Acquired resistance of pancreatic cancer cells to treatment with gemcitabine and HER-inhibitors is accompanied by increased sensitivity to STAT3 inhibition

Drug-resistance is a major contributing factor for the poor prognosis in patients with pancreatic cancer. We have shown previously that the irreversible ErbB family blocker afatinib, is more effective than the reversible EGFR tyrosine kinase inhibitor erlotinib in inhibiting the growth of human pancreatic cancer cells. The aim of this study was to develop human pancreatic cancer cell (BxPc3) variants with acquired resistance to treatment with gemcitabine, afatinib, or erlotinib, and to investigate the molecular changes that accompany the acquisition of a drug-resistant phenotype. We also investigated the therapeutic potential of various agents in the treatment of such drug-resistant variants. Three variant forms of BxPc3 cells with acquired resistance to gemcitabine (BxPc3GEM), afatinib (BxPc3AFR) or erlotinib (BxPc3OSIR) were developed following treatment with increasing doses of such drugs. The expression level, mutational and phosphorylation status of various growth factor receptors and downstream cell signaling molecules were determined by FACS, human phopsho-RTK array, and western blot analysis while the sulforhodamine B assay was used for determining the effect of various agents on the growth of such tumours. We found that all three BxPc3 variants with acquired resistance to gemcitabine (BxPc3GEM), afatinib (BxPc3AFR) or erlotinib (BxPc3OSIR) also become less sensitive to treatment with the two other agents. Acquisition of resistance to these agents was accompanied by upregulation of p-c-MET, p-STAT3, CD44, increased autocrine production of EGFR ligand amphiregulin and differential activation status of EGFR tyrosine residues as well as downregulation of total and p-SRC. Of all therapeutic interventions examined, including the addition of an anti-EGFR antibody ICR62, an anti-CD44 monoclonal antibody, and of STAT3 or c-MET inhibitors, only treatment with the STAT3 inhibitor Stattic produced a higher growth inhibitory effect in all three drug-resistant variants. In addition, treatment with a combination of afatinib with either c-MET inhibitor Crizotinib or Stattic resulted in an additive or synergistic growth inhibition in all three variants. Our results suggest that activation of STAT3 may play an important role in the acquisition of resistance to gemcitabine and HER inhibitors in pancreatic cancer and warrant further studies on the therapeutic potential of STAT3 inhibitors in such a setting

Afatinib efficacy against squamous cell carcinoma of the head and neck cell lines in vitro and in vivo.

Epidermal growth factor receptor (EGFR) inhibitors have demonstrated efficacy in squamous cell carcinoma of the head and neck (SCCHN). In addition to EGFR, other ErbB family members are expressed and activated in SCCHN. Afatinib is an ErbB family blocker that has been approved for treating patients with EGFR-mutated nonsmall cell lung cancer. We sought to determine the efficacy of afatinib in preclinical models and compare this to other EGFR-targeted agents. Afatinib efficacy was characterized in a panel of ten SCCHN cell lines and found to be most effective against cell lines amplified for EGFR. Afatinib had lower IC(50) values than did gefitinib against the same panel. Two EGFR-amplified cell lines that are resistant to gefitinib are sensitive to afatinib. Cetuximab was not found to have a synergistic effect with afatinib either in vitro or in vivo. Both afatinib and cetuximab were effective in tumor xenograft model. Afatinib is an effective agent in SCCHN especially in models with EGFR amplification

Crustaceans and Marine Heterobranchia: A New Symbiotic Relationship in the Mediterranean Sea

The "solar-powered" Elysia timida (Risso, 1818) is an endemic Mediterranean sacoglossan living in rocky substrates at shallow water. During a scuba dive, one E. timida was photographed and collected. The observation revealed the presence of a small crustacean of the Pinnotheridae family. It was not possible to assign the crustacean to a genus, but it shares typical ecological and external morphological features with other Pinnotherinae species. This is the first report of a pea crab hosted by a non-shelled Heterobranchia and the first case of a symbiotic association between crustaceans and marine Heterobranchia reported in the Mediterranean Sea

Analisi di rischio con metodologia ALBA, studio microclimatico e sue applicazioni in Florette Italia, una piccola-media azienda di prodotti di IV gamma

