Structural Investigations on Novel Non-Nucleoside Inhibitors of Human Norovirus Polymerase

Human norovirus is the first cause of foodborne disease worldwide, leading to extensive outbreaks of acute gastroenteritis, and causing around 200,000 children to die annually in developing countries. No specific vaccines or antiviral agents are currently available, with therapeutic options limited to supportive care to prevent dehydration. The infection can become severe and lead to life-threatening complications in young children, the elderly and immunocompromised individuals, leading to a clear need for antiviral agents, to be used as treatments and as prophylactic measures in case of outbreaks. Due to the key role played by the viral RNA-dependent RNA polymerase (RdRp) in the virus life cycle, this enzyme is a promising target for antiviral drug discovery. In previous studies, following in silico investigations, we identified different small-molecule inhibitors of this enzyme. In this study, we rationally modified five identified scaffolds, to further explore structure–activity relationships, and to enhance binding to the RdRp. The newly designed compounds were synthesized according to multiple-step synthetic routes and evaluated for their inhibition of the enzyme in vitro. New inhibitors with low micromolar inhibitory activity of the RdRp were identified, which provide a promising basis for further hit-to-lead optimization

Multi-Criteria Decision Analysis (MCDA) for sustainability assessment in food sector. A systematic literature review on methods, indicators and tools

In the last decades, several studies have highlighted the significant impacts of the food sector. Therefore, enhancing sustainability within this sector has become of paramount importance. A crucial step towards achieving this goal involves the definition and implementation of effective sustainability metric and measurements. In this regard, the adoption of multi-criteria decision analysis (MCDA) methods can be seen as one of the most suitable and promising approach to comprehensively capture the complex and broad-ranging effects of agricultural practices and food supply chains. In such context, a systematic review of the scientific literature on multi-criteria approaches and tools for measuring the sustainability of food supply chains (harvest and post-harvest stages) has been carried out, resulting in the selection and analysis of 42 articles. To delve into the selected articles, three main areas of focus have been identified. The first about MCDA methods and their features, revealing the most adopted methods for sustainability assessments of food supply chains. The second, focusing on the participatory approach, led to the definition of a stakeholder's engagement map, highlighting the typology of stakeholders involved, the reasons of their involvement and engagement methods. Lastly, the third focus is related to the analysis and classification of indicators adopted in each study and the sustainability dimensions to which they refer to. The results of the present review study provide a comprehensive overview of the essential aspects to be considered when developing a MCDA for sustainability assessment in the food sector, serving as a valuable resource for both scholars and practitioners

A particular anatomic variation of the course of the left pulmonary artery

n/

Targeting the Complement Serine Protease MASP-2 as a Therapeutic Strategy for Coronavirus Infections

MASP-2, mannose-binding protein-associated serine protease 2, is a key enzyme in the lectin pathway of complement activation. Hyperactivation of this protein by human coronaviruses SARS-CoV, MERS-CoV and SARS-CoV-2 has been found to contribute to aberrant complement activation in patients, leading to aggravated lung injury with potentially fatal consequences. This hyperactivation is triggered in the lungs through a conserved, direct interaction between MASP-2 and coronavirus nucleocapsid (N) proteins. Blocking this interaction with monoclonal antibodies and interfering directly with the catalytic activity of MASP-2, have been found to alleviate coronavirus-induced lung injury both in vitro and in vivo. In this study, a virtual library of 8736 licensed drugs and clinical agents has been screened in silico according to two parallel strategies. The first strategy aims at identifying direct inhibitors of MASP-2 catalytic activity, while the second strategy focusses on finding protein-protein interaction inhibitors (PPIs) of MASP-2 and coronaviral N proteins. Such agents could represent promising support treatment options to prevent lung injury and reduce mortality rates of infections caused by both present and future-emerging coronaviruses. Forty-six drug repurposing candidates were purchased and, for the ones selected as potential direct inhibitors of MASP-2, a preliminary in vitro assay was conducted to assess their interference with the lectin pathway of complement activation. Some of the tested agents displayed a dose-response inhibitory activity of the lectin pathway, potentially providing the basis for a viable support strategy to prevent the severe complications of coronavirus infections

Structure–Activity Relationship Studies on Novel Antiviral Agents for Norovirus Infections

