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Introduction: Microwave imaging presents several potential advantages including its non-ionising and harmless nature. This open, multicentric, interventional, prospective, non-randomised trial aims to validate MammoWave's artificial intelligence (AI)-based classification algorithm, leveraging microwave imaging, to achieve a sensitivity exceeding 75% and a specificity exceeding 90% in breast screening., Methods and Analysis: 10 000 volunteers undergoing regular mammographic breast cancer screening will be recruited across 9 European centres and invited to participate in the clinical study, involving MammoWave testing on both breasts. MammoWave results will be checked against the reference standard, to be intended as the output of conventional breast examination path (with histological confirmation of cancer cases) with 2 years follow-up. Anonymised clinical and MammoWave's results, including microwave images, associated features and a label provided by the AI-based classification algorithm, will be collected and stored in a dedicated electronic case report form. The prospective study will involve a comparative analysis between the output of the conventional breast examination path (control intervention) and the labels provided by MammoWave's AI system (experimental intervention). These labels will categorise breasts into two groups: breast With Suspicious Finding, indicating the presence of a suspicious lesion or No Suspicious Finding, indicating the absence of a lesion or the presence of a low-suspicion lesion. This trial aims to provide evidence regarding the novel MammoWave's AI system for detecting breast cancer in asymptomatic populations during screening., Ethics and Dissemination: This study was approved by the Research Ethics Committee of the Liguria Region (CET), Italy (CET-Liguria: 524/2023-DB id 13399), the Research Ethics Committee of Complejo Hospitalario de Toledo (CEIC), Spain (CEIC-1094), the National Ethics Committee for Clinical Research (CEIC), Portugal (CEIC-2311KC814), the Bioethical Committee of Pomeranian Medical University in Szczecin, Poland (KB-006/23/2024) and the Zurich Cantonal Ethics Commission, Switzerland (BASEC 2023-D0101). The findings of this study will be disseminated through academic and scientific conferences as well as peer-reviewed journals., Trial Registration Number: NCT06291896., Competing Interests: Competing interests: NG, GT, BK and Alessandra Bigotti are employed by UBT—Umbria Bioengineering Technologies Srl. NG and GT are shareholders of UBT—Umbria Bioengineering Technologies. UBT—Umbria Bioengineering Technologies Srl manufactured the microwave imaging device known as MammoWave. This does not alter our adherence to BMJ Open policies., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY. Published by BMJ.)

SARS-CoV-2 (SCoV2) and its variants of concern pose serious challenges to the public health. The variants increased challenges to vaccines, thus necessitating for development of new intervention strategies including anti-virals. Within the international Covid19-NMR consortium, we have identified binders targeting the RNA genome of SCoV2. We established protocols for the production and NMR characterization of more than 80 % of all SCoV2 proteins. Here, we performed an NMR screening using a fragment library for binding to 25 SCoV2 proteins and identified hits also against previously unexplored SCoV2 proteins. Computational mapping was used to predict binding sites and identify functional moieties (chemotypes) of the ligands occupying these pockets. Striking consensus was observed between NMR-detected binding sites of the main protease and the computational procedure. Our investigation provides novel structural and chemical space for structure-based drug design against the SCoV2 proteome., (© 2022 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Novel and efficient strategies need to be developed to interfere with the SARS-CoV-2 virus. One of the most promising pharmaceutical targets is the nucleocapsid protein (N), responsible for genomic RNA packaging. N is composed of two folded domains and three intrinsically disordered regions (IDRs). The globular RNA binding domain (NTD) and the tethered IDRs are rich in positively charged residues. The study of the interaction of N with polyanions can thus help to elucidate one of the key driving forces responsible for its function, i.e., electrostatics. Heparin, one of the most negatively charged natural polyanions, has been used to contrast serious cases of COVID-19 infection, and we decided to study its interaction with N at the molecular level. We focused on the NTR construct, which comprises the NTD and two flanking IDRs, and on the NTD construct in isolation. We characterized this interaction using different nuclear magnetic resonance approaches and isothermal titration calorimetry. With these tools, we were able to identify an extended surface of NTD involved in the interaction. Moreover, we assessed the importance of the IDRs in increasing the affinity for heparin, highlighting how different tracts of these flexible regions modulate the interaction.

The SARS-CoV-2 nucleocapsid (N) protein is crucial for the highly organized packaging and transcription of the genomic RNA. Studying atomic details of the role of its intrinsically disordered regions (IDRs) in RNA recognition is challenging due to the absence of structure and to the repetitive nature of their primary sequence. IDRs are known to act in concert with the folded domains of N and here we use NMR spectroscopy to identify the priming events of N interacting with a regulatory SARS-CoV-2 RNA element. 13 C-detected NMR experiments, acquired simultaneously to 1 H detected ones, provide information on the two IDRs flanking the N-terminal RNA binding domain (NTD) within the N-terminal region of the protein (NTR, 1-248). We identify specific tracts of the IDRs that most rapidly sense and engage with RNA, and thus provide an atom-resolved picture of the interplay between the folded and disordered regions of N during RNA interaction.

