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Neuroblastoma (NB) is the most common extracranial solid tumor in early childhood. Despite intensive multimodal therapy, nearly half of children with high-risk disease will relapse with therapy-resistant tumors. Dysregulation of MAPK pathway has been implicated in the pathogenesis of relapsed and refractory NB patients, which underscores the possibility of targeting MAPK signaling cascade as a novel therapeutic strategy. In this study, we found that high expressions of RAF family kinases correlated with advanced tumor stage, high-risk disease, tumor progression, and poor overall survival. Targeted inhibition of RAF family kinases with the novel small molecule inhibitor agerafenib abrogated the activation of ERK MAPK pathway in NB cells. Agerafenib significantly inhibited the cell proliferation and colony formation ability of NB cells in vitro, and its combination with traditional chemotherapy showed a synergistic pro-apoptotic effect. More importantly, agerafenib exhibited a favorable toxicity profile, potently suppressed tumor growth, and prolonged survival in NB mouse models. In conclusion, our preclinical data suggest that agerafenib might be an effective therapeutic agent for NB treatment, both as a single-agent and in combination with chemotherapy.

Hepatoblastoma (HB) is the most common pediatric liver malignancy. High-risk patients have poor survival, and current chemotherapies are associated with significant toxicities. Targeted therapies are needed to improve outcomes and patient quality of life. Most HB cases are TP53 wild-type; therefore, we hypothesized that targeting the p53 regulator Murine double minute 4 (MDM4) to reactivate p53 signaling may show efficacy. MDM4 expression was elevated in HB patient samples, and increased expression was strongly correlated with decreased expression of p53 target genes. Treatment with NSC207895 (XI-006), which inhibits MDM4 expression, or ATSP-7041, a stapled peptide dual inhibitor of MDM2 and MDM4, showed significant cytotoxic and antiproliferative effects in HB cells. Similar phenotypes were seen with short hairpin RNA (shRNA)-mediated inhibition of MDM4. Both NSC207895 and ATSP-7041 caused significant upregulation of p53 targets in HB cells. Knocking-down TP53 with shRNA or overexpressing MDM4 led to resistance to NSC207895-mediated cytotoxicity, suggesting that this phenotype is dependent on the MDM4-p53 axis. MDM4 inhibition also showed efficacy in a murine model of HB with significantly decreased tumor weight and increased apoptosis observed in the treatment group. This study demonstrates that inhibition of MDM4 is efficacious in HB by upregulating p53 tumor suppressor signaling.

Hepatoblastoma (HB) is the most common pediatric liver malignancy. Management of HB requires multidisciplinary efforts. The 5-year overall survival of this disease is about 80% in developed countries. Despite advances in the care of these patients, survival in recurrent or treatment-refractory disease is lower than 50%. This is due to more complex tumor biology, including hepatocellular carcinoma (HCC)-like mutations and expression of aggressive gene signatures leading to chemoresistance, vascular invasion, and metastatic spread. The current treatment protocols for pediatric liver cancer do not incorporate targeted therapies, and the ability to test these therapies is limited due to the inaccessibility of cell lines and mouse models. In this review, we discuss the current status of preclinical animal modeling in pediatric liver cancer, primarily HB. Although HB is a rare cancer, the research community has worked together to develop a range of interesting and relevant mouse models for diverse preclinical studies.

