The Role of p75NTR and its signaling pathways in Fenretinide (4-hydroxyphenyl retinamide)-induced apoptosis in neuroblastoma cells

OBJECTIVE: Neuroblastoma is a common, frequently fatal, neural crest tumor of childhood. Chemotherapy-resistant neuroblastoma cells typically have Schwann cell-like ("S-type") morphology and express the p75 neurotrophin receptor (p75NTR) and are often refractory to chemotherapy. p75NTR has been previously shown to modulate the redox state of neural crest tumor cells. The retinoic acid analogue, fenretinide(4HPR), has been shown to cause apoptosis by increasing generation of reactive oxygen species in cancer cells. We tested the hypothesis that expression of the p75 neurotrophin receptor (p75NTR), enhances neuroblastoma responsiveness to 4HPR.;METHODS: Neuroblastoma cell lines derived from minimal residual disease were used to determine the effect of p75NTR signaling on susceptibility to 4HPR-induced apoptosis. p75NTR signaling was knocked down using transfection and concentration- and time-cell survival studies were performed to determine sensitivity to 4HPR. Changes in oxidative stress as a result of 4HPR treatment were determined using redox-active dye staining and site-specific antioxidants. Western blotting and site-specific stains were used to determine the components of the signaling pathway involved in 4HPR-induced apoptosis.;RESULTS: Knockdown of p75NTR attenuated 4HPR-induced accumulation of mitochondrial superoxide and apoptosis. Overexpression of p75NTR had the opposite effects. Treatment of SH-EP1 neuroblastoma cells with 4HPR for 72 hrs revealed no change in production of cellular hydroxyl radical or cytosolic superoxide, but a sharp increase was observed in mitochondrial superoxide production. Pretreatment of cells with 2-thenoyltrifluoroacetone or dehydroascorbic acid uniquely prevented mitochondrial superoxide accumulation and cell death after 4HPR treatment. Mitochondrial complex II is the likely site of 4HPR-induced superoxide generation and p75NTR-induced potentiation of these phenomena. MAPK signaling resulting in JNK phosphorylation is one of the central pathways regulating apoptosis following 4HPR treatment. Akt signaling is a protective signal which is enhanced in p75NTR knockdown neuroblastoma cells.;CONCLUSION: Modification of expression of p75NTR in a particular neuroblastoma cell line modifies its susceptibility to 4HPR. Enhancers of p75NTR expression or signaling could be potential drugs for use as adjuncts to chemotherapy of neural tumors. Mitochondrial oxidative stress, JNK phosphorylation and suppression of Akt signaling are responsible for this potentiation. p75NTR is a biomarker for efficacy of 4HPR against neuroblastoma.