The highly infectious disease COVID-19 caused by the Betacoronavirus SARS-CoV-2 poses a severe threat to humanity and demands the redirection of scientific efforts and criteria to organized research projects. The international COVID19-NMR consortium seeks to provide such new approaches by gathering scientific expertise worldwide. In particular, making available viral proteins and RNAs will pave the way to understanding the SARS-CoV-2 molecular components in detail. The research in COVID19-NMR and the resources provided through the consortium are fully disclosed to accelerate access and exploitation. NMR investigations of the viral molecular components are designated to provide the essential basis for further work, including macromolecular interaction studies and high-throughput drug screening. Here, we present the extensive catalog of a holistic SARS-CoV-2 protein preparation approach based on the consortium's collective efforts. We provide protocols for the large-scale production of more than 80% of all SARS-CoV-2 proteins or essential parts of them. Several of the proteins were produced in more than one laboratory, demonstrating the high interoperability between NMR groups worldwide. For the majority of proteins, we can produce isotope-labeled samples of HSQC-grade. Together with several NMR chemical shift assignments made publicly available on covid19-nmr.com , we here provide highly valuable resources for the production of SARS-CoV-2 proteins in isotope-labeled form., Competing Interests: CH was employed by Signals GmbH & Co. KG. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Altincekic, Korn, Qureshi, Dujardin, Ninot-Pedrosa, Abele, Abi Saad, Alfano, Almeida, Alshamleh, de Amorim, Anderson, Anobom, Anorma, Bains, Bax, Blackledge, Blechar, Böckmann, Brigandat, Bula, Bütikofer, Camacho-Zarco, Carlomagno, Caruso, Ceylan, Chaikuad, Chu, Cole, Crosby, de Jesus, Dhamotharan, Felli, Ferner, Fleischmann, Fogeron, Fourkiotis, Fuks, Fürtig, Gallo, Gande, Gerez, Ghosh, Gomes-Neto, Gorbatyuk, Guseva, Hacker, Häfner, Hao, Hargittay, Henzler-Wildman, Hoch, Hohmann, Hutchison, Jaudzems, Jović, Kaderli, Kalniņš, Kaņepe, Kirchdoerfer, Kirkpatrick, Knapp, Krishnathas, Kutz, zur Lage, Lambertz, Lang, Laurents, Lecoq, Linhard, Löhr, Malki, Bessa, Martin, Matzel, Maurin, McNutt, Mebus-Antunes, Meier, Meiser, Mompeán, Monaca, Montserret, Mariño Perez, Moser, Muhle-Goll, Neves-Martins, Ni, Norton-Baker, Pierattelli, Pontoriero, Pustovalova, Ohlenschläger, Orts, Da Poian, Pyper, Richter, Riek, Rienstra, Robertson, Pinheiro, Sabbatella, Salvi, Saxena, Schulte, Schiavina, Schwalbe, Silber, Almeida, Sprague-Piercy, Spyroulias, Sreeramulu, Tants, Tārs, Torres, Töws, Treviño, Trucks, Tsika, Varga, Wang, Weber, Weigand, Wiedemann, Wirmer-Bartoschek, Wirtz Martin, Zehnder, Hengesbach and Schlundt.)

The nucleocapsid protein N from SARS-CoV-2 is one of the most highly expressed proteins by the virus and plays a number of important roles in the transcription and assembly of the virion within the infected host cell. It is expected to be characterized by a highly dynamic and heterogeneous structure as can be inferred by bioinformatics analyses as well as from the data available for the homologous protein from SARS-CoV. The two globular domains of the protein (NTD and CTD) have been investigated while no high-resolution information is available yet for the flexible regions of the protein. We focus here on the 1-248 construct which comprises two disordered fragments (IDR1 and IDR2) in addition to the N-terminal globular domain (NTD) and report the sequence-specific assignment of the two disordered regions, a step forward towards the complete characterization of the whole protein.

Many properties of intrinsically disordered proteins (IDPs), or protein regions (IDRs), are modulated by the nature of amino acid side chains as well as by local solvent exposure. We propose a set of exclusively heteronuclear NMR experiments to investigate these features in different experimental conditions that are relevant for physiological function. The proposed approach is generally applicable to many IDPs/IDRs whose assignment is available in the Biological Magnetic Resonance Bank (BMRB) to investigate how their properties are modulated by different, physiologically relevant conditions. The experiments, tested on α-synuclein, are then used to investigate how α-synuclein senses Ca 2+ concentration jumps associated with the transmission of nerve signals. Novel modules in the primary sequence of α-synuclein optimized for calcium sensing in highly flexible, disordered protein segments are identified., (© 2020 Wiley-VCH GmbH.)

Intrinsically disordered proteins (IDPs) as well as intrinsically disordered regions (IDRs) of complex protein machineries have recently been recognized as key players in many cellular functions. NMR represents a unique tool to access atomic resolution structural and dynamic information on highly flexible IDPs/IDRs. Improvements in instrumental sensitivity made heteronuclear direct detection possible for biomolecular NMR applications. The CON experiment has become one of the most useful NMR experiments to get a snapshot of an IDP/IDR in conditions approaching physiological ones. The availability of NMR spectrometers equipped with multiple receivers now enables the acquisition of several experiments simultaneously instead of one after the other. Here, we propose several variants of the CON experiment in which, during the recovery delay, a second two-dimensional experiment is acquired, either based on 1 H detection (CON//HN) or on 15 N detection (CON//btNH, CON//(H)CAN). The possibility to collect simultaneous snapshots of an IDP/IDR through different two-dimensional spectra provides a novel tool to follow chemical reactions, such as the occurrence of posttranslational modifications, as well as to study samples of limited lifetime such as cell lysates or whole cells., (Copyright © 2019 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