// Wenjing Sun 1, 5 , Yesenia Rojas 1 , Hao Wang 2, 6 , Yang Yu 1 , Yongfeng Wang 3 , Zhenghu Chen 3 , Kimal Rajapakshe 4 , Xin Xu 3 , Wei Huang 3 , Saurabh Agarwal 3 , Roma H. Patel 1 , Sarah Woodfield 1 , Yanling Zhao 3 , Jingling Jin 1 , Hong Zhang 2 , Angela Major 7 , M. John Hicks 7 , Jason M. Shohet 3 , Sanjeev A. Vasudevan 1 , Cristian Coarfa 4 , Jianhua Yang 3 and Jed G. Nuchtern 1 1 Pediatric Surgery Division, Michael E. Debakey Department of Surgery, Texas Children’s Hospital, Baylor College of Medicine, Houston, TX 77030, USA 2 Department of Pathology, MD Anderson Cancer Center, Houston, TX 77030, USA 3 Department of Pediatrics, Texas Children’s Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA 4 Department of Molecular and Cell Biology, Baylor College of Medicine, Houston, TX 77030, USA 5 Laboratory of Medical Genetics, Harbin Medical University, Harbin, Heilongjiang 150081, China 6 Department of Hepatopancreatobiliary Surgery, The Second Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150086, China 7 Department of Pathology, Texas Children’s Cancer Center, Baylor College of Medicine, Houston, TX 77030, USA Correspondence to: Jed G. Nuchtern, email: nuchtern@bcm.edu Keywords: neuroblastoma; YK-4-279; chemotherapy; doxorubicin; EWSR1 Abbreviations: NB, neuroblastoma; RHA, RNA helicase A; Dox, doxorubicin Received: July 12, 2017 Accepted: August 21, 2017 Published: October 19, 2017 ABSTRACT Treatment failure in high risk neuroblastoma (NB) is largely due to the development of chemotherapy resistance. We analyzed the gene expression changes associated with exposure to chemotherapy in six high risk NB tumors with the aid of the Connectivity Map bioinformatics platform. Ten therapeutic agents were predicted to have a high probability of reversing the transcriptome changes associated with neoadjuvant chemotherapy treatment. Among these agents, initial screening showed the EWS-FLI1 and RNA helicase A interaction inhibitor YK-4-279, had obvious cytotoxic effects on NB cell lines. Using a panel of NB cell lines, including MYCN nonamplified (SK-N-AS, SH-SY5Y, and CHLA-255), and MYCN amplified (NB-19, NGP, and IMR-32) cell lines, we found that YK-4-279 had cytotoxic effects on all lines tested. In addition, YK-4-279 also inhibited cell proliferation and anchorage-independent growth and induced cell apoptosis of these cells. YK-4-279 enhanced the cytotoxic effect of doxorubicin (Dox). Moreover, YK-4-279 was able to overcome the established chemoresistance of LA-N-6 NB cells. In an orthotopic xenograft NB mouse model, YK-4-279 inhibited NB tumor growth and induced apoptosis in tumor cells through PARP and Caspase 3 cleavage in vivo . While EWS-FLI1 fusion protein is not frequently found in NB, using the R2 public database of neuroblastoma outcome and gene expression, we found that high expression of EWSR1 was associated with poor patient outcome. Knockdown of EWSR1 inhibited the oncogenic potential of neuroblastoma cell lines. Taken together, our results indicate that YK-4-279 might be a promising agent for treatment of NB that merits further exploration.

Neuroblastoma (NB), the most common extracranial pediatric solid tumor, continues to cause significant cancer-related morbidity and mortality in children. Dysregulation of oncogenic receptor tyrosine kinases (RTKs) has been shown to contribute to tumorigenesis in various human cancers and targeting these RTKs has had therapeutic benefit. RET is an RTK which is commonly expressed in NB, and high expression of RET correlates with poor outcomes in patients with NB. Herein we report that RET is required for NB cell proliferation and that the small molecule inhibitor regorafenib (BAY 73-4506) blocks glial cell derived neurotrophic factor (GDNF)-induced RET signaling in NB cells and inhibits NB growth both in vitro and in vivo. We found that regorafenib significantly inhibited cell proliferation and colony formation ability of NB cells. Moreover, regorafenib suppressed tumor growth in both an orthotopic xenograft NB mouse model and a TH-MYCN transgenic NB mouse model. Finally, regorafenib markedly improved the overall survival of TH-MYCN transgenic tumor-bearing mice. In summary, our study suggests that RET is a potential therapeutic target in NB, and that using a novel RET inhibitor, like regorafenib, should be investigated as a therapeutic treatment option for children with NB.

Activating germline mutations of anaplastic lymphoma kinase (ALK) occur in most cases of hereditary neuroblastoma (NB) and the constitutively active kinase activity of ALK promotes cell proliferation and survival in NB. Therefore, ALK kinase is a potential therapeutic target for NB. In this study, we show that the novel ALK inhibitor alectinib effectively suppressed cell proliferation and induces apoptosis in NB cell lines with either wild-type ALK or mutated ALK (F1174L and D1091N) by blocking ALK-mediated PI3K/Akt/mTOR signaling. In addition, alectinib enhanced doxorubicin-induced cytotoxicity and apoptosis in NB cells. Furthermore, alectinib induced apoptosis in an orthotopic xenograft NB mouse model. Also, in the TH-MYCN transgenic mouse model, alectinib resulted in decreased tumor growth and prolonged survival time. These results indicate that alectinib may be a promising therapeutic agent for the treatment of NB.

