ABSTRACT
Most of the anaplastic large-cell lymphoma (ALCL) cases carry the t(2;5; p23;q35) that produces the fusion protein NPM-ALK (nucleophosmin-anaplastic lymphoma kinase). 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 because of 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 relocalization of NPM-ALK to the cytoplasm by NPM genetic knockout or knockdown caused ERK1/2 (extracellular signal-regulated protein kinases 1 and 2) increased phosphorylation and cell death through the engagement of an ATM/Chk2- and γH2AX (phosphorylated H2A histone family member X)-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.
Subject(s)
Apoptosis , Lymphoma, Large-Cell, Anaplastic/metabolism , Oncogene Proteins, Fusion/metabolism , Protein-Tyrosine Kinases/metabolism , Signal Transduction , Animals , Blotting, Western , Cell Line, Tumor , Cell Survival/drug effects , Cell Survival/genetics , Cells, Cultured , Crizotinib , Dose-Response Relationship, Drug , Drug Synergism , Extracellular Signal-Regulated MAP Kinases/metabolism , Histones/metabolism , Humans , Hydrazines/pharmacology , Lymphoma, Large-Cell, Anaplastic/genetics , Lymphoma, Large-Cell, Anaplastic/pathology , Mice, Inbred NOD , Mice, Knockout , Mice, SCID , Microscopy, Confocal , Nucleophosmin , Oncogene Proteins, Fusion/genetics , Protein Kinase Inhibitors/pharmacology , Protein-Tyrosine Kinases/genetics , Pyrazoles/pharmacology , Pyridines/pharmacology , RNA Interference , Transplantation, Heterologous , Triazoles/pharmacologyABSTRACT
Non-small cell lung cancer (NSCLC) harboring chromosomal rearrangements of the anaplastic lymphoma kinase (ALK) gene is treated with ALK tyrosine kinase inhibitors (TKI), but the treatment is successful for only a limited amount of time; most patients experience a relapse due to the development of drug resistance. Here, we show that a vaccine against ALK induced a strong and specific immune response that both prophylactically and therapeutically impaired the growth of ALK-positive lung tumors in mouse models. The ALK vaccine was efficacious also in combination with ALK TKI treatment and significantly delayed tumor relapses after TKI suspension. We found that lung tumors containing ALK rearrangements induced an immunosuppressive microenvironment, regulating the expression of PD-L1 on the surface of lung tumor cells. High PD-L1 expression reduced ALK vaccine efficacy, which could be restored by administration of anti-PD-1 immunotherapy. Thus, combinations of ALK vaccine with TKIs and immune checkpoint blockade therapies might represent a powerful strategy for the treatment of ALK-driven NSCLC.
