Improved synthesis of the hypoxia probe 5-deutero-5-fluoro-5-deoxy-azomycin arabinoside (FAZA) as a model process for tritium radiolabeling

Peer reviewedPublisher PD

Investigating Flexibility as a Performance Dimension of a Manufacturing Value Modeling Methodology (MVMM): A Framework for Identifying Flexibility Types in Manufacturing Systems☆

Abstract In recent years manufacturing companies have been faced with various challenges related to volatile demand and changing requirements from customer as well as suppliers. This trend is now even accelerating with a direct impact on the value chain. New technological roadmaps and suggested interventions in manufacturing systems try to solve these challenges and solutions such as the German high tech strategy "Industrie 4.0" or the Italian cluster "Fabbrica Intelligente" which often aimed at enhancing the flexibility of manufacturing systems among many other competitive dimensions. However, these approaches often do not provide a detailed definition of flexibility and its different manifestations. Therefore, the question rises if different types of flexibility, that have an impact on the complete manufacturing system, can be better identified with the existing Manufacturing Value Modeling Methodology (MVMM). This question becomes even more important when considering the potential that smart machines interacting with humans, such as cyber-physical systems (CPS), and the possibility to increase connectivity and data access through technologies, such as the internet of things (IoT), offer for increasing flexibility. Especially due to the various possibilities it becomes even more important to understand, which kind of flexibility is needed for a given problem. Implementing flexibility into the MVMM requires a 'catalog' that makes use of the MVMM framework presenting an overview of internal and external influence factors in order to support the identification of correct solutions and improvements related to functional areas in the manufacturing environment. Starting from a qualitative literature review on manufacturing flexibility, a 'flexibility catalog' is designed, which provides a structural definition of existing flexibility types and their composition as well as providing decision support. In conclusion, the scope of the 'flexibility catalog' is to verify that the flexibility demand fits into the market trends and is aligned to the manufacturing and company strategy, in order to help firms to take decisions and delivering value

Cereblon versus VHL:Hijacking E3 ligases against each other using PROTACs

<div> <div> <div> <p>The von Hippel-Lindau (VHL) and cereblon (CRBN) proteins are substrate recognition subunits of two ubiquitously expressed and biologically important Cullin RING E3 ubiquitin ligase complexes. VHL and CRBN are also the two most popular E3 ligases being recruited by bifunctional Proteolysis-targeting chimeras (PROTACs) to induce ubiquitination and subsequent proteasomal degradation of a target protein. Using homo-PROTACs, VHL and CRBN have been independently dimerized to induce their own degradation. Here we report the design, synthesis and cellular activity of VHL-CRBN hetero-dimerizing PROTACs featuring diverse conjugation patterns. We found that the most active compound 14a induced potent, rapid and profound preferential degradation of CRBN over VHL in cancer cell lines. At lower concentrations, weaker degradation of VHL was instead observed. This work demonstrates proof of concept of designing PROTACs to hijack different E3 ligases against each other, and highlights a powerful and generalizable proximity-induced strategy to achieve E3 ligase knockdown. </p> </div> </div> </div

The Italian Validation of the Gender Minority Stress and Resilience Measure

Transgender and gender nonconforming (TGNC) people experience high levels of minority stress and associated risk for negative mental health outcomes. Notwithstanding, TGNC people may resist the negative effects of minority stress on health through the resilience factors. As no comprehensive measures of gender minority stress and resilience exist in Italy, this study evaluated the psychometric characteristics of an Italian language version of the Gender and Minority Stress and Resilience Measure (GMSR) in an Italian sample of 203 TGNC individuals ranged from 18 to 66 years of age (M = 30.70, SD = 10.79). The GMSR, developed in the United States in 2015, assesses distal stressors (discrimination, rejection, victimization, and nonaffirmation), proximal stressors (internalized transphobia, negative expectations, and nondisclosure), and resilience factors (pride and community connectedness). Confirmatory factor analysis showed that the original 9-factor model had adequate fit to the data obtained from the Italian sample. Criterion validity was partially confirmed, as the stress scales positively correlated with anxiety and depression, and pride negatively correlated with depression, but not anxiety. On the contrary, community connectedness did not correlate with any of the mental health measures. Instead, both convergent and discriminant validity were confirmed as both distal and proximal stressors positively correlated with perceived stress, community connectedness was positively associated with perceived support from friends, and all correlations were below .60. This study offers evidence of the reliability and validity of the GMSR in the Italian context, providing Italian clinicians and researchers with a comprehensive tool to assess gender minority stress in TGNC individuals

Rapid and reversible knockdown of endogenously tagged endosomal proteins via an optimized HaloPROTAC degrader