LAUREA MAGISTRALEL’analisi di rischio è il miglior strumento per individuare tutti i rischi in un'azienda e studiare soluzioni atte alla eliminazione o alla mitigazione degli stessi: lo studio, e l’applicazione delle eventuali soluzioni individuate, è quindi di primaria importanza per la salute e la salvaguardia dei lavoratori. Durante lo stage presso l’azienda Florette Italia, si è svolto un processo di analisi dei rischi e del sistema con la metodologia ALBA (Algorithm Logic Bayesian Analysis), un generatore dinamico di alberi degli eventi. Questa metodologia si basa su un'approfondita analisi del sistema, che è stata svolta con le tecniche più diffuse e utilizzate, quali Functional Analysis e analisi dei flussi interni, di cui si tratta nel Capitolo 1 e Capitolo 2. In questa prima parte del progetto, condotta nelle fasi iniziali dello stage, si sono redatti anche i primi documenti necessari al conseguimento della certificazione OHSAS 18001, lo standard internazionale per l’implementazione di un sistema di gestione della sicurezza e della salute dei lavoratori. Grazie a questa approfondita analisi, al continuo e giornaliero confronto con i lavoratori e dirigenti, e allo studio del Documento Valutazione Rischi (D.lgs. n. 81/2008), sono emersi diversi elementi critici per la salute e la sicurezza dei lavoratori. In particolare, sono state individuate procedure inadeguate, o mancanti, per l’utilizzo di sostanze chimiche in determinate zone dello stabilimento, e un insufficiente fornitura di vestiario ai lavoratori, per affrontare le basse temperature nei reparti di produzione. Nel Capitolo 3 vengono quindi descritte le procedure mancanti, redatte grazie alle schede di sicurezza e alla collaborazione del Responsabile Qualità e dei responsabili di linea e prontamente messe in atto, e le modifiche suggerite all’azienda, in parte già attuate dalla direzione. Nel capitolo 4 viene proposto lo svolgimento di un'analisi microclimatica approfondita degli ambienti, eseguita seguendo la norma UNI ENV ISO 11079:2007 e norme ad essa collegate: con questo studio, sono stati individuati i limiti di protezione richiesta (IREQ – Insulation Required) e i necessari indumenti volti a garantire il corretto isolamento termico dei lavoratori. Con le indicazioni fornite, sono state individuate diverse combinazioni di vestiario e, dopo alcune settimane di valutazione da parte dei lavoratori, la direzione ha proceduto all’acquisto. Nel capitolo 5 infine viene descritta l’analisi ALBA: questa metodologia induttiva è fondata sui tre principi di coerenza della statistica e permette di descrivere in modo completo, da un punto di vista logico e stocastico, l’intero sviluppo del sistema. Ciò conduce all'individuazione di tutti i possibili percorsi in cui esso può evolvere tramite l’identificazione di un numero adeguato e coerente di domande e vincoli. L'analisi dei risultati ottenuti con la metodologia ALBA ha evidenziato tre possibili ambiti di intervento da parte dell'azienda per migliorare la sicurezza. Nello specifico si propone la realizzazione di una formazione di più alto livello e aggiornamento dei dipendenti; una maggiore supervisione del lavoro e la fornitura di più adeguati Dispositivi di Protezione Individuali (DPI). La simulazione dell'introduzione di queste tre proposte mostra come, con un minimo impiego di risorse, sia facilmente raggiungibile un miglioramento generale della sicurezza. Inoltre queste procedure potrebbero essere inserite nel manuale per la certificazione OHSAS 18001, qualora l'azienda riprendesse in considerazione il conseguimento della stessa. A completamento di questo studio, emerge che sarebbe inoltre possibile realizzare queste proposte attraverso la reintroduzione di una figura operativa, già presente nei vecchi organigrammi ma da tempo non più implementata: il responsabile di produzione. Su tale figura ricadrebbero la responsabilità della formazione e dell'aggiornamento specialistico dei lavoratori, la supervisione dei turni lavorativi e le misure di correzione adeguate di comportamenti non idonei. Inoltre, si otterrebbe una semplificazione dei mansionari in uso attualmente, e un'ottimizzazione nella implementazione e successiva gestione della eventuale certificazione OHSAS 18001.Risk analysis is the best tool to identify all the risks in a company and to study and propose solutions suitable for removing or mitigating risks. Therefore, this assessment and the application of any of the identified solutions are of primary importance to the health and protection of the workers. During the internship in Florette Italia, it was conducted a risk assessment and system analysis with ALBA methodology (Algorithm Logic Bayesian Analysis), a dynamic event tree generator, which led to a number of proposals for the improvement of the activities. ALBA methodology works at its best when an accurate and complete system analysis is performed: as described in the first and second chapter, the analysis has been conducted with the classical methods of investigation, like Functional and System analysis. This was completed during the first part of the internship, when the first documents needed for the OHSAS 18001 certification, the international applied standard for occupational health and safety management systems, were also drafted. Thanks to this complete and detailed analysis, to the continued and daily talking with workers and executives, and to the study of the DVR (the Italian certification of the risk assessment, due to the state decree 81/2008), it was possible to identify some criticalities regarding health and safety of the employees. In particular, inadequate or missing standard procedures were identified, especially for the use of chemicals (Sodium metabisulfite and Sodium metabisulfite) in certain areas of the factory, and the absence of proper and protective clothes against the cold environment, which were not provided by the company. In Chapter 3, the missing procedures and some proposals for a process modification are described. Thanks to safety data sheets and the collaboration with the Quality Manager and Line Manager of the company, these procedures were designed and immediately implemented. Other procedures and improvements for the safety in working areas were proposed and were included; the management has already carried out some of them. The risk analysis of the thermal environment, conducted in compliance with the UNI ENV ISO 11079:2007 and its references normative, is described in Chapter 4: effective clothing insulation, required to maintain the thermal balance of the body, was achieved after proper data gathering, and a solution of the desired clothes was provided. After preliminary testing, the executives decide to buy new uniforms and work clothes. Finally, Chapter 5 describes the Risk assessment conducted with ALBA methodology, a new method that generates the entire spectrum of the risk, and links together the critical components and magnitudes of the scenario. From the analysis of the results obtained with ALBA methodology, three new system procedures can be suggested: 1. put in place better trainings and wider skill learning update for workers; 2. continuous supervision of the work environment; 3. provide better Individual Protection Devices (IDP). The simulation with ALBA shows how, thanks to the strategy of implementing these three procedures leads to the achievement of significant risks reduction with a small economic effort. Furthermore, if the process for the OHSAS 18001 certification is resumed, these procedures could be easily added to the Occupational, Health and Safety Management System (OHSMS). The strategy can also be implemented with the reintroduction of the Plant Manager, who (as used to be some years ago) should be empowered to supervise workers' training and update, and plant and production control. This will eventually help in reducing risks and achieving an overall better plant management, providing better communication and relationship between workers and executives

