MARS: A Multipurpose Software for Untargeted LC-MS-Based Metabolomics and Exposomics.

Untargeted metabolomics is a growing field, in which recent advances in high-resolution mass spectrometry coupled with liquid chromatography (LC-MS) have facilitated untargeted approaches as a result of improvements in sensitivity, mass accuracy, and resolving power. However, a very large amount of data are generated. Consequently, using computational tools is now mandatory for the in-depth analysis of untargeted metabolomics data. This article describes MetAbolomics ReSearch (MARS), an all-in-one vendor-agnostic graphical user interface-based software applying LC-MS analysis to untargeted metabolomics. All of the analytical steps are described (from instrument data conversion and processing to statistical analysis, annotation/identification, quantification, and preliminary biological interpretation), and tools developed to improve annotation accuracy (e.g., multiple adducts and in-source fragmentation detection, trends across samples, and the MS/MS validator) are highlighted. In addition, MARS allows in-house building of reference databases, to bypass the limits of freely available MS/MS spectra collections. Focusing on the flexibility of the software and its user-friendliness, which are two important features in multipurpose software, MARS could provide new perspectives in untargeted metabolomics data analysis.

Discovery and Structure-Activity Relationships of Novel ssDAF-12 Receptor Modulators.

The nuclear receptor ssDAF-12 has been recognized as the key molecular player regulating the life cycle of the nematode parasite Strongyloides stercoralis. ssDAF-12 ligands permit the receptor to function as an on/off switch modulating infection, making it vulnerable to therapeutic intervention. In this study, we report the design and synthesis of a set of novel dafachronic acid derivatives, which were used to outline the first structure-activity relationship targeting the ssDAF-12 receptor and to unveil hidden properties shared by the molecular shape of steroidal ligands that are relevant to the receptor binding and modulation. Moreover, biological results led to the discovery of sulfonamide 3 as a submicromolar ssDAF-12 agonist endowed with a high receptor selectivity, no toxicity, and improved properties, as well as to the identification of unprecedented ssDAF-12 antagonists that can be exploited in the search for novel chemical tools and alternative therapeutic approaches for treating parasitism such as Strongyloidiasis.

VHL-Modified PROteolysis TArgeting Chimeras (PROTACs) as a Strategy to Evade Metabolic Degradation in In Vitro Applications.

PROteolysis TArgeting Chimeras (PROTACs) are tripartite molecules consisting of a linker connecting a ligand for a protein of interest to an E3 ligase recruiter, whose rationale relies on proteasome-based protein degradation. PROTACs have expanded as a therapeutic strategy to open new avenues for unmet medical needs. Leveraging our expertise, we undertook a series of in vitro experiments aimed at elucidating PROTAC metabolism. In particular, we focused on PROTACs recruiting the von Hippel-Lindau (VHL) E3 ligase. After high-resolution mass spectrometry measurements, a characteristic metabolite with mass reduction of 200 units was detected and successively confirmed as a product deriving from the cleavage of the VHL ligand moiety. Subsequently, we identified hepatic and extrahepatic prolyl endopeptidases as the main putative metabolic enzymes involved. Finally, we designed and synthesized analogs of the VHL ligands that we further exploited for the synthesis of novel VHL-directed PROTACs with an improved metabolic stability in in vitro applications.

Effective and Selective Extraction of Quercetin from Onion (Allium cepa L.) Skin Waste Using Water Dilutions of Acid-Based Deep Eutectic Solvents.

Deep Eutectic Solvents (DESs) are experiencing growing interest as substitutes of polluting organic solvents for their low or absent toxicity and volatility. Moreover, they can be formed with natural bioavailable and biodegradable molecules; they are synthesized in absence of hazardous solvents. DESs are, inter alia, successfully used for the extraction/preconcentration of biofunctional molecules from complex vegetal matrices. Onion skin is a highly abundant waste material which represents a reservoir of molecules endowed with valuable biological properties such as quercetin and its glycosylated forms. An efficient extraction of these molecules from dry onion skin from "Dorata di Parma" cultivar was obtained with water dilution of acid-based DESs. Glycolic acid (with betaine 2/1 molar ratio and L-Proline 3/1 molar ratio as counterparts) and of p-toluensulphonic acid (with benzyltrimethylammonium methanesulfonate 1/1 molar ratio)-based DESs exhibited more than 3-fold higher extraction efficiency than methanol (14.79 µg/mL, 18.56 µg/mL, 14.83 µg/mL vs. 5.84 µg/mL, respectively). The extracted quercetin was also recovered efficaciously (81% of recovery) from the original extraction mixture. The proposed extraction protocol revealed to be green, efficacious and selective for the extraction of quercetin from onion skin and it could be useful for the development of other extraction procedures from other biological matrixes.

Synthesis and characterization of 1,2,4-triazolo[1,5-a]pyrimidine-2-carboxamide-based compounds targeting the PA-PB1 interface of influenza A virus polymerase.

Influenza viruses (Flu) are responsible for seasonal epidemics causing high rates of morbidity, which can dramatically increase during severe pandemic outbreaks. Antiviral drugs are an indispensable weapon to treat infected people and reduce the impact on human health, nevertheless anti-Flu armamentarium still remains inadequate. In search for new anti-Flu drugs, our group has focused on viral RNA-dependent RNA polymerase (RdRP) developing disruptors of PA-PB1 subunits interface with the best compounds characterized by cycloheptathiophene-3-carboxamide and 1,2,4-triazolo[1,5-a]pyrimidine-2-carboxamide scaffolds. By merging these moieties, two very interesting hybrid compounds were recently identified, starting from which, in this paper, a series of analogues were designed and synthesized. In particular, a thorough exploration of the cycloheptathiophene-3-carboxamide moiety led to acquire important SAR insight and identify new active compounds showing both the ability to inhibit PA-PB1 interaction and viral replication in the micromolar range and at non-toxic concentrations. For few compounds, the ability to efficiently inhibit PA-PB1 subunits interaction did not translate into anti-Flu activity. Chemical/physical properties were investigated for a couple of compounds suggesting that the low solubility of compound 14, due to a strong crystal lattice, may have impaired its antiviral activity. Finally, computational studies performed on compound 23, in which the phenyl ring suitably replaced the cycloheptathiophene, suggested that, in addition to hydrophobic interactions, H-bonds enhanced its binding within the PAC cavity.