Aim: To summarize the current knowledge about the potential relationship between hepatitis C virus (HCV) infection and the risk of several extra-liver cancers., Methods: We performed a systematic review of the literature, according to the Preferred Reporting Items for Systematic reviews and Meta-Analysis (PRISMA) Statement. We extracted the pertinent articles, published in MEDLINE and the Cochrane Library, using the following search terms: neoplasm/cancer/malignancy/tumor/carcinoma/adeno-carcinoma and non-Hodgkin lymphomas, kidney/renal-, cholangio-, pancreatic-, thyroid-, breast-,oral-, skin-, prostate-, lung-, colon-, stomach-, haematologic. Case series, case-series with control-group, case-control, cohort-studies as well as meta-analyses, written in English were collected. Some of the main characteristics of retrieved trials, which were designed to investigate the prevalence of HCV infection in each type of the above-mentioned human malignancies were summarised. A main table was defined and included a short description in the text for each of these tumours, whether at least five studies about a specific neoplasm, meeting inclusion criteria, were available in literature. According to these criteria, we created the following sections and the corresponding tables and we indicated the number of included or excluded articles, as well as of meta-analyses and reviews: (1) HCV and haematopoietic malignancies; (2) HCV and cholangiocarcinoma; (3) HCV and pancreatic cancer; (4) HCV and breast cancer; (5) HCV and kidney cancer; (6) HCV and skin or oral cancer; and (7) HCV and thyroid cancer., Results: According to available data, a clear correlation between regions of HCV prevalence and risk of extra-liver cancers has emerged only for a very small group of types and histological subtypes of malignancies. In particular, HCV infection has been associated with: (1) a higher incidence of some B-cell Non-Hodgkin-Lymphoma types, in countries, where an elevated prevalence of this pathogen is detectable, accounting to a percentage of about 10%; (2) an increased risk of intra-hepatic cholangiocarcinoma; and (3) a correlation between HCV prevalence and pancreatic cancer (PAC) incidence., Conclusion: To date no definitive conclusions may be obtained from the analysis of relationship between HCV and extra-hepatic cancers. Further studies, recruiting an adequate number of patients are required to confirm or deny this association.

Differential scanning fluorimetry (DSF) is a technique that reports protein thermal stability via the selective recognition of unfolded states by fluorogenic dyes. However, DSF applications remain limited by protein incompatibilities with existing DSF dyes. Here we overcome this obstacle with the development of a protein-adaptive DSF platform (paDSF) that combines a dye library 'Aurora' with a streamlined procedure to identify protein–dye pairs on demand. paDSF was successfully applied to 94% (66 of 70) of proteins, tripling the previous compatibility and delivering assays for 66 functionally and biochemically diverse proteins, including 10 from severe acute respiratory syndrome coronavirus 2. We find that paDSF can be used to monitor biological processes that were previously inaccessible, demonstrated for the interdomain allostery of O-GlcNAc transferase. The chemical diversity and varied selectivities of Aurora dyes suggest that paDSF functionality may be readily extended. paDSF is a generalizable tool to interrogate protein stability, dynamics and ligand binding. A library of fluorogenic dyes enables the measurement of protein unfolding for a large range of proteins. [ABSTRACT FROM AUTHOR]

Nuclear magnetic resonance (NMR) spectroscopy is a powerful technique for characterizing biomolecules such as proteins and nucleic acids at atomic resolution. Increased magnetic field strengths drive progress in biomolecular NMR applications, leading to improved performance, e.g., higher resolution. A new class of NMR spectrometers with a 28.2 T magnetic field (1.2 GHz 1 H frequency) has been commercially available since the end of 2019. The availability of ultra-high-field NMR instrumentation makes it possible to investigate more complex systems using NMR. This is especially true for highly flexible intrinsically disordered proteins (IDPs) and highly flexible regions (IDRs) of complex multidomain proteins. Indeed, the investigation of these proteins is frequently hampered by the crowding of NMR spectra. The advantages, however, are accompanied by challenges that the user must overcome when conducting experiments at such a high field (e.g., large spectral widths, radio frequency bandwidth, performance of decoupling schemes). This protocol presents strategies and tricks for optimising high-field NMR experiments for IDPs/IDRs based on the analysis of the relaxation properties of the investigated protein. The protocol, tested on three IDPs of different molecular weight and structural complexity, focuses on 13 C-detected NMR at 1.2 GHz. A set of experiments, including some multiple receiver experiments, and tips to implement versions tailored for IDPs/IDRs are described. However, the general approach and most considerations can also be applied to experiments that acquire 1 H or 15 N nuclei and to experiments performed at lower field strengths., (© 2023. Springer Nature Limited.)

Context: Hepatitis B (HBV) and hepatitis C virus (HCV) possess well-known oncogenic properties and may promote carcinogenesis in liver. However antigens and replicative sequences of HBV/HCV have been also detected in different extra-hepatic tissues, including the pancreas. Although epidemiological studies and meta-analyses have recently suggested that HBV/HCV may be also risk factors for pancreatic cancer and several researches have investigated the possible mechanisms and intra-/extra-cellular paths involved in pancreatic and hepatic carcinogenesis, to date, these complex processes remain largely unexplained., Objectives: In our paper, we aimed to propose a comprehensive and qualitative hypothetical model, describing how HBV/HCV may exert their oncogenic role., Methods: We performed a systematic research of scientific literature, by searching MEDLINE, the Cochrane Library and EMBASE databases. The used keywords were: "chronic HBV/HCV", "pancreatic cancer", "liver carcinoma", "carcinogenesis mechanisms", "tensional integrity", "cytoskeleton", and "extracellular matrix"., Results: Taking advantage from available studies, we suggest an unifying hypothesis based on results and data, obtained from different areas of research. In particular we considered the well-defined model of tensional integrity and correlated it to changes induced by HBV/HCV in viscoelastic properties/stiffness of cellular/extracellular microenvironments. These events perturb the tightly-regulated feedback loop, which usually couples the intracellular-generated forces to substrate rigidity of extracellular compartments. Therefore, such a change strongly affects intracellular functions and cellular fate, by promoting a substantial deregulation of critical intracellular biochemical activities and genome expression., Conclusions: Our hypothesis might provide for the first time a reliable system, which correlates tensional integrity model with intra-/extra-cellular modifications, occurring in liver and pancreas during HBV/HCV-induced carcinogenesis. This approach might improve our understanding of pathogenetic mechanisms involved in the development of pancreatic and hepatic carcinogenesis , enhancing the possibility of their treatment. Furthermore, should the usefulness of this model be definitively confirmed, it might be also helpful to extend its field of application to other viruses-related cancers.

