Excess of NPM-ALK oncogenic signaling promotes cellular apoptosis and drug dependency

Abstract

Most of Anaplastic Large Cell Lymphoma (ALCL) cases carry the t(2;5; p23;q35) that produces the fusion protein NPM-ALK. NPM-ALK deregulated kinase activity drives several pathways that support malignant transformation of lymphoma cells. We found that in ALK-rearranged ALCL cell lines NPM-ALK was distributed in equal amounts between the cytoplasm and the nucleus. Only the cytoplasmic portion was catalytically active in both cell lines and primary ALCL, whereas the nuclear portion was inactive due to heterodimerization with NPM1. Thus, about 50% of the NPM-ALK is not active and sequestered as NPM-ALK/NPM1 heterodimers in the nucleus. Overexpression or re-localization of NPM-ALK to the cytoplasm by NPM genetic knock-out or knock-down caused ERK1/2 increased phosphorylation and cell death through the engagement of an ATM/Chk2 and γH2AX mediated DNA damage response. Remarkably, human NPM-ALK amplified cell lines resistant to ALK tyrosine kinase inhibitors (TKIs) underwent apoptosis upon drug withdrawal as a consequence of ERK1/2 hyperactivation. Altogether, these findings indicate that an excess of NPM-ALK activation and signaling induces apoptosis via oncogenic stress responses. A “drug holiday” where the ALK TKI treatment is suspended could represent a therapeutic option in cells that become resistant by NPM-ALK amplification.

Lorlatinib Treatment Elicits Multiple On- and Off-Target Mechanisms of Resistance in ALK-Driven Cancer

“…Therefore, these cells were kept in culture in the presence of 3-10 nmol/L lorlatinib. However, in contrast to previously described drug-addicted cells (26), NPM/ALK transcript levels were not substantially upregulated (Table 1; Supplementary Fig. S2).…”

“…Whether this is due to inefficient dimerization with normal NPM1, or preferential binding to cytoplasmic proteins, is unclear at present and is under investigation. We speculate that the aberrant cytoplasmic localization of N1178H mutants functionally mimics an overexpressed fusion kinase, since the cytoplasmic fraction of NPM/ALK is thought to be the oncogenic driver (26), thus leading to resistance. This behavior may also explain the curious difference in Tyr1604 versus Tyr1278 uniquely observed in N1178H-mutated BD1 cells.…”

“…The new K299-LR1000 and SUPM2-LR1000 cell lines displayed a drug-addicted phenotype ( Fig. 1E and F), that is, their viability progressively decreased at lower lorlatinib doses (26). They showed persistent ALK phosphorylation at high drug concentrations ( Fig.…”

Non-translational Connections of eEF1B in the Cytoplasm and Nucleus of Cancer Cells

“…An EEF1G-anaplastic lymphoma kinase (ALK) gene fusion was recently found in two pediatric patients with anaplastic large cell lymphoma (Palacios et al, 2017). In the context of the review, it is important that a fusion of the eEF1Bγ N-terminal domain provided exclusively cytoplasmic localization of the fusion protein (Ceccon et al, 2016) suggesting that the N-terminal domain of eEF1Bγ could not possibly be related to nuclear localization of eEF1Bγ alone.…”

TRAF1-ALK fusion indicates poor prognosis to ALK tyrosine kinase inhibitors in relapsed/refractory ALK-positive anaplastic large-cell lymphoma patients

“…We also observed that patients who harbored the NPM1-ALK fusion would be likely to achieve CR, especially when the fusion-type was exon 4 of the NPM1 gene aligned to exon 20 of the ALK gene. based on a preclinical model, that temporary suspension of TKI administration might bene t patients with the NPM-ALK fusion who develop resistance, which is the so-called "drug holiday" therapy 27 . There is no clinical evidence supporting the effectiveness of such therapy.…”