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

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

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

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

Boron Doped Diamond Electrodes for Direct Measurement in Biological Fluids: An In Situ Regeneration Approach

International audienceBoron doped diamond (BDD) electrodes are extremely promising in the field of biomedical applications as they exhibit a unique combination of properties. Despite these advantages, BDD electrodes are prone to fouling when used in biological fluids (urine, blood plasma), and synthetic fluids. We propose a electrochemical (EC) treatment where a train of short cathodic and/or anodic pulses are applied to clean fouled electrodes. This technique can be used to retrieve the lost reactivity, characterized by electron transfer rate k0 of the boron doped diamond electrodes, thereby enhancing their reusability over long period of measurements without degradation of the signal, thus significantly extending the field of monitoring and surveying applications. The technique does not require the use of a specific medium and thus can be directly performed in the probed fluid. Although an aqueous electrolyte containing non-electroactive species is preferred for EC activation, it can also be done in biological fluids such as blood, urine etc, thereby opening the field for $in-vivo$ analysis. Through Electrochemical impedance spectroscopy (EIS) it was observed that the k$_0$ value was increased up to 0.1 cm s$^{−1}$ after the activation process. This technique improves the sensitivity, reproducibility and lifetime of the electrodes to a considerable extent

Experiences that \u201creach the heart\u201d. Taking part in a whole body dissection course at the University of Malta

This article summarizes the activities of the four-week whole body dissection course the main authors participated in in August 2016 at the dissection hall of the University of Malta (UoM). Our team comprised 10 second-year medicine students from University of Palermo chosen among who had passed the Human Anatomy exam brilliantly. The need to move to the UoM to take part in such activity derives from the lack of practice approach in Italian schools of medicine, focused mostly on the theoretical studies, neglecting practical experience. The heart dissection reveal itself as a huge opportunity to finally apply our anatomical knowledge, improving it and enabling us to compare images took from books to the actual organ. We had the chance to handle a real heart, to appreciate its weight and consistence. We took part in coronary artery courses focusing on their functions within the heart machinery.This article summarizes the activities of the four-week whole body dissection course the main authors partecipated in August 2016 at the dissection hall of the University of Malta (UoM). Our team comprised 10 second-year medicine students from University of Palermo chosen among who had passed the Human Anatomy exam brilliantly. The need to molve to the UoM to take part in such activity derives from the lack of practice approach in Italian schools of medicine, focused mostly on the theoretical studies, neglecting practical experience. The heart dissection reveal itself as a huge opportunity to finally apply our anatomical knowledge, improving it and enabling us to compare image took from books to the actual organ. We had the chance to handle a real heart, to appreciate its weight and consistence. We took part in coronary artery courses focusing on their functions within the heart machinery

Timing performance of a Micro-Channel-Plate Photomultiplier Tube

The spatial dependence of the timing performance of the R3809U-50 Micro-Channel-Plate PMT (MCP-PMT) by Hamamatsu was studied in high energy muon beams. Particle position information is provided by a GEM tracker telescope, while timing is measured relative to a second MCP-PMT, identical in construction. In the inner part of the circular active area (radius r5.5 mm) the time resolution of the two MCP-PMTs combined is better than 10 ps. The signal amplitude decreases in the outer region due to less light reaching the photocathode, resulting in a worse time resolution. The observed radial dependence is in quantitative agreement with a dedicated simulation. With this characterization, the suitability of MCP-PMTs as t0 reference detectors has been validated.Peer reviewe

Precise timing with the PICOSEC-Micromegas detector

This work presents the concept of the PICOSEC-Micromegas de-tector to achieve a time resolution below 30 ps. PICOSEC consists of a two-stageMicromegas detector coupled to a Cherenkov radiator and equipped with a photo-cathode. The results from single-channel prototypes as well as the understanding ofthe detector in terms of detailed simulations and preliminary results from a multi-channel prototype are presented.Peer reviewe

FICD acts bifunctionally to AMPylate and de-AMPylate the endoplasmic reticulum chaperone BiP

Protein folding homeostasis in the endoplasmic reticulum (ER) is defended by an unfolded protein response that matches ER chaperone capacity to the burden of unfolded proteins. As levels of unfolded proteins decline, a metazoan-specific FIC-domain-containing ER-localized enzyme (FICD) rapidly inactivates the major ER chaperone BiP by AMPylating T518. Here we show that the single catalytic domain of FICD can also release the attached AMP, restoring functionality to BiP. Consistent with a role for endogenous FICD in de-AMPylating BiP, FICD$_{-/-}$ hamster cells are hypersensitive to introduction of a constitutively AMPylating, de-AMPylation-defective mutant FICD. These opposing activities hinge on a regulatory residue, E234, whose default state renders FICD a constitutive de-AMPylase $\textit{in vitro}$. The location of E234 on a conserved regulatory helix and the mutually antagonistic activities of FICD $\textit{in vivo}$, suggest a mechanism whereby fluctuating unfolded protein load actively switches FICD from a de-AMPylase to an AMPylase.Supported by Wellcome Trust Principal Research Fellowship to D.R. (Wellcome 200848/Z/16/Z), a UK Medical Research Council PhD studentship to L.A.P. and a Wellcome Trust Strategic Award to the Cambridge Institute for Medical Research (Wellcome 100140)

Comparing Local Governments' Performance Internationally: A Mission Impossible?

While some attempt has been made to address cross-national trends in performance measurement systems (PMSs) in local governments, very few systemic efforts have been attempted to uncover similarities and differences. This article seeks to advance the understanding of the variables researchers should consider when comparing the adoption characteristics and process of PMSs across countries at the local level. Specifically, a framework is constructed to argue that the intersection of the level of performance regime, the types of intergovernmental relationships and the focus of PMS in use on the different types of measure, shape a specific \u2018performance measurement context\u2019 that affects comparability. The framework is then empirically motivated by a comparison between Italy and Michigan, USA. For practitioners and analysts, the framework is a tool to guide effective comparisons across service areas for applied research and organizational learning purposes