Background: Although the number of elderly people is progressively increasing in the world, old and very old patients have been under-represented and understudied in trials evaluating the efficacy of chronic illness management models. The usual hospital indicators and practice guidelines do not consider the effects of complexity - co-morbidity, social support, functional and cognitive status, patient adherence to therapy, risk of adverse drug reactions - in these subjects. The aim of this observational, multi-centric cohort study was to carefully assess factors contributing to the complexity of care for patients admitted to internal medicine wards. This was done by evaluating the severity of disease and degree of stability at admission, co-morbidity, age-related impairments, and the need for discharge planning plus post-discharge support., Methods: A total of 386 patients from 11 internal medicine wards in Emilia-Romagna and Marche, Italy, enrolled in a given week were evaluated. At admission, the following variables were recorded: demographic characteristics, medical history, global clinical-social prognostic evaluation, co-morbidity, severity of illness, presence of shock or hemodynamic instability, coma, and frequencies and causes of unscheduled hospital re-admission., Results: Cancer, congestive heart failure, pneumonia, stroke, and chronic obstructive pulmonary disease were the most frequent primary diagnoses. The complexity of our case study was characterized by several concomitant diseases. Over 50% of the patients were considered severe or more than severe, and over 20% extremely severe, with very high co-morbidity indices and illness severity scores. Some 55% of our patients were in need of partial or total care; 10% had some speech impairment, and 63% needed in-home health care after hospital discharge., Conclusions: The increasing numbers of elderly patients admitted to internal medicine departments suggests the need for a chronic illness management model, integrating gerontological and geriatric care to improve outcomes. For effective care, future protocols need to take a multi-dimensional, interdisciplinary approach to these patients and to develop a coordinated, integrated plan for treatment and long-term follow-up.

The SARS-CoV-2 nucleocapsid protein is indispensable for viral RNA genome processing. Although the N-terminal domain (NTD) is suggested to mediate specific RNA-interactions, high-resolution structures with viral RNA are still lacking. Available hybrid structures of the NTD with ssRNA and dsRNA provide valuable insights; however, the precise mechanism of complex formation remains elusive. Similarly, the molecular impact of nucleocapsid NTD mutations that have emerged since 2019 has not yet been fully explored. Using crystallography and solution NMR, we investigate how NTD mutations influence structural integrity and RNA-binding. We find that both features rely on a core network of residues conserved in Betacoronaviruses, crucial for protein stability and communication among flexible loop-regions that facilitate RNA-recognition. Our comprehensive structural analysis demonstrates that contacts within this network guide selective RNA-interactions. We propose that the core network renders the NTD evolutionarily robust in stability and plasticity for its versatile RNA processing roles. The authors use solution-state NMR and X-ray crystallography to study mutations in the SARS-CoV-2 Nucleocapsid NTD, uncovering a network essential for protein integrity and the selectively of RNA binding. [ABSTRACT FROM AUTHOR]

The nucleocapsid (N) protein of SARS-CoV-2 is a multifunctional protein involved in nucleocapsid assembly and various regulatory functions. It is the most abundant protein during viral infection. Its functionality is closely related to its structure, which comprises two globular domains, the N-terminal domain (NTD) and the C-terminal domain (CTD), flanked by intrinsically disordered regions. The linker between the NTD and CTD includes a Serine-Arginine rich (SR) region, which is crucial for the regulation of the N protein's function. Here, we report the near-complete assignment of the construct containing the NTD followed by the SR region (NTD-SR). Additionally, we describe the dynamic nature of the SR region and compare it with all other available chemical shift assignments reported for the SR region., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

The robustness of NMR coherence transfer in proximity of a paramagnetic center depends on the relaxation properties of the nuclei involved. In the case of Iron-Sulfur Proteins, different pulse schemes or different parameter sets often provide complementary results. Tailored versions of HCACO and CACO experiments significantly increase the number of observed Cα/C' connectivities in highly paramagnetic systems, by recovering many resonances that were lost due to paramagnetic relaxation. Optimized 13C direct detected experiments can significantly extend the available assignments, improving the overall knowledge of these systems. The different relaxation properties of Cα and C' nuclei are exploited in CACO vs COCA experiments and the complementarity of the two experiments is used to obtain structural information. The two [Fe2S2]+ clusters containing NEET protein CISD3 and the one [Fe4S4]2+ cluster containing HiPIP protein PioC have been taken as model systems. We show that tailored experiments contribute to decrease the blind sphere around the cluster, to extend resonance assignment of cluster bound cysteine residues and to retrieve details on the topology of the iron-bound ligand residues. [ABSTRACT FROM AUTHOR]

