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

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.