Sofosbuvir Selects for Drug-Resistant Amino Acid Variants in the Zika Virus RNA-Dependent RNA-Polymerase Complex In Vitro

The nucleotide analog sofosbuvir, licensed for the treatment of hepatitis C, recently revealed activity against the Zika virus (ZIKV) in vitro and in animal models. However, the ZIKV genetic barrier to sofosbuvir has not yet been characterized. In this study, in vitro selection experiments were performed in infected human hepatoma cell lines. Increasing drug pressure significantly delayed viral breakthrough (p = 0.029). A double mutant in the NS5 gene (V360L/V607I) emerged in 3 independent experiments at 40–80 µM sofosbuvir resulting in a 3.9 ± 0.9-fold half- maximal inhibitory concentration (IC₅₀) shift with respect to the wild type (WT) virus. A triple mutant (C269Y/V360L/V607I), detected in one experiment at 80 µM, conferred a 6.8-fold IC₅₀ shift with respect to the WT. Molecular dynamics simulations confirmed that the double mutant V360L/V607I impacts the binding mode of sofosbuvir, supporting its role in sofosbuvir resistance. Due to the distance from the catalytic site and to the lack of reliable structural data, the contribution of C269Y was not investigated in silico. By a combination of sequence analysis, phenotypic susceptibility testing, and molecular modeling, we characterized a double ZIKV NS5 mutant with decreased sofosbuvir susceptibility. These data add important information to the profile of sofosbuvir as a possible lead for anti-ZIKV drug development.     

Performance evaluation of a region growing procedure for mammographic breast lesion identification

At present, mammography is the most effective examination for an early diagnosis of breast cancer. Nevertheless, the detection of cancer signs in mammograms is a difficult procedure owing to the great number of non-pathological structures which are also present in the image. Recent statistics show that in current breast cancer screenings 10%-25% of the tumors are missed by the radiologists. For this reason, a lot of research is currently being done to develop systems for Computer Aided Detection (CADe). Probably, some causes of the false-negative screening examinations are that tumoral masses have varying dimension and irregular shape, their borders are often ill-defined and their contrast is very low, thus making difficult the discrimination from parenchymal structures. Therefore, in a CADe system a preliminary segmentation procedure has to be implemented in order to separate the mass from the background tissue. In this way, various characteristics of the segmented mass can be evaluated and used in a classification step to discriminate benign and malignant cases. In this paper, we describe an effective algorithm for massive lesions segmentation based on a region-growing technique and we provide full details the performance evaluation procedure used in this specific context.     

Integrating web service and semantic dialogue model for user models interoperability on the web

Nowadays there is a great number of Web information systems that build a model of the user and adapt their services according to the needs and preferences maintained by the user model (UM). One of the most challenging issues of this scenario is the possibility to enable different systems to cooperate in order to exchange the available information about a user. Our aim is to create rich (and scalable) communication protocols and infrastructures to enable consumers and providers of UM data to interact. Our solution for dealing with such an issue is to exploit Web standards for interoperability (i.e. Semantic Web and Web Services) for implementing simple atomic communication, and a dialogue model for implementing enhanced communication capabilities. In particular, two systems can start a semantics-enhanced Dialogue Game as a form of negotiation to clarify the meaning of the requested concepts when a shared knowledge model does not exist, and to approximate the response when the exact one is not available. We propose a distributed semantic conversation framework based on the Sesame semantic environment for the exchange of user model knowledge on the Web. Systems have to expose their user model data as a Web Service, and to exploit a public dialogue knowledge base to start the dialogue. The main advantage of the approach is to allow systems to deal with difficult situations by starting an appropriate dialogue game instead of stopping the communication as in the traditional “all-or-nothing” Web Service approach. On the basis of a preliminary evaluation, the approach has shown an improvement of the adaptation results provided by the systems we tested.     

Bio-PEPA: A framework for the modelling and analysis of biological systems

In this work we present Bio-PEPA, a process algebra for the modelling and the analysis of biochemical networks. It is a modification of PEPA, originally defined for the performance analysis of computer systems, in order to handle some features of biological models, such as stoichiometry and the use of general kinetic laws. Bio-PEPA may be seen as an intermediate, formal, compositional representation of biological systems, on which different kinds of analyses can be carried out. Bio-PEPA is enriched with some notions of equivalence. Specifically, the isomorphism and strong bisimulation for PEPA have been considered and extended to our language. Finally, we show the translation of a biological model into the new language and we report some analysis results.     