We learned that proteins with unique 3D structures can switch folds (referred to as "metamorphic" or shapeshifting proteins), and a new class of proteins, called intrinsically disordered proteins (IDPs), that lack structure but can either transition to order, or in many cases, be functional in the absence of any structure ("fuzzy" logic), was discovered. This paper showed that structured proteins and IDPs could be separated by a straight line on a plot of a protein's net charge ( I Y i -axis) versus its overall hydropathy ( I X i -axis) using the Kyte and Doolittle hydropathy scale. Until the late 1990s, we believed that protein function required a unique, well-defined 3D structure encrypted in the amino acid sequence. This collection is a small token of the respect, admiration, and affection the contributors have for Prof. Vladimir (Volodya) Uversky. [Extracted from the article]

In paramagnetic metalloproteins, longitudinal relaxation rates of 13C′ and 13Cα nuclei can be measured using 13C detected experiments and converted into electron spin-nuclear spin distance restraints, also known as Paramagnetic Relaxation Enhancement (PRE) restraints. 13C are less sensitive to paramagnetism than 1H nuclei, therefore, 13C based PREs constitute an additional, non-redundant, structural information. We will discuss the complementarity of 13C PRE restraints with 1H PRE restraints in the case of the High Potential Iron Sulfur Protein (HiPIP) PioC, for which the NMR structure of PioC has been already solved by a combination of classical and paramagnetism-based restraints. We will show here that 13C R1 values can be measured also at very short distances from the paramagnetic center and that the obtained set of 13C based restraints can be added to 1H PREs and to other classical and paramagnetism based NMR restraints to improve quality and quantity of the NMR information. [ABSTRACT FROM AUTHOR]

Coronavirus disease 2019 (COVID-19) continues to take a heavy toll on personal health, healthcare systems, and economies around the globe. Scientists are expending tremendous effort to develop diagnostic technologies for detecting positive infections within the shortest possible time, and vaccines and drugs specifically for the prevention and treatment of COVID-19 disease. At the same time, emerging novel variants have raised serious concerns about vaccine efficacy. The SARS-CoV-2 nucleocapsid (N) protein plays an important role in the coronavirus life cycle, and participates in various vital activities after virus invasion. It has attracted a large amount of attention for vaccine and drug development. Here, we summarize the latest research of the N protein, including its role in the SARS-CoV-2 life cycle, structure and function, and post-translational modifications in addition to its involvement in liquid–liquid phase separation (LLPS) and use as a basis for the development of vaccines and diagnostic techniques. [ABSTRACT FROM AUTHOR]

The dysbindin domain-containing protein 1 (DBNDD1) is a conserved protein among higher eukaryotes whose structure and function are poorly investigated so far. Here, we present the backbone and side chain nuclear magnetic resonance assignments for the human DBNDD1 protein. Our chemical-shift based secondary structure analysis reveals the human DBNDD1 as an intrinsically disordered protein. [ABSTRACT FROM AUTHOR]

Nickel exposure is associated with tumors of the respiratory tract such as lung and nasal cancers, acting through still-uncharacterized mechanisms. Understanding the molecular basis of nickel-induced carcinogenesis requires unraveling the mode and the effects of Ni(II) binding to its intracellular targets. A possible Ni(II)-binding protein and a potential focus for cancer treatment is hNDRG1, a protein induced by Ni(II) through the hypoxia response pathway, whose expression correlates with higher cancer aggressiveness and resistance to chemotherapy in lung tissue. The protein sequence contains a unique C-terminal sequence of 83 residues (hNDRG1*C), featuring a three-times-repeated decapeptide, involved in metal binding, lipid interaction and post-translational phosphorylation. In the present work, the biochemical and biophysical characterization of unmodified hNDRG1*C was performed. Bioinformatic analysis assigned it to the family of the intrinsically disordered regions and the absence of secondary and tertiary structure was experimentally proven by circular dichroism and NMR. Isothermal titration calorimetry revealed the occurrence of a Ni(II)-binding event with micromolar affinity. Detailed information on the Ni(II)-binding site and on the residues involved was obtained in an extensive NMR study, revealing an octahedral paramagnetic metal coordination that does not cause any major change of the protein backbone, which is coherent with CD analysis. hNDRG1*C was found in a monomeric form by light-scattering experiments, while the full-length hNDRG1 monomer was found in equilibrium between the dimer and tetramer, both in solution and in human cell lines. The results are the first essential step for understanding the cellular function of hNDRG1*C at the molecular level, with potential future applications to clarify its role and the role of Ni(II) in cancer development. [ABSTRACT FROM AUTHOR]

Severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) nucleocapsid protein is an essential structural component of mature virions, encapsulating the genomic RNA and modulating RNA transcription and replication. Several of its activities might be associated with the protein's ability to undergo liquid–liquid phase separation. NSARS‐CoV‐2 contains an intrinsically disordered region at its N‐terminus (NTE) that can be phosphorylated and is affected by mutations found in human COVID‐19 infections, including in the Omicron variant of concern. Here, we show that NTE deletion decreases the range of RNA concentrations that can induce phase separation of NSARS‐CoV‐2. In addition, deletion of the prion‐like NTE allows NSARS‐CoV‐2 droplets to retain their liquid‐like nature during incubation. We further demonstrate that RNA‐binding engages multiple parts of the NTE and changes NTE's structural properties. The results form the foundation to characterize the impact of N‐terminal mutations and post‐translational modifications on the molecular properties of the SARS‐CoV‐2 nucleocapsid protein. Statement: The nucleocapsid protein of SARS‐CoV‐2 plays an important role in both genome packaging and viral replication upon host infection. Replication has been associated with RNA‐induced liquid–liquid phase separation of the nucleocapsid protein. We present insights into the role of the N‐terminal part of the nucleocapsid protein in the protein's RNA‐mediated liquid–liquid phase separation. [ABSTRACT FROM AUTHOR]