Neuroblastoma is the most common extracranial malignant solid tumor in children, and drug resistance is a major reason for poor outcomes. Elevated proteasome activity plays an important role in neuroblastoma tumor development and resistance to conventional chemotherapy. Ubiquitin-specific protease 14 (USP14), one of three deubiquitinases associated with the regulatory subunit of the proteasome, is emerging as a potential therapeutic target in multiple tumor types. However, the role of USP14 in neuroblastoma is yet to be elucidated. We found that USP14 inhibition in neuroblastoma via knockdown or a specific inhibitor such as b-AP15 suppressed cell proliferation by inducing cell apoptosis. Furthermore, b-AP15 significantly inhibited neuroblastoma tumor growth in NGP and SH-SY5Y xenograft mouse models. For combination treatment, b-AP15 plus conventional chemotherapeutic agents such as doxorubicin or VP-16 resulted in synergistic antitumor effects on neuroblastoma. Our study demonstrates that USP14 is required for cell viability and is a novel therapeutic target in neuroblastoma. Moreover, USP14 inhibition may add value in combination therapy due to its powerful synergistic effects in treating neuroblastoma.

// Shayahati Bieerkehazhi 1, 2, * , Zhenghu Chen 3, 4, * , Yanling Zhao 4 , Yang Yu 4 , Huiyuan Zhang 4 , Sanjeev A. Vasudevan 5 , Sarah E. Woodfield 5 , Ling Tao 4 , Joanna S. Yi 4 , Jodi A. Muscal 4 , Jonathan C. Pang 4, 6 , Shan Guan 4 , Hong Zhang 2 , Jed G. Nuchtern 5 , Hui Li 7 , Huiwu Li 8 , Jianhua Yang 4 1 Department of Labour Hygiene and Sanitary Science, College of Public Health, Xinjiang Medical University, Urumqi, Xinjiang 830011, P.R. China 2 Department of Pathology, University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA 3 Department of Ophthalmology, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai 200072, P. R. China 4 Texas Children's Cancer Center, Department of Pediatrics, Dan L. Duncan Cancer Center, Baylor College of Medicine, Houston, Texas 77030, USA 5 Division of Pediatric Surgery, Texas Children’s Hospital Department of Surgery, Michael E. DeBakey Department of Surgery, Dan L. Duncan Cancer Center, Baylor College of Medicine, Houston, Texas 77030, USA 6 Department of Biosciences, Weiss School of Natural Sciences, Rice University, Houston, Texas 77005, USA 7 Central Laboratory of Xinjiang Medical University, Urumqi, Xinjiang 830011, P.R. China 8 Cancer Prevention and Research Institute, The Affiliated Tumor Hospital of Xinjiang Medical University, Urumqi, Xinjiang 830011, P.R. China * These authors have contributed equally to this work and should be considered co-first authors Correspondence to: Huiwu Li, email: huiwuli1234@163.com Jianhua Yang, email: jianhuay@bcm.edu Keywords: neuroblastoma, bosutinib, SKI-606, Bosulif, chemotherapy Received: October 29, 2016 Accepted: November 12, 2016 Published: November 26, 2016 ABSTRACT Neuroblastoma (NB) is the most common extracranial solid tumor in children. Aberrant activation of the non-receptor tyrosine kinases Src and c-Abl contributes to the progression of NB. Thus, targeting these kinases could be a promising strategy for NB therapy. In this paper, we report that the potent dual Src/Abl inhibitor bosutinib exerts anti-tumor effects on NB. Bosutinib inhibited NB cell proliferation in a dose-dependent manner and suppressed colony formation ability of NB cells. Mechanistically, bosutinib effectively decreased the activity of Src/Abl and PI3K/AKT/mTOR, MAPK/ERK, and JAK/STAT3 signaling pathways. In addition, bosutinib enhanced doxorubicin (Dox)- and etoposide (VP-16)-induced cytotoxicity in NB cells. Furthermore, bosutinib demonstrated anti-tumor efficacy in an orthotopic xenograft NB mouse model in a similar mechanism as of that in vitro . In summary, our results reveal that Src and c-Abl are potential therapeutic targets in NB and that the novel Src/Abl inhibitor bosutinib alone or in combination with other chemotherapeutic agents may be a valuable therapeutic option for NB patients.