Human norovirus is the leading cause of acute gastroenteritis worldwide, affecting every year 685 million people. Norovirus outbreaks are associated with very significant economic losses, with an estimated societal cost of 60 billion USD per year. Despite this, no therapeutic options or vaccines are currently available to treat or prevent this infection. An antiviral therapy that can be used as treatment and as a prophylactic measure in the case of outbreaks is urgently needed. We previously described the computer-aided design and synthesis of novel small-molecule agents able to inhibit the replication of human norovirus in cell-based systems. These compounds are non-nucleoside inhibitors of the viral polymerase and are characterized by a terminal para-substituted phenyl group connected to a central phenyl ring by an amide-thioamide linker, and a terminal thiophene ring. Here we describe new modifications of these scaffolds focused on exploring the role of the substituent at the para position of the terminal phenyl ring and on removing the thioamide portion of the amide-thioamide linker, to further explore structure-activity relationships (SARs) and improve antiviral properties. According to three to four-step synthetic routes, we prepared thirty novel compounds, which were then evaluated against the replication of both murine (MNV) and human (HuNoV) norovirus in cells. Derivatives in which the terminal phenyl group has been replaced by an unsubstituted benzoxazole or indole, and the thioamide component of the amide-thioamide linker has been removed, showed promising results in inhibiting HuNoV replication at low micromolar concentrations. Particularly, compound 28 was found to have an EC50 against HuNoV of 0.9 µM. Although the most active novel derivatives were also associated with an increased cytotoxicity in the human cell line, these compounds represent a very promising starting point for the development of new analogues with reduced cytotoxicity and improved selectivity indexes. In addition, the experimental biological data have been used to create an initial 3D quantitative structure-activity relationship model, which could be used to guide the future design of novel potential anti-norovirus agents

Therapeutic homology-independent targeted integration in retina and liver

Challenges to the widespread application of gene therapy with adeno-associated viral (AAV) vectors include dominant conditions due to gain-of-function mutations which require allele-specific knockout, as well as long-term transgene expression from proliferating tissues, which is hampered by AAV DNA episomal status. To overcome these challenges, we used CRISPR/Cas9-mediated homology-independent targeted integration (HITI) in retina and liver as paradigmatic target tissues. We show that AAV-HITI targets photoreceptors of both mouse and pig retina, and this results in significant improvements to retinal morphology and function in mice with autosomal dominant retinitis pigmentosa. In addition, we show that neonatal systemic AAV-HITI delivery achieves stable liver transgene expression and phenotypic improvement in a mouse model of a severe lysosomal storage disease. We also show that HITI applications predominantly result in on-target editing. These results lay the groundwork for the application of AAV-HITI for the treatment of diseases affecting various organs

Energy retrofit of rural protected buildings. The case of a new agri-food hub in a peripheral context in Milan.

The research origins in the framework of “UIA OpenAgri”, a European project of urban regeneration. The present research focuses on the renovation of two historical buildings within Cascina Nosedo (Milan). First aim of our study is to support and address the energy retrofit project of the buildings toward energy efficiency and integration of the local renewable potential, in the respect of the constraints related to the historical value of the buildings. In this framework, after a preliminary analysis of the actual conservation state, two envelope scenarios, with internal and external insulation, were developed, simulated with IESVE tool and evaluated. As main results we obtained that, for the demand side there are not significant difference in terms of energy loads on yearly basis (sum of heating and cooling loads) in the two retrofit scenarios. Several considerations on the supply side were carried out to assess the local renewable potential

ROPfuscator: Robust Obfuscation with ROP

Software obfuscation is crucial in protecting intellectual property in software from reverse engineering attempts. While some obfuscation techniques originate from the obfuscation-reverse engineering arms race, others stem from different research areas, such as binary software exploitation. Return-oriented programming (ROP) became one of the most effective exploitation techniques for memory error vulnerabilities. ROP interferes with our natural perception of a process control flow, inspiring us to repurpose ROP as a robust and effective form of software obfuscation. Although previous work already explores ROP’s effectiveness as an obfuscation technique, evolving reverse engineering research raises the need for principled reasoning to understand the strengths and limitations of ROP-based mechanisms against man-at-the-end (MATE) attacks. To this end, we present ROPFuscator, a compiler-driven obfuscation pass based on ROP for any programming language supported by LLVM. We incorporate opaque predicates and constants and a novel instruction hiding technique to withstand sophisticated MATE attacks. More importantly, we introduce a realistic and unified threat model to thoroughly evaluate ROPFuscator and provide principled reasoning on ROP-based obfuscation techniques that answer to code coverage, incurred overhead, correctness, robustness, and practicality challenges. The project’s source code is published online to aid further research

An innovative application of 5GDHC: A techno-economic assessment of shallow geothermal systems potential in different European climates

Energy consumption for thermal purposes represents the most impacting energy issue in the European building sector. In addition to space heating, space cooling is constantly growing, due also to climate change, which provokes extreme hot events in summer even in moderate climates. To approach this challenge, it is essential to invest in lowering the overall energy demand, to increase the energy conversion efficiencies and to replace fossil fuels with renewable energy sources. The research here presented deals with the European decarbonization goals and focuses on shallow geothermal technology in district thermal systems (DTS), i.e. Geo-5GDHC. The research investigates whether Geo-5GDHC can be cost-effective in different scenarios based on climatic contexts, insulation levels and the possible integration of photovoltaic and thermal technology (PVT). Through the elaboration of proper KPIs and the implementation and use of a tool specifically developed to couple a Geo-5GDHC energy assessment model with an economic analysis (PILEDHC), the research highlights results and guidelines for providing solutions for new and innovative DTS, considering both energy and economic aspects in different contexts