NEW FRONTIERS IN THE COGNITIVE ASSESSMENT OF AMYOTROPHIC LATERAL SCLEROSIS: BRAIN COMPUTER INTERFACE AND EYE TRACKING

Background: Over the last 20 years, cognitive and behavioural alterations in amyotrophic lateral sclerosis (ALS) have been recognized as an integral part of the disease. A proportion of patients present with a full-blown frontotemporal dementia syndrome, while selective cognitive changes are more commonly found, especially regarding frontal-executive abilities. Moreover, recent studies have highlighted a broader cognitive involvement in this population, concerning language and social cognition. Despite the increased awareness of ALS as a multisystem disorder and the importance of an accurate cognitive evaluation of these patients, the traditional neuropsychological \u2018paper and pencil\u2019 tools do not compensate for patients\u2019 physical disability and can not be adequately used in the moderate-advanced stages of the disease. Objective: To investigate the use of P300-based Brain Computer Interface (BCI) and Eye Tracking (ET) technology for the administration of motor-verbal free cognitive measures in ALS. Materials and Methods: 34 patients diagnosed with ALS and 30 healthy subjects have been recruited. All participants underwent the BCI and ET-based neuropsychological assessment, together with three traditional cognitive screening tools (Frontal Assessment Battery - FAB; Montreal Cognitive Assessment \u2013 MoCA; Working Memory subtest of the Brief Assessment of Cognition in Schizophrenia), two psychological questionnaires (Beck Depression Inventory - BDI; State-Trate Anxiety Inventory - STAI-Y) and a usability questionnaire. For patients, also respiratory examination was performed, and the Frontal Behavioural Inventory - FBI was carried out with caregivers. Results: Significant correlations were observed between the traditional cognitive measures and the BCI- and ET-based neuropsychological assessment, mainly concerning accuracy and time-related variables in the ALS patients sample. Patients provided comparable rates than controls with regard to the BCI and ET usability. Conclusions: The developed motor-verbal free neuropsychological battery allows a longitudinal cognitive assessment during the course of the disease, also when traditional measures are not fully administrable, providing relevant information for clinical practice and ethical issues. Further work will be aimed at refining the developed system and enlarging the cognitive spectrum investigated

Bulk Properties of Liquids and Molecular Properties in Liquids from a Combination of Quantum Chemical Calculations and Classical Simulations

In the first Column of this series we presented an introduction to the title subject including a historical background and some educational aspects. Here, we show how to calculate bulk properties of liquids without claiming to be comprehensive. The next Column will end the cycle with an introduction to the calculation of solvent effects on nuclear and molecular properties