// Xinfang Mao 1, 2, * , Zhenghu Chen 2, 3, * , Yanling Zhao 2 , Yang Yu 2 , Shan Guan 1, 2 , Sarah E. Woodfield 4 , Sanjeev A. Vasudevan 4 , Ling Tao 2 , Jonathan C. Pang 2 , Jiaxiong Lu 2 , Huiyuan Zhang 2 , Fuchun Zhang 1 , Jianhua Yang 2 1 Xinjiang Key Laboratory of Biological Resources and Genetic Engineering, College of Life Science and Technology, Xinjiang University, Urumqi 830046, P. R. China 2 Texas Children’s Cancer Center, Department of Pediatrics, Dan L. Duncan Cancer Center, Baylor College of Medicine, Houston, Texas 77030, USA 3 Department of Ophthalmology, Shanghai Tenth People’s Hospital, Tongji University School of Medicine, Shanghai 200072, P. R. China 4 Division of Pediatric Surgery, Texas Children’s Hospital Department of Surgery, Michael E. DeBakey Department of Surgery, Dan L. Duncan Cancer Center, Baylor College of Medicine, Houston, Texas 77030, USA * These authors have contributed equally to this work and should be considered co-first authors Correspondence to: Fuchun Zhang, email: zfcxju@qq.com Jianhua Yang, email: jianhuay@bcm.edu Keywords: neuroblastoma, EGFR inhibitor, afatinib, apoptosis, chemotherapy Received: October 12, 2016 Accepted: November 08, 2016 Published: November 26, 2016 ABSTRACT Neuroblastoma is the most common extracranial solid tumor in children. The ErbB family of proteins is a group of receptor tyrosine kinases that promote the progression of various malignant cancers including neuroblastoma. Thus, targeting them with small molecule inhibitors is a promising strategy for neuroblastoma therapy. In this study, we investigated the anti-tumor effect of afatinib, an irreversible inhibitor of members of the ErbB family, on neuroblastoma. We found that afatinib suppressed the proliferation and colony formation ability of neuroblastoma cell lines in a dose-dependent manner. Afatinib also induced apoptosis and blocked EGF-induced activation of PI3K/AKT/mTOR signaling in all neuroblastoma cell lines tested. In addition, afatinib enhanced doxorubicin-induced cytotoxicity in neuroblastoma cells, including the chemoresistant LA-N-6 cell line. Finally, afatinib exhibited antitumor efficacy in vivo by inducing apoptosis in an orthotopic xenograft neuroblastoma mouse model. Taken together, these results show that afatinib inhibits neuroblastoma growth both in vitro and in vivo by suppressing EGFR-mediated PI3K/AKT/mTOR signaling. Our study supports the idea that EGFR is a potential therapeutic target in neuroblastoma. And targeting ErbB family protein kinases with small molecule inhibitors like afatinib alone or in combination with doxorubicin is a viable option for treating neuroblastoma.

Background High-risk neuroblastoma has poor outcomes with high rates of relapse despite aggressive treatment, and novel therapies are needed to improve these outcomes. Ponatinib is a multi-tyrosine kinase inhibitor that targets many pathways implicated in neuroblastoma pathogenesis. We hypothesized that ponatinib would be effective against neuroblastoma in preclinical models. Methods We evaluated the effects of ponatinib on survival and migration of human neuroblastoma cells in vitro. Using orthotopic xenograft mouse models of human neuroblastoma, we analyzed tumors treated with ponatinib for growth, gross and histologic appearance, and vascularity. Results Ponatinib treatment of neuroblastoma cells resulted in decreased cell viability and migration in vitro. In mice with orthotopic xenograft neuroblastoma tumors, treatment with ponatinib resulted in decreased growth and vascularity. Conclusions Ponatinib reduces neuroblastoma cell viability in vitro and reduces tumor growth and vascularity in vivo. The antitumor effects of ponatinib suggest its potential as a novel therapeutic agent for neuroblastoma, and further preclinical testing is warranted.

