with empty vector or with uncleavable TrkC D945N D641N double mutant that does not trigger apoptosis, and with scrambled siRNA or TrkC siRNA (siRNA TrkC). (C) Cell death induction in the CLB-Ge2 cell line was quantified after transfection with either scrambled siRNA (siScr), TrkC siRNA (siTrkC), NT-3 siRNA (siNT-3), or a mix of TrkC and NT-3 siRNA, using relative caspase-3 activity assay. (D) Phospho-Akt and phospho-Erk levels of CLB-Ge2 cells were monitored by Western blot after 16 hours of treatment with 2 μg/ml anti-TrkC blocking antibody 20 nM Ly29402, 100 nM U0126, or 100 ng/ml NT-3, in absence of serum. (E) Detection of TrkC cleavage band (20 kDa, indicated by the arrow) by Western blot, using an anti-TrkC antibody on cells treated (or not) with anti-TrkC blocking antibody, with or without the general caspase inhibitor BAF. (A and C) Data represent mean ± SEM. *P < 0.05, 2-sided Mann-Whitney test, compared with control.