High-Energy Physics on the Grid: the ATLAS and CMS Experience

In this paper we present the experience of the ATLAS and CMS High-Energy Physics (HEP) experiments at the Large Hadron Collider (LHC) with the LCG/EGEE Grid infrastructure. The activity developed around the following two main lines: large-scale physics and detector simulations and end-user analysis. The LCG/EGEE Grid infrastructure offers a large amount of computing and storage resources and is growing very rapidly. It provides the natural environment for large-scale physics and detector simulations. Also, the analysis of these detector simulation data (and in the near future of the reconstructed data from physics collisions) requires efficient end-users access to Grid resources. In this paper, the main findings and lessons learned in terms of performance, robustness and scalability of the whole system are discussed in detail.     

Metabolic Syndrome and Male Fertility: Beyond Heart Consequences of a Complex Cardiometabolic Endocrinopathy

Metabolic syndrome (MetS) is a highly prevalent condition among adult males, affecting up to 41% of men in Europe. It is characterized by the association of obesity, hypertension, and atherogenic dyslipidemia, which lead to premature morbidity and mortality due to cardiovascular disease (CVD). Male infertility is another common condition which accounts for about 50% of cases of couple infertility worldwide. Interestingly, male infertility and MetS shares several risk factors (e.g., smoking, ageing, physical inactivity, and excessive alcohol consumption), leading to reactive oxygen species (ROS) production and increased oxidative stress (OS), and resulting in endothelial dysfunction and altered semen quality. Thus, the present narrative review aims to discuss the pathophysiological mechanisms which link male infertility and MetS and to investigate the latest available evidence on the reproductive consequences of MetS.     

Engineered Sleeping Beauty Transposon as Efficient System to Optimize Chimp Adenoviral Production

Sleeping Beauty (SB) is the first DNA transposon employed for efficient transposition in vertebrate cells, opening new applications for genetic engineering and gene therapies. A transposon-based gene delivery system holds the favourable features of non-viral vectors and an attractive safety profile. Here, we employed SB to engineer HEK293 cells for optimizing the production of a chimpanzee Adenovector (chAd) belonging to the Human Mastadenovirus C species. To date, chAd vectors are employed in several clinical settings for infectious diseases, last but not least COVID-19. A robust, efficient and quick viral vector production could advance the clinical application of chAd vectors. To this aim, we firstly swapped the hAd5 E1 with chAd-C E1 gene by using the CRISPR/Cas9 system. We demonstrated that in the absence of human Ad5 E1, chimp Ad-C E1 gene did not support HEK293 survival. To improve chAd-C vector production, we engineered HEK293 cells to stably express the chAd-C precursor terminal protein (ch.pTP), which plays a crucial role in chimpanzee Adenoviral DNA replication. The results indicate that exogenous ch.pTP expression significantly ameliorate the packaging and amplification of recombinant chAd-C vectors thus, the engineered HEK293ch.pTP cells could represent a superior packaging cell line for the production of these vectors.     

Chronic Hyperkaliemia in Chronic Kidney Disease: An Old Concern with New Answers

Increasing potassium intake ameliorates blood pressure (BP) and cardiovascular (CV) prognoses in the general population; therefore the World Health Organization recommends a high-potassium diet (90–120 mEq/day). Hyperkalaemia is a rare condition in healthy individuals due to the ability of the kidneys to effectively excrete dietary potassium load in urine, while an increase in serum K⁺ is prevalent in patients with chronic kidney disease (CKD). Hyperkalaemia prevalence increases in more advanced CKD stages, and is associated with a poor prognosis. This scenario generates controversy on the correct nutritional approach to hyperkalaemia in CKD patients, considering the unproven link between potassium intake and serum K⁺ levels. Another concern is that drug-induced hyperkalaemia leads to the down-titration or withdrawal of renin-angiotensin system inhibitors (RASI) and mineralocorticoids receptors antagonists (MRA) in patients with CKD, depriving these patients of central therapeutic interventions aimed at delaying CKD progression and decreasing CV mortality. The new K⁺-binder drugs (Patiromer and Sodium-Zirconium Cyclosilicate) have proven to be adequate and safe therapeutic options to control serum K⁺ in CKD patients, enabling RASI and MRA therapy, and possibly, a more liberal intake of fruit and vegetables.