Inducing post-translational protein knockdown is an important approach to probe biology and validate drug targets. An efficient strategy to achieve this involves expression of a protein of interest fused to an exogenous tag, allowing tag-directed chemical degraders to mediate protein ubiquitylation and proteasomal degradation. Here, we combine improved HaloPROTAC degrader probes with CRISPR/Cas9 genome editing technology to trigger rapid degradation of endogenous target proteins. Our optimized probe, HaloPROTAC-E, a chloroalkane conjugate of high-affinity VHL binder VH298, induced reversible degradation of two endosomally localized proteins, SGK3 and VPS34, with a DC₅₀ of 3–10 nM. HaloPROTAC-E induced rapid (∼50% degradation after 30 min) and complete (<i>D</i>_max of ∼95% at 48 h) depletion of Halo-tagged SGK3, blocking downstream phosphorylation of the SGK3 substrate NDRG1. HaloPROTAC-E more potently induced greater steady state degradation of Halo tagged endogenous VPS34 than the previously reported HaloPROTAC3 compound. Quantitative global proteomics revealed that HaloPROTAC-E is remarkably selective inducing only degradation of the Halo tagged endogenous VPS34 complex (VPS34, VPS15, Beclin1, and ATG14) and no other proteins were significantly degraded. This study exemplifies the combination of HaloPROTACs with CRISPR/Cas9 endogenous protein tagging as a useful method to induce rapid and reversible degradation of endogenous proteins to interrogate their function

Factors Affecting Mortality in 1022 COVID-19 Patients Referred to an Emergency Department in Bergamo during the Peak of the Pandemic

The unexpected outbreak of COVID-19 in the area of Bergamo and the general crisis of personnel and devices has been managed as well as possible during the maximum peak of epidemic; Humanitas Gavazzeni Hospital implemented its facilities and organization in order to optimize the treatment of patients. The number of beds in the Intensive Care Unit (ICU) was doubled (from 16 to 33), and more than 220 beds were dedicated to the COVID-19 patients. This paper analyzes the factors affecting mortality in 1022 COVID-19 patients who referred to Humanitas Gavazzeni between February 25 and March 26, 2020. A total of 274 (34.9%) fatal events were registered: 202 among those admitted to the Intensive Care Unit (ICU) and COVID department and 72 among those treated in Acute Admission Unit Level II (AAUl-2) who died before hospital admission. This paper studies 274 dead cases by analyzing patient's characteristics, physiological and laboratory parameters, symptoms, and the scores of severity of the disease. Patients who had fatal events in the AAUL-2 showed the worst parameters of risk. The most important differences regarded the Apache II score, Glasgow Coma Score (GCS), CRP (C-reactive protein), pH, creatinine, RR (respiratory rate), and asthenia

The bromodomain and extra-terminal domain degrader MZ1 exhibits preclinical anti-tumoral activity in diffuse large B-cell lymphoma of the activated B cell-like type

AIM: Bromodomain and extra-terminal domain (BET) proteins are epigenetic readers that play a fundamental role in transcription regulation. Preclinical and early clinical evidence sustain BET targeting as an anti-cancer approach. BET degraders are chimeric compounds comprising of a BET inhibitor, which allows the binding to BET bromodomains, linked to a small molecule, binder for an E3 ubiquitin ligase complex, triggering BET proteins degradation via the proteasome. These degraders, called proteolysis-targeting chimeras (PROTACs), can exhibit greater target specificity compared to BET inhibitors and overcome some of their limitations, such as the upregulation of the BET proteins themselves. Here are presented data on the anti-tumor activity and the mechanism of action of the BET degrader MZ1 in diffuse large B cell lymphoma (DLBCL) of the activated B-cell like (ABC, ABC DLBCL), using a BET inhibitor as a comparison. METHODS: Established lymphoma cell lines were exposed for 72 h to increasing doses of the compounds. Cell proliferation was evaluated by using an 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazoliumbromide (MTT) assay. Fluorescent-Activated Cell Sorter (FACS) analysis was performed to measure apoptotic activation and RNA sequencing (RNA-Seq) to study the transcriptional changes induced by the compounds. RESULTS: MZ1, and not its negative control epimer cisMZ1, was very active with a median half maximal inhibitory concentration (IC(50)) of 49 nmol/L. MZ1 was more in vitro active than the BET inhibitor birabresib (OTX015). Importantly, MZ1 induced cell death in all the ABC DLBCL cell lines, while the BET inhibitor was cytotoxic only in a fraction of them. BET degrader and inhibitor shared partially similar changes at transcriptome level but the MZ1 effect was stronger and overlapped with that caused cyclin-dependent kinase 9 (CDK9) inhibition. CONCLUSIONS: The BET degrader MZ1 had strong cytotoxic activity in all the ABC DLBCL cell lines that were tested, and, at least in vitro, it elicited more profound effects than BET inhibitors, and encourages further investigations