Activating mutations in ALK provide a therapeutic target in neuroblastoma

Abstract

Neuroblastoma, an embryonal tumor of the peripheral sympathetic nervous system, accounts for approximately 15% of all deaths due to childhood cancer1. High-risk neuroblastomas, prevalent in the majority of patients, are rapidly progressive; even with intensive myeloablative chemotherapy, relapse is common and almost uniformly fatal2,3. Here we report the detection of previously unknown mutations in the ALK gene, which encodes a receptor tyrosine kinase, in 8% of primary neuroblastomas. Five non-synonymous sequence variations were identified in the kinase domain of ALK, of which three were somatic and two were germline. The most frequent mutation, F1174L, was also identified in three different neuroblastoma cell lines. ALK cDNAs encoding the F1174L and R1275Q variants, but not the wild-type ALK cDNA, transformed IL-3-dependent murine hematopoietic Ba/F3 cells to cytokine-independent growth. Ba/F3 cells expressing these mutations were sensitive to a small-molecule inhibitor of ALK, TAE6844. Furthermore, two human neuroblastoma cell lines harboring the F1174L mutation were sensitive to the inhibitor. Cytotoxicity was associated with increased levels of apoptosis as measured by TUNEL-labeling. shRNA-mediated knockdown of ALK expression in neuroblastoma cell lines with the F1174L mutation also resulted in apoptosis and impaired cell proliferation. Thus, activating alleles of the ALK receptor tyrosine kinase are present in primary neuroblastoma tumors and in established neuroblastoma cell lines, and confer sensitivity to ALK inhibition with small molecules, providing a molecular rationale for targeted therapy of this disease.

Aberrant activation of ALK kinase by a novel truncated form ALK protein in neuroblastoma

“…ALK inhibition by an ALK inhibitor and siRNA-mediated knockdown in neuroblastoma cells A partial ALK-deleted neuroblastoma-derived cell line (NB-1), ALK-mutated neuroblastoma-derived cell lines (SK-N-SH and TGW) and a glioblastomaderived cell line (H4) were cultured with varying concentrations of the ALK inhibitor TAE684, 8 and cell growth was measured using the CellTiter-Glo Luminescent Cell Viability Assay (Promega, Tokyo, Japan). NIH3T3 cells were used as a control.…”

Activity of tyrosine kinase inhibitor Dasatinib in neuroblastoma cells in vitro and in orthotopic mouse model

“…9,10 ALK inhibition resulted in profound inhibition of growth in all cell lines harbouring mutant or amplified ALK, pointing to this cell-surface kinase as a possible novel target for NB treatment. 11 The development of less toxic therapies is a compelling priority in paediatric oncology since the young age of patients and the likelihood of long-term side effects pose an additional problem of clinical management. At present, Stage 4 NB patients are treated with an aggressive multimodal treatment 12 that causes severe or life-threatening acute side effects, and it is only partially effective in achieving long-term survival (reviewed in Ref.…”

Exposure of neuroblastoma cell lines to imatinib results in the upregulation of the CDK inhibitor p27KIP1 as a consequence of c-Abl inhibition

“…IMR5, SK-N-AS, and SK-N-F1 cells have the wild-type and unamplified ALK gene [18,19]. IMR5 and SK-N-F1 are of N-type [20,21], SK-N-AS cells are of S-type [22].…”