Expression of the oncogenic NPM-ALK chimeric protein in human lymphoid T-cells inhibits drug-induced, but not Fas-induced apoptosis

Oncogene. 2001 Nov 1;20(50):7386-97. doi: 10.1038/sj.onc.1204870.

Abstract

Anaplastic large cell lymphomas (ALCLs) are frequently associated with the t(2;5)(p23;q35) translocation, leading to the expression of NPM-ALK, a fusion protein linking nucleophosmin and anaplastic lymphoma kinase, a receptor tyrosine kinase. In ALCLs, dimerization of NPM-ALK leads to constitutive autophosphorylation and activation of the kinase, necessary for NPM-ALK oncogenicity. To investigate whether NPM-ALK, like other oncogenic tyrosine kinases, can inhibit drug-induced apoptosis, we permanently transfected NPM-ALK into Jurkat T-cells. As in ALCLs, NPM-ALK was expressed as a constitutively kinase-active 80 kDa protein, and could be detected by immunocytochemistry in nucleoli, nuclei and cytoplasm. Doxorubicin-induced apoptosis (assessed by cell morphology and annexin V-FITC binding) was significantly inhibited in two independent NPM-ALK-expressing clones (5.2+/-1.8 and 7.5+/-0.8% apoptosis), compared to control vector-transduced cells (36+/-6.7%). Similar results were observed with etoposide. In contrast, Fas-induced apoptosis was not inhibited. Cytochrome c release into the cytosol was delayed in doxorubicin-, but not anti-Fas-treated transfectant cells, indicating that apoptosis inhibition occurred upstream of mitochondrial events. Using NPM-ALK mutants, we demonstrated that inhibition of drug-induced apoptosis: (1) requires functional kinase activity, (2) does not involve phospholipase C-gamma, essential for NPM-ALK-mediated mitogenicity and (3) appears to be phosphoinositide 3-kinase independent, despite a strong Akt/PKB activation observed in wild type NPM-ALK-expressing cells. These results suggest that the NPM-ALK antiapoptotic and mitogenic pathways are distinct.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Antineoplastic Agents / pharmacology
  • Apoptosis / drug effects*
  • Apoptosis / genetics
  • Apoptosis / physiology
  • Binding Sites
  • Chromones / pharmacology
  • Cytochrome c Group / metabolism
  • Doxorubicin / pharmacology
  • Enzyme Inhibitors / pharmacology
  • Etoposide / pharmacology
  • Fas Ligand Protein
  • Humans
  • Isoenzymes / metabolism
  • Jurkat Cells / drug effects
  • Jurkat Cells / metabolism
  • Membrane Glycoproteins / physiology*
  • Mitochondria / drug effects
  • Mitochondria / enzymology
  • Morpholines / pharmacology
  • Mutagenesis, Site-Directed
  • Neoplasm Proteins / biosynthesis
  • Neoplasm Proteins / genetics
  • Neoplasm Proteins / physiology
  • Phosphatidylinositol 3-Kinases / metabolism
  • Phosphoinositide-3 Kinase Inhibitors
  • Phospholipase C gamma
  • Phosphorylation
  • Protein Processing, Post-Translational
  • Protein-Serine-Threonine Kinases*
  • Protein-Tyrosine Kinases / biosynthesis
  • Protein-Tyrosine Kinases / chemistry
  • Protein-Tyrosine Kinases / genetics
  • Protein-Tyrosine Kinases / physiology*
  • Proto-Oncogene Proteins / metabolism
  • Proto-Oncogene Proteins c-akt
  • Recombinant Fusion Proteins / physiology
  • T-Lymphocytes / drug effects
  • T-Lymphocytes / metabolism*
  • Transfection
  • Type C Phospholipases / metabolism
  • fas Receptor / physiology*

Substances

  • Antineoplastic Agents
  • Chromones
  • Cytochrome c Group
  • Enzyme Inhibitors
  • FASLG protein, human
  • Fas Ligand Protein
  • Isoenzymes
  • Membrane Glycoproteins
  • Morpholines
  • Neoplasm Proteins
  • Phosphoinositide-3 Kinase Inhibitors
  • Proto-Oncogene Proteins
  • Recombinant Fusion Proteins
  • fas Receptor
  • 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
  • Etoposide
  • Doxorubicin
  • Adenosine Triphosphate
  • p80(NPM-ALK) protein
  • Protein-Tyrosine Kinases
  • AKT1 protein, human
  • Protein-Serine-Threonine Kinases
  • Proto-Oncogene Proteins c-akt
  • Type C Phospholipases
  • Phospholipase C gamma