Severe Acute Respiratory Syndrome Coronavirus Type 2 (SARS-CoV-2) emerged at the end of 2019, causing a highly infectious and pathogenic disease known as 2019 coronavirus disease. This disease poses a serious threat to human health and public safety. The SARS-CoV-2 main protease (M pro ) is a highly sought-after target for developing drugs against COVID-19 due to its exceptional specificity. Its crystal structure has been extensively documented. Numerous strategies have been employed in the investigation of M pro inhibitors. This paper is primarily concerned with Fragment-based Drug Discovery (FBDD), which has emerged as an effective approach to drug design in recent times. Here, we summarize the research on the approach of FBDD and its application in developing inhibitors for SARS-CoV-2 M pro ., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The current work investigated SARS‐CoV‐2 Nucleocapsid (NCAP or N protein) interactors in A549 human lung cancer cells using a SILAC‐based mass spectrometry approach. NCAP interactors included proteins of the stress granule (SG) machinery and immunoregulators. NCAP showed specific interaction with the SG proteins G3BP1, G3BP2, YTHDF3, USP10 and PKR, and translocated to SGs following oxidative stress and heat shock. Treatment of recombinant NCAP with RNA isolated from A549 cells exposed to oxidative stress‐stimulated NCAP to undergo liquid–liquid phase separation (LLPS). RNA degradation using RNase A treatment completely blocked the LLPS property of NCAP as well as its SG association. The RNA intercalator mitoxantrone also disrupted NCAP assembly in vitro and in cells. This study provides insight into the biological processes and biophysical properties of the SARS‐CoV‐2 NCAP. [ABSTRACT FROM AUTHOR]

The relationship between hepatitis B virus (HBV) and pancreatic cancer has been controversial for years, but more recently new information on this relationship has been updated Therefore, we performed a meta-analysis to provide summary estimates of the risk of pancreatic cancer associated with HBV infection. A systematic literature search on HBV and pancreatic cancer in English was performed in Pubmed, Cochrane library and Embase up to July 2020. Pooled rate ratios (RRs) and 95% confidence intervals (CIs) were calculated by the random-effects model. Stata software version 15.1 was used to perform this meta-analysis of the 17 studies considered to be eligible. 17 studies including 7 case-control and 10 cohort studies met the selection criteria. Begg's and Egger's test results indicated that there was no publication bias. Individuals with Hepatitis B surface antigen (HBsAg) or HBV DNA seropositivity had a significantly increased risk of pancreatic cancer showing an RR (95% CI) of 1.39 (1.19, 1.63). Similar conclusions were drawn from the results of the subgroup analysis (subgroup by study design, population, sex ratio) except when subgrouped by patient's region: the RR and 95% CI in Europe and Oceania were 1.44 (0.88, 2.34) and 1.47(0.38, 5.71) respectively. The findings of this meta-analysis suggest that HBV infections may increase the risk of pancreatic cancer under most conditions, while there remains some doubt when comparison is made between European and Oceania patients. [ABSTRACT FROM AUTHOR]

Pancreatic cancer is one of the most common digestive system cancers. Early diagnosis is difficult owing to the lack of specific symptoms and reliable biomarkers. The cause of pancreatic cancer remains ambiguous. Smoking, drinking, new-onset diabetes, and chronic pancreatitis have been proven to be associated with the occurrence of pancreatic cancer. In recent years, a large number of studies have clarified that a variety of microorganisms colonized in pancreatic cancer tissues are also closely related to the occurrence and development of pancreatic cancer, and the specific mechanisms include inflammatory induction, immune regulation, metabolism, and microenvironment changes caused by microorganism. The mechanism of action of the pancreatic colonized microbiome in the tumor microenvironment, as well as immunotherapy approaches require further study in order to find more evidence to explain the complex relationship between the pancreatic colonized microbiome and PDAC. Relevant studies targeting the microbiome may provide insight into the mechanisms of PDAC development and progression, improving treatment effectiveness and overall patient prognosis. In this article, we focus on the research relating to the microorganisms colonized in pancreatic cancer tissues, including viruses, bacteria, and fungi. We also highlight the microbial diversity in the occurrence, invasion, metastasis, treatment, and prognosis of pancreatic cancer in order to elucidate its significance in the early diagnosis and new therapeutic treatment of pancreatic cancer, which urgently need to be improved in clinical practice. The elimination or increase in diversity of the pancreatic microbiome is beneficial for prolonging the survival of PDAC patients, improving the response to chemotherapy drugs, and reducing tumor burden. The colonization of microorganisms in the pancreas may become a new hotspot in the diagnosis and treatment of pancreatic cancer. [ABSTRACT FROM AUTHOR]

NMR represents a key spectroscopic technique that contributes to the emerging field of highly flexible, intrinsically disordered proteins (IDPs) or protein regions (IDRs) that lack a stable three-dimensional structure. A set of exclusively heteronuclear NMR experiments tailored for proline residues, highly abundant in IDPs/IDRs, are presented here. They provide a valuable complement to the widely used approach based on amide proton detection, filling the gap introduced by the lack of amide protons in proline residues within polypeptide chains. The novel experiments have very interesting properties for the investigations of IDPs/IDRs of increasing complexity. [ABSTRACT FROM AUTHOR]