Author(s): Woodfield, Sarah E; Guo, Rong Jun; Liu, Yin; Major, Angela M; Hollingsworth, Emporia Faith; Indiviglio, Sandra; Whittle, Sarah B; Mo, Qianxing; Bean, Andrew J; Ittmann, Michael; Lopez-Terrada, Dolores; Zage, Peter E | Abstract: BackgroundUBE4B is an E3/E4 ubiquitin ligase whose gene is located in chromosome 1p36.22. We analyzed the associations of UBE4B gene and protein expression with neuroblastoma patient outcomes and with tumor prognostic features and histology.MethodsWe evaluated the association of UBE4B gene expression with neuroblastoma patient outcomes using the R2 Platform. We screened neuroblastoma tumor samples for UBE4B protein expression using immunohistochemistry. FISH for UBE4B and 1p36 deletion was performed on tumor samples. We then evaluated UBE4B expression for associations with prognostic factors and with levels of phosphorylated ERK in neuroblastoma tumors and cell lines.ResultsLow UBE4B gene expression is associated with poor outcomes in patients with neuroblastoma and with worse outcomes in all patient subgroups. UBE4B protein expression was associated with neuroblastoma tumor differentiation, and decreased UBE4B protein levels were associated with high-risk features. UBE4B protein levels were also associated with levels of phosphorylated ERK.ConclusionsWe have demonstrated associations between UBE4B gene expression and neuroblastoma patient outcomes and prognostic features. Reduced UBE4B protein expression in neuroblastoma tumors was associated with high-risk features, a lack of differentiation, and with ERK activation. These results suggest UBE4B may contribute to the poor prognosis of neuroblastoma tumors with 1p36 deletions and that UBE4B expression may mediate neuroblastoma differentiation.

Hepatoblastoma is the most common malignant liver tumor in children. Since it is often unresectable and exhibits drug resistance, the treatment of advanced hepatoblastoma is challenging. The orphan nuclear receptor liver receptor homolog-1 (LRH-1) serves prominent roles in malignancy; however, to the best of our knowledge, the role of LRH-1 in hepatoblastoma remains unknown. In the present study, human hepatoblastoma cell lines were analyzed; the mRNA and protein expression levels of LRH-1 were significantly higher in HepG2 and HuH6 cells compared with those in HepT1 cells and control THLE-2 cells. Knockdown of LRH-1 resulted in decreased HepG2 and HuH6 cell proliferation via downregulation of cyclin D1 (CCND1) and c-Myc. Furthermore, treatment with an LRH-1 antagonist (LRA) inhibited the proliferation and colony formation of cell lines in a dose-dependent manner, and induced cell cycle arrest at G(1) phase through inhibition of CCND1 expression. Finally, LRA treatment enhanced the cytotoxic effects of doxorubicin on hepatoblastoma cells. Collectively, these findings suggested that LRH-1 may have an important role in the progression of hepatoblastoma and implicated LRA as a novel, potential therapeutic agent for the treatment of hepatoblastoma.

BACKGROUND Outcomes for children with high-risk neuroblastoma are poor, and improved understanding of the mechanisms underlying neuroblastoma pathogenesis, recurrence, and treatment resistance will lead to improved outcomes. Aberrant growth factor receptor expression and receptor tyrosine kinase signaling are associated with the pathogenesis of many malignancies. A germline polymorphism in the FGFR4 gene is associated with increased receptor expression and activity and with decreased survival, treatment resistance, and aggressive disease for many malignancies. We therefore investigated the role of this FGFR4 polymorphism in neuroblastoma pathogenesis. MATERIALS AND METHODS Germline DNA from neuroblastoma patients and matched controls was assessed for the FGFR4 Gly/Arg388 polymorphism by RT-PCR. Allele frequencies were assessed for association with neuroblastoma patient outcomes and prognostic features. Degradation rates of the FGFR4 Arg388 and Gly388 receptors and rates of receptor internalization into the late endosomal compartment were measured. RESULTS Frequency of the FGFR4 AA genotype and the prevalence of the A allele were significantly higher in patients with neuroblastoma than in matched controls. The Arg388 receptor demonstrated slower degradation than the Gly388 receptor in neuroblastoma cells and reduced internalization into multivesicular bodies. CONCLUSIONS The FGFR4 Arg388 polymorphism is associated with an increased prevalence of neuroblastoma in children, and this association may be linked to differences in FGFR4 degradation rates. Our study provides the first evidence of a role for FGFR4 in neuroblastoma, suggesting that FGFR4 genotype and the pathways regulating FGFR4 trafficking and degradation may be relevant for neuroblastoma pathogenesis.

