Oncogenic tyrosine kinase NPM/ALK induces activation of the MEK/ERK signaling pathway independently of c-Raf

Abstract

The mechanisms of cell transformation mediated by the highly oncogenic, chimeric NPM/ALK tyrosine kinase remain only partially understood. Here we report that cell lines and native tissues derived from the NPM/ALKexpressing T-cell lymphoma (ALK þ TCL) display phosphorylation of the extracellular signal-regulated protein kinase (ERK) 1/2 complex. Transfection of BaF3 cells with NPM/ALK induces phosphorylation of EKR1/2 and of its direct activator mitogen-induced extracellular kinase (MEK) 1/2. Depletion of NPM/ ALK by small interfering RNA (siRNA) or its inhibition by WHI-154 abrogates the MEK1/2 and ERK1/2 phosphorylation. The NPM/ALK-induced MEK/ERK activation is independent of c-Raf as evidenced by the lack of MEK1/2 and ERK1/2 phosphorylation upon c-Raf inactivation by two different inhibitors, RI and ZM336372, and by its siRNA-mediated depletion. In contrast, ERK1/2 activation is strictly MEK1/2 dependent as shown by suppression of the ERK1/2 phosphorylation by the MEK1/2 inhibitor U0126. The U0126-mediated inhibition of ERK1/2 activation impaired proliferation and viability of the ALK þ TCL cells and expression of antiapoptotic factor Bcl-xL and cell cyclepromoting CDK4 and phospho-RB. Finally, siRNAmediated depletion of both ERK1 and ERK2 inhibited cell proliferation, whereas depletion of ERK 1 (but not ERK2) markedly increased cell apoptosis. These findings identify MEK/ERK as a new signaling pathway activated by NPM/ALK and indicate that the pathway represents a novel therapeutic target in the ALK-induced malignancies.

Oncogenic tyrosine kinase NPM/ALK induces activation of the rapamycin-sensitive mTOR signaling pathway

“…However, relatively few NPM/ ALK-triggered pathways have been identified so far, with the roles of PI3K/Akt, STAT3, STAT5 and MEK/ ERK being the best characterized (Wasik, 2002;Chiarle et al, 2005;Kasprzycka et al, 2006;Marzec et al, 2007). In this report, we provide several lines of evidence that NPM/ALK induces also activation of mTOR by the mechanisms schematically depicted in Figure 6.…”

“…As the MEK/ERK signaling pathway is induced by NPM/ALK (Marzec et al, 2007) and can activate mTOR (Tee et al, 2003;Roux et al, 2004;Ma et al, 2005;Arvisais et al, 2006), we examined whether inhibition of MEK would impact mTOR activation in ALK þ TCL cells. Treatment of Sudhl-1 cells with two different MEK inhibitors, U0126 and PD98059, markedly suppressed phosphorylation of not only ERK1/2 but also the mTOR target S6rp (Figure 4a).…”

“…Considering that mTOR inhibitors rapamycin and RAD001 are used as immunosuppressive drugs known to affect primarily T lymphocytes, ALK þ TCL cells may be quite sensitive to therapies targeting mTOR. Although it could be argued that NPM/ALK itself represents an attractive therapeutic target (Marzec et al, 2007), combination of an ALK inhibitor with inhibitor(s) of its key effectors such as mTOR may be even more effective. Finally, given the strong antiproliferative and proapoptotic effect of rapamycin on ALK þ TCL cells (Figure 5), targeting mTOR alone may have a significant therapeutic effect in ALK þ TCL patients.…”

“…The t(2;5)(p23;q35) translocation that occurs in >80% of ALK þ TCL fuses a distal portion of the ALK gene with the proximal part of the nucleophosmin (NPM) gene (Morris et al, 1994;Shiota et al, 1994). The chimeric NPM/ALK protein is constitutively active due to autophosphorylation and promotes malignant cell transformation as shown both in vitro (Fujimoto et al, 1996;Bischof et al, 1997) and in vivo (Kuefer et al, 1997) by phosphorylating and activating signaling transmitters and pathways, such as phospholipase C (PLC)gphosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt), Stat3, Stat5 and mitogen-induced extracellular kinase (MEK)/extracellular signal-regulated kinase (ERK) (Wasik, 2002;Marzec et al, 2007).…”

Activation of Rac1 and the exchange factor Vav3 are involved in NPM-ALK signaling in anaplastic large cell lymphomas

“…Pro-mitogenic functions include binding of adaptors, such as Shc, Grb2 and IRS1, to regulators of the Erk pathway, phospholipase Cg (PLCg), tyrosine phosphatases and the protooncogene pp60 c-src (Fujimoto et al, 1996;Bai et al, 1998;Cussac et al, 2004;Honorat et al, 2006;Marzec et al, 2007). Antiapoptotic functions are related to the activation of the survival phosphatidylinositol 3-kinase (PI3K)/AKT pathway and of the Jak/STAT3-5 module (Bai et al, 2000;Amin et al, 2003;Chiarle et al, 2005).…”