Sensitivity of neuroblastoma to the novel kinase inhibitor cabozantinib is mediated by ERK inhibition

Zhang, Linna; Scorsone, Kathleen A.; Woodfield, Sarah E.; Zage, Peter E.

doi:10.1007/s00280-015-2871-z

Cited by 26 publications

(34 citation statements)

References 42 publications

(57 reference statements)

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“…both of our luciferase transfected cell lines are sensitive to proliferation and migration inhibiting effects of cabozantinib, an inhibitor of VEGFR2 and MET kinases, with IC 50 s in the low micromolar range. Our findings were consistent with the recent results described by Zhang et al (37) where IMR-32 was one of the most sensitive neuroblastoma cell lines tested. In addition, we demonstrated that wound repair capacity was significantly reduced in IGR-N91-Luc cells and more importantly in the IMR-32-Luc cell line.…”

Section: Discussionsupporting

confidence: 94%

Dual inhibition using cabozantinib overcomes HGF/MET signaling mediated resistance to pan-VEGFR inhibition in orthotopic and metastatic neuroblastoma tumors

Daudigeos-Dubus

Dret

Bawa

et al. 2016

International Journal of Oncology

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MET is expressed on neuroblastoma cells and may trigger tumor growth, neoangiogenesis and metastasis. MET upregulation further represents an escape mechanism to various anticancer treatments including VEGF signaling inhibitors. We developed in vitro a resistance model to pan-VEGFR inhibition and explored the simultaneous inhibition of VEGFR and MET in neuroblastoma models in vitro and in vivo using cabozantinib, an inhibitor of the tyrosine kinases including VEGFR2, MET, AXL and RET. Resistance in IGR-N91-Luc neuroblastoma cells under continuous in vitro exposure pressure to VEGFR1-3 inhibition using axitinib was associated with HGF and p-ERK overexpression. Cabozantinib exhibited anti-proliferative effects in neuroblastoma cells and reduced cell migration in vitro as measured by phase-contrast with IncuCyte system. In vivo, an enhanced number of animals with IGR-N91-Luc metastases was noted following axitinib treatment as compared to control animals. Orally administered cabozantinib per gavage at 30 and 60 mg/kg/day significantly inhibited tumor growth of orthotopic adrenal IGR-N91-Luc and metastatic IMR-32-Luc xenografts. Antitumor activity was associated with decreased vascularization, inhibition of p-SRC and induction of apoptotic cell death. Activation of the HGF-mediated MET pathway is involved in escape to selective VEGFR inhibition in neuroblastoma suggesting combined inhibition of MET and VEGFR signaling to reduce secondary resistance and enhanced invasiveness.

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Section: Discussionsupporting

confidence: 94%

Dual inhibition using cabozantinib overcomes HGF/MET signaling mediated resistance to pan-VEGFR inhibition in orthotopic and metastatic neuroblastoma tumors

Daudigeos-Dubus

Dret

Bawa

et al. 2016

International Journal of Oncology

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“…The neuroblastoma cell lines used in this study have been previously described [ 30 – 38 ] and were generously provided by Shahab Asgharzadeh (Children’s Hospital Los Angeles, Los Angeles, CA), Susan Cohn (The University of Chicago Children’s Hospital, Chicago, IL), Jill Lahti (St. Jude Children’s Research Hospital, Memphis, TN), John Maris (Children’s Hospital of Philadelphia, Philadelphia, PA), William Weiss (The University of California, San Francisco, San Francisco, CA) or were purchased from the American Type Culture Collection (ATCC; Rockville, MD). Cell lines were grown at 37° in 5 % CO 2 in appropriate media (Invitrogen, Carlsbad, CA) supplemented with 10 % heat-inactivated fetal bovine serum (FBS) (Life Technologies, Grand Island, NY), L-glutamine, sodium pyruvate, and non-essential amino acids [ 39 ]. All cell lines were authenticated by deoxyribonucleic acid (DNA) profiling prior to use.…”

Section: Methodsmentioning

confidence: 99%

“…Patient tumor samples were homogenized and incubated for 30 min in radioimmunoprecipitation assay (RIPA) protein lysis buffer containing protease inhibitors (Sigma) and phosphatase inhibitors (Roche, San Francisco, CA) with homogenization every 10 min as previously described [ 39 ]. Lysates were centrifuged and supernatants were collected.…”

Section: Methodsmentioning

confidence: 99%

Binimetinib inhibits MEK and is effective against neuroblastoma tumor cells with low NF1 expression

et al. 2016

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BackgroundNovel 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.MethodsLevels 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.ResultsBoth 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.ConclusionsNeuroblastoma 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-016-2199-z) contains supplementary material, which is available to authorized users.

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“…The neuroblastoma cell lines used in this study have been previously utilized by our laboratory2223, and were either purchased from American Type Culture Collection (ATCC, www.atcc.org) or were generously provided by Susan Cohn (The University of Chicago Children’s Hospital, Chicago, IL, USA), John Maris (Children’s Hospital of Philadelphia, Philadelphia, PA, USA), or the Children’s Oncology Group (COG) Cell Culture and Xenograft Repository (www.cogcell.org). Neural tumor cell lines U-87, U373, CHLA-02-ATRT and PFSK-1 were purchased from ATCC.…”

Section: Methodsmentioning

confidence: 99%

“…The viability of cells exposed to nifurtimox and BSO was determined using a modified methyl tetrazolium (MTT; Sigma) assay as previously described2223. 0.5–1.0 × 10 4 cells/well of exponentially growing cells were plated in 96-well plates.…”

Section: Methodsmentioning

confidence: 99%

Nifurtimox Is Effective Against Neural Tumor Cells and Is Synergistic with Buthionine Sulfoximine

Zhang

Scorsone

et al. 2016

Sci Rep

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Children with aggressive neural tumors have poor survival rates and novel therapies are needed. Previous studies have identified nifurtimox and buthionine sulfoximine (BSO) as effective agents in children with neuroblastoma and medulloblastoma. We hypothesized that nifurtimox would be effective against other neural tumor cells and would be synergistic with BSO. We determined neural tumor cell viability before and after treatment with nifurtimox using MTT assays. Assays for DNA ladder formation and poly-ADP ribose polymerase (PARP) cleavage were performed to measure the induction of apoptosis after nifurtimox treatment. Inhibition of intracellular signaling was measured by Western blot analysis of treated and untreated cells. Tumor cells were then treated with combinations of nifurtimox and BSO and evaluated for viability using MTT assays. All neural tumor cell lines were sensitive to nifurtimox, and IC50 values ranged from approximately 20 to 210 μM. Nifurtimox treatment inhibited ERK phosphorylation and induced apoptosis in tumor cells. Furthermore, the combination of nifurtimox and BSO demonstrated significant synergistic efficacy in all tested cell lines. Additional preclinical and clinical studies of the combination of nifurtimox and BSO in patients with neural tumors are warranted.

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Sensitivity of neuroblastoma to the novel kinase inhibitor cabozantinib is mediated by ERK inhibition

Cited by 26 publications

References 42 publications

Dual inhibition using cabozantinib overcomes HGF/MET signaling mediated resistance to pan-VEGFR inhibition in orthotopic and metastatic neuroblastoma tumors

Dual inhibition using cabozantinib overcomes HGF/MET signaling mediated resistance to pan-VEGFR inhibition in orthotopic and metastatic neuroblastoma tumors

Binimetinib inhibits MEK and is effective against neuroblastoma tumor cells with low NF1 expression

Nifurtimox Is Effective Against Neural Tumor Cells and Is Synergistic with Buthionine Sulfoximine

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