Phosphoproteomics reveals ALK promote cell progress via RAS/ JNK pathway in neuroblastoma

Oncotarget. 2016 Nov 15;7(46):75968-75980. doi: 10.18632/oncotarget.12513.

Abstract

Emerging evidence suggests receptor tyrosine kinase ALK as a promising therapeutic target in neuroblastoma. However, clinical trials reveal that a limited proportion of ALK-positive neuroblastoma patients experience clinical benefits from Crizotinib, a clinically approved specific inhibitor of ALK. The precise molecular mechanisms of aberrant ALK activity in neuroblastoma remain elusive, limiting the clinical application of ALK as a therapeutic target in neuroblastoma. Here, we describe a deep quantitative phosphoproteomic approach in which Crizotinib-treated neuroblastoma cell lines bearing aberrant ALK are used to investigate downstream regulated phosphoproteins. We identified more than 19,500-and quantitatively analyzed approximately 10,000-phosphorylation sites from each cell line, ultimately detecting 450-790 significantly-regulated phosphorylation sites. Multiple layers of bioinformatic analysis of the significantly-regulated phosphoproteins identified RAS/JNK as a downstream signaling pathway of ALK, independent of the ALK variant present. Further experiments demonstrated that ALK/JNK signaling could be inactivated by either ALK- or JNK-specific inhibitors, resulting in cell growth inhibition by induction of cell cycle arrest and cell apoptosis. Our study broadly defines the phosphoproteome in response to ALK inhibition and provides a resource for further clinical investigation of ALK as therapeutic target for the treatment of neuroblastoma.

Keywords: ALK; neuroblastoma; pediatric oncology; phosphoproteomics; target therapy.

MeSH terms

  • Anaplastic Lymphoma Kinase
  • Cell Cycle Checkpoints / drug effects
  • Cell Line, Tumor
  • Cell Survival / drug effects
  • Computational Biology / methods
  • Disease Progression
  • Humans
  • JNK Mitogen-Activated Protein Kinases / metabolism*
  • Neuroblastoma / metabolism*
  • Neuroblastoma / pathology
  • Phosphoproteins / metabolism*
  • Protein Kinase Inhibitors
  • Proteome
  • Proteomics* / methods
  • Receptor Protein-Tyrosine Kinases / metabolism*
  • Signal Transduction*
  • ras Proteins / metabolism*

Substances

  • Phosphoproteins
  • Protein Kinase Inhibitors
  • Proteome
  • ALK protein, human
  • Anaplastic Lymphoma Kinase
  • Receptor Protein-Tyrosine Kinases
  • JNK Mitogen-Activated Protein Kinases
  • ras Proteins