Neuroblastoma (NB) is the most common extracranial malignant solid tumor seen in children and continues to lead to the death of many pediatric cancer patients. The poor outcome in high risk NB is largely attributed to the development of chemoresistant tumor cells. Doxorubicin (dox) has been widely employed as a potent anti-cancer agent in chemotherapeutic regimens; however, it also leads to chemoresistance in many cancer types including NB. Thus, developing novel small molecules that can overcome dox-induced chemoresistance is a promising strategy in cancer therapy. Here we show that the second generation proteasome inhibitor ixazomib (MLN9708) not only inhibits NB cell proliferation and induces apoptosis in vitro but also enhances dox-induced cytotoxicity in NB cells. Ixazomib inhibits dox-induced NF-κB activity and sensitizes NB cells to dox-induced apoptosis. More importantly, ixazomib demonstrated potent anti-tumor efficacy in vivo by enhancing dox-induced apoptosis in an orthotopic xenograft NB mouse model. Collectively, our study illustrates the anti-tumor efficacy of ixazomib in NB both alone and in combination with dox, suggesting that combination therapy including ixazomib with traditional therapeutic agents such as dox is a viable strategy that may achieve better outcomes for NB patients.

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.

Background 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. Methods 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. Results 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. Conclusions 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. Electronic supplementary material The online version of this article (doi:10.1186/s12885-016-2199-z) contains supplementary material, which is available to authorized users.

Multiple genes involved in endocytosis and endosomal protein trafficking in Drosophila have been shown to function as neoplastic tumor suppressor genes (nTSGs), including Endosomal Sorting Complex Required for Transport-II (ESCRT-II) components vacuolar protein sorting 22 (vps22), vps25, and vps36. However, most studies of endocytic nTSGs have been done in mosaic tissues containing both mutant and non-mutant populations of cells, and interactions among mutant and non-mutant cells greatly influence the final phenotype. Thus, the true autonomous phenotype of tissues mutant for endocytic nTSGs remains unclear. Here, we show that tissues predominantly mutant for ESCRT-II components display characteristics of neoplastic transformation and then undergo apoptosis. These neoplastic tissues show upregulation of c-Jun N-terminal Kinase (JNK), Notch, and Janus Kinase (JAK)/Signal Transducer and Activator of Transcription (STAT) signaling. Significantly, while inhibition of JNK signaling in mutant tissues partially inhibits proliferation, inhibition of JAK/STAT signaling rescues other aspects of the neoplastic phenotype. This is the first rigorous study of tissues predominantly mutant for endocytic nTSGs and provides clear evidence for cooperation among de-regulated signaling pathways leading to tumorigenesis.

Malignant hepatocellular cancers are the most common primary liver malignancies in children, and hepatoblastomas (HBs) account for more than two-thirds of these cases. While most HBs respond to chemotherapy and have favorable outcomes, the 3-year overall survival rate for high-risk HBs is below 50% and guidelines for their classification and treatment are still evolving. HB risk-stratification efforts using clinical, histological, and molecular parameters have been reported to help identify patients that require more or less aggressive therapies in retrospective studies, and are being validated in clinical trials. However, risk assessment is particularly challenging for cancers with certain histologies, including tumors in the recently proposed provisional hepatocellular neoplasm not otherwise specified (HCN NOS) category. HCN NOSs exhibit either intermediate or combined HB and hepatocellular carcinoma (HCC) histological features, and while neoplasms with such features were observed over a decade ago, only a handful have been characterized and little is known about their biology and clinical features. Here, we molecularly characterized a series of clinically annotated HCN NOSs that demonstrated either intermediate HB/HCC histology or distinct coexisting areas with HB and HCC histological features. In addition, molecular profiling of HBs demonstrating focal pleomorphism or anaplasia (HB FPA) revealed underlying biological features previously observed in HCCs. Our study suggested that HCN NOSs and HB FPAs are aggressive tumors, irrespective of patient age or resectability. Consequently, we designated them collectively as HBs with carcinoma features (HBCs) and outlined histological and molecular characteristics for their diagnosis and treatment. In our single-institution study, transplanted HBC patients were significantly and more than twice as likely to have good outcomes, highlighting the importance of molecular testing and aggressive early intervention.