binimetinib and Neuroblastoma

Novel therapies are needed for children with high-risk and relapsed neuroblastoma. We hypothesized that MAPK/ERK kinase (MEK) inhibition with the novel MEK1/2 inhibitor binimetinib would be effective in neuroblastoma preclinical models.. Levels of total and phosphorylated MEK and extracellular signal-regulated kinase (ERK) were examined in primary neuroblastoma tumor samples and in neuroblastoma cell lines by Western blot. A panel of established neuroblastoma tumor cell lines was treated with increasing concentrations of binimetinib, and their viability was determined using MTT assays. Western blot analyses were performed to examine changes in total and phosphorylated MEK and ERK and to measure apoptosis in neuroblastoma tumor cells after binimetinib treatment. NF1 protein levels in neuroblastoma cell lines were determined using Western blot assays. Gene expression of NF1 and MEK1 was examined in relationship to neuroblastoma patient outcomes.. Both primary neuroblastoma tumor samples and cell lines showed detectable levels of total and phosphorylated MEK and ERK. IC50 values for cells sensitive to binimetinib ranged from 8 nM to 1.16 μM, while resistant cells did not demonstrate any significant reduction in cell viability with doses exceeding 15 μM. Sensitive cells showed higher endogenous expression of phosphorylated MEK and ERK. Gene expression of NF1, but not MEK1, correlated with patient outcomes in neuroblastoma, and NF1 protein expression also correlated with responses to binimetinib.. Neuroblastoma tumor cells show a range of sensitivities to the novel MEK inhibitor binimetinib. In response to binimetinib, sensitive cells demonstrated complete loss of phosphorylated ERK, while resistant cells demonstrated either incomplete loss of ERK phosphorylation or minimal effects on MEK phosphorylation, suggesting alternative mechanisms of resistance. NF1 protein expression correlated with responses to binimetinib, supporting the use of NF1 as a biomarker to identify patients that may respond to MEK inhibition. MEK inhibition therefore represents a potential new therapeutic strategy for neuroblastoma.

Topics: Benzimidazoles; Cell Line, Tumor; Dose-Response Relationship, Drug; Extracellular Signal-Regulated MAP Kinases; Gene Expression Regulation, Neoplastic; Humans; Neuroblastoma; Neurofibromin 1; Protein Kinase Inhibitors; ras Proteins; Signal Transduction

The majority of patients with neuroblastoma have tumors that initially respond to chemotherapy, but a large proportion will experience therapy-resistant relapses. The molecular basis of this aggressive phenotype is unknown. Whole-genome sequencing of 23 paired diagnostic and relapse neuroblastomas showed clonal evolution from the diagnostic tumor, with a median of 29 somatic mutations unique to the relapse sample. Eighteen of the 23 relapse tumors (78%) showed mutations predicted to activate the RAS-MAPK pathway. Seven of these events were detected only in the relapse tumor, whereas the others showed clonal enrichment. In neuroblastoma cell lines, we also detected a high frequency of activating mutations in the RAS-MAPK pathway (11/18; 61%), and these lesions predicted sensitivity to MEK inhibition in vitro and in vivo. Our findings provide a rationale for genetic characterization of relapse neuroblastomas and show that RAS-MAPK pathway mutations may function as a biomarker for new therapeutic approaches to refractory disease.

Topics: Anaplastic Lymphoma Kinase; Animals; Benzimidazoles; Blotting, Western; Cell Line, Tumor; Child; Child, Preschool; Chromosome Aberrations; Cyclin-Dependent Kinase Inhibitor p16; Female; Gene Expression Regulation, Neoplastic; HEK293 Cells; Humans; Infant; Male; MAP Kinase Signaling System; Mice, SCID; Mitogen-Activated Protein Kinases; Mutation; Neoplasm Recurrence, Local; Neuroblastoma; Phosphorylation; ras Proteins; Receptor Protein-Tyrosine Kinases; Reverse Transcriptase Polymerase Chain Reaction; Xenograft Model Antitumor Assays