Tumors are complex networks of constantly interacting elements: tumor cells, stromal cells, immune and stem cells, blood/lympathic vessels, nerve fibers and extracellular matrix components. These elements can influence their microenvironment through mechanical and physical signals to promote tumor cell growth. To get a better understanding of tumor biology, cooperation between multidisciplinary fields is needed. Diverse mathematic computations and algorithms have been designed to find prognostic targets and enhance diagnostic assessment. In this work, we use computational digital tools to study the topology of vitronectin, a glycoprotein of the extracellular matrix. Vitronectin is linked to angiogenesis and migration, two processes closely related to tumor cell spread. Here, we investigate whether the distribution of this molecule in the tumor stroma may confer mechanical properties affecting neuroblastoma aggressiveness. Combining image analysis and graph theory, we analyze different topological features that capture the organizational cues of vitronectin in histopathological images taken from human samples. We find that the Euler number and the branching of territorial vitronectin, two topological features, could allow for a more precise pretreatment risk stratification to guide treatment strategies in neuroblastoma patients. A large amount of recently synthesized VN would create migration tracks, pinpointed by both topological features, for malignant neuroblasts, so that dramatic change in the extracellular matrix would increase tumor aggressiveness and worsen patient outcomes. What's new? The tumor microenvironment has a strong influence on cancer malignancy. Here, the authors investigate whether the organization of the extracellular matrix glycoprotein vitronectin in the tumor stroma may confer mechanical properties affecting neuroblastoma aggressiveness. Combining image analysis and graph theory, they identify two topological features of vitronectin that could potentially be used to improve patient pre‐treatment risk stratification. The data also point to the creation of vitronectin migration tracks for malignant neuroblasts, so that dramatic changes in the extracellular matrix would increase tumor stiffness and aggressiveness and worsen patient outcomes. [ABSTRACT FROM AUTHOR]

Microbiota is just beginning to be recognized as an important player in carcinogenesis and the interplay among microbes is greater than expected. Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease for which mortality closely parallels incidence. Early detection would provide the best opportunity to increase survival rates. Specific well-studied oral, gastrointestinal, and intrapancreatic microbes and some kinds of hepatotropic viruses and bactibilia may have potential etiological roles in pancreatic carcinogenesis, or modulating individual responses to oncotherapy. Concrete mechanisms mainly involve perpetuating inflammation, regulating the immune system-microbe-tumor axis, affecting metabolism, and altering the tumor microenvironment. The revolutionary technology of omics has generated insight into cancer microbiomes. A better understanding of the microbiota in PDAC might lead to the establishment of screening or early-stage diagnosis methods, implementation of cancer bacteriotherapy, adjustment of therapeutic efficacy even alleviating the adverse effects, creating new opportunities and fostering hope for desperate PDAC patients. [ABSTRACT FROM AUTHOR]

Today liver damage caused by Hepatitis C Virus (HCV) is widespread and included in the list of diseases of a social character. The aim of this research is to identify the peculiarities and summarize the epidemiological characteristics of HCV-infection in Ukraine, based on a retrospective analysis of detecting HCV based on antibodies, RNA and its genotyping, taking into account the sex and age of infected people in regions of Ukraine and particularly in Dniprо and Dnipropetrovs’k region. The research materials were provided by sampling the results of screening tests and qualitative detection of RNA-HCV genotyping (1a, 1b, 2 and 3). The survey involved 10,394 people from different regions of Ukraine (2014–2016). HCV-infection was detected in 3,436 of them, accounting on average for 32.5% of those surveyed. We analysed further 1,113 cases with positive RNA-HCV genotyping. Detection of infection by age periods was: 0–9 years old – 0.8%, 10–19 – 0.5%, 20–29 – 11%, 30–39 – 32%, 40–49 – 30%, 50–59 – 19.5%, 60–69 – 5.0% and 70–79 – 1.0%. 603 (54.2%) infected people belonged to genotype 1; 107 (9.6%) belonged to genotype 2; 403 (36.2%) belonged to genotype 3. Women were more frequently infected by genotype 1b, with increased percentage of 1ab; men often had genotype 3. 1,513 inhabitants of Dniprо and Dnipropetrovs’k region were sent to be tested for anti-HCV in 2011–2015. The percentage of positive anti-HCV showed a sharp increase from 28% to 64% over the observation period. Detection of HCV by age periods was: 0–9 years old – 0.9%, 10–19 – 1.5%, 20–29 – 9.5%, 30– 39 – 27.6%, 40–49 – 27.8%, 50–59 – 23.2%, 60–69 – 8.8% and 70–79 – 0.7%. 1 genotype was identified in 262 (56.8%) infected patients among 461 HCV-infected, 38 (8.2%) had genotype 2, 61 (35%) had genotype 3. 2.6% were diagnosed 1ab. In both populations that were tested, the frequency of detection of HCV was similar among men and women. Gender differences in the spread of genotype 2 haven’t been identified. The disease HCV remains one of the urgent problems of public health in Ukraine. [ABSTRACT FROM AUTHOR]

Increased stromal stiffness is associated with hepatocellular carcinoma (HCC) development and progression. However, the molecular mechanism by which matrix stiffness stimuli modulate HCC progress is largely unknown. In this study, we explored whether matrix stiffness-mediated effects on osteopontin (OPN) expression occur in HCC cells. We used a previously reported in vitro culture system with tunable matrix stiffness and found that OPN expression was remarkably upregulated in HCC cells with increasing matrix stiffness. Furthermore, the phosphorylation level of GSK3β and the expression of nuclear β-catenin were also elevated, indicating that GSK3β/β-catenin pathway might be involved in OPN regulation. Knock-down analysis of integrin β1 showed that OPN expression and p-GSK3β level were downregulated in HCC cells grown on high stiffness substrate compared with controls. Simultaneously, inhibition of GSK-3β led to accumulation of β-catenin in the cytoplasm and its enhanced nuclear translocation, further triggered the rescue of OPN expression, suggesting that the integrin β1/GSK-3β/β-catenin pathway is specifically activated for matrix stiffness-mediated OPN upregulation in HCC cells. Tissue microarray analysis confirmed that OPN expression was positively correlated with the expression of LOX and COL1. Taken together, high matrix stiffness upregulated OPN expression in HCC cells via the integrin β1/GSK-3β/β-catenin signaling pathway. It highlights a new insight into a pathway involving physical mechanical signal and biochemical signal molecules which contributes to OPN expression in HCC cells. [ABSTRACT FROM AUTHOR]

This section presents various orthopedic studies. One study considers arthroscopy as a valuable diagnostic tool and therapeutic technique in treatment process of reactive synovitis and arthrofibrosis. According to another study, suture-anchor arthroscopic technique has the capacity to identify intra-articular associated lesions. Another study concludes that overall Local Recurrence Rate (LRR) can be reduced by radiotherapy.

Objective: Evaluation of the interaction between alcohol intake and cofactors [hepatitis B virus (HBV), hepatitis C virus (HCV), body mass index] and coffee consumption on the risk of cirrhosis. Design: Seven hundred and forty-nine consecutive patients with chronic liver disease referring to units for liver or alcohol diseases in Italy during a 6-months period. Teetotalers were excluded. The odds ratios (OR) for cirrhosis were evaluated using chronic hepatitis cases as the control group. Results: An alcohol intake of more than 3 units/day resulted associated with the likelihood of cirrhosis both in males (OR 4.3; 95% CI=2.5–7.3) and in females (OR 5.7; 95% CI=2.3–14.5). A multiplicative interaction on the risk of cirrhosis between risky alcohol intake and HBsAg or HCV-Ab/HCV-RNA positivity was observed. A reduction of cirrhosis risk was observed in subjects consuming more than 3 alcohol units/day with increasing coffee intake. The OR for the association with cirrhosis decreased from 2.3 (95% CI=1.2–4.4) in subjects drinking 0–2 cups of coffee/day to 1.4 (95% CI=0.6–3.6) in those drinking more than 2 cups/day. Conclusions: In subjects with an alcohol intake >3 units/day the coexistence of HBV or HCV multiplies the risk of cirrhosis. Coffee represents a modulator of alcoholic cirrhosis risk. [ABSTRACT FROM AUTHOR]

The impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on the world is still expanding. Thus, there is an urgent need to better understand this novel virus and find a way to control its spread. Like other coronaviruses, the nucleocapsid (N) protein is one of the most crucial structural components of SARS-CoV-2. This protein shares 90% homology with the severe acute respiratory syndrome coronavirus N protein, implying functional significance. Based on the evolutionary conservation of the N protein in coronavirus, we reviewed the currently available knowledge regarding the SARS-CoV-2 N protein in terms of structure, biological functions, and clinical application as a drug target or vaccine candidate. [ABSTRACT FROM AUTHOR]

Using SAXS and NMR spectroscopy, we herein provide a high-resolution description of the intrinsically disordered N-terminal domain (PNT, aa 1–406) shared by the Nipah virus (NiV) phosphoprotein (P) and V protein, two key players in viral genome replication and in evasion of the host innate immune response, respectively. The use of multidimensional NMR spectroscopy allowed us to assign as much as 91% of the residues of this intrinsically disordered domain whose size constitutes a technical challenge for NMR studies. Chemical shifts and nuclear relaxation measurements provide the picture of a highly flexible protein. The combination of SAXS and NMR information enabled the description of the conformational ensemble of the protein in solution. The present results, beyond providing an overall description of the conformational behavior of this intrinsically disordered region, also constitute an asset for obtaining atomistic information in future interaction studies with viral and/or cellular partners. The present study can thus be regarded as the starting point towards the design of inhibitors that by targeting crucial protein–protein interactions involving PNT might be instrumental to combat this deadly virus. [ABSTRACT FROM AUTHOR]

Background: A hallmark of pancreatic ductal adenocarcinoma is the desmoplastic reaction, but its impact on the tumor behavior remains controversial. Our aim was to introduce a computer -aided method to precisely quantify the amount of pancreatic collagenic extra-cellular matrix, its spatial distribution pattern, and the degradation process.Methods: A series of normal, inflammatory and neoplastic pancreatic ductal adenocarcinoma formalin-fixed and paraffin-embedded Sirius red stained sections were automatically digitized and analyzed using a computer-aided method.Results: We found a progressive increase of pancreatic collagenic extra-cellular matrix from normal to the inflammatory and pancreatic ductal adenocarcinoma. The two-dimensional fractal dimension showed a significant difference in the collagenic extra-cellular matrix spatial complexity between normal versus inflammatory and pancreatic ductal adenocarcinoma. A significant difference when comparing the number of cycles necessary to degrade the pancreatic collagenic extra-cellular matrix in normal versus inflammatory and pancreatic ductal adenocarcinoma was also found. The difference between inflammatory and pancreatic ductal adenocarcinoma was also significant. Furthermore, the mean velocity of collagenic extra-cellular matrix degradation was found to be faster in inflammatory and pancreatic ductal adenocarcinoma than in normal.Conclusion: These findings demonstrate that inflammatory and pancreatic ductal adenocarcinomas are characterized by an increased amount of pancreatic collagenic extra-cellular matrix and by changes in their spatial complexity and degradation. Our study defines new features about the pancreatic collagenic extra-cellular matrix, and represents a basis for further investigations into the clinical behavior of pancreatic ductal adenocarcinoma and the development of therapeutic strategies. [ABSTRACT FROM AUTHOR]