Identification of FasL as a crucial host factor driving COVID-19 pathology and lethality.

The dysregulated immune response and inflammation resulting in severe COVID-19 are still incompletely understood. Having recently determined that aberrant death-ligand-induced cell death can cause lethal inflammation, we hypothesized that this process might also cause or contribute to inflammatory disease and lung failure following SARS-CoV-2 infection. To test this hypothesis, we developed a novel mouse-adapted SARS-CoV-2 model (MA20) that recapitulates key pathological features of COVID-19. Concomitantly with occurrence of cell death and inflammation, FasL expression was significantly increased on inflammatory monocytic macrophages and NK cells in the lungs of MA20-infected mice. Importantly, therapeutic FasL inhibition markedly increased survival of both, young and old MA20-infected mice coincident with substantially reduced cell death and inflammation in their lungs. Intriguingly, FasL was also increased in the bronchoalveolar lavage fluid of critically-ill COVID-19 patients. Together, these results identify FasL as a crucial host factor driving the immuno-pathology that underlies COVID-19 severity and lethality, and imply that patients with severe COVID-19 may significantly benefit from therapeutic inhibition of FasL.

A Photochemical Strategy for the Conversion of Nitroarenes into Rigidified Pyrrolidine Analogues.

Bicyclic amines are important motifs for the preparation of bioactive materials. These species have well-defined exit vectors that enable accurate disposition of substituents toward specific areas of chemical space. Of all possible skeletons, the 2-azabicyclo[3.2.0]heptane framework is virtually absent from MedChem libraries due to a paucity of synthetic methods for its preparation. Here, we report a modular synthetic strategy that utilizes nitroarenes as flat and easy-to-functionalize feedstocks for the assembly of these sp3-rich materials. Mechanistically, this approach exploits two concomitant photochemical processes that sequentially ring-expand the nitroarene into an azepine and then fold it into a rigid bicycle pyrroline by means of singlet nitrene-mediated nitrogen insertion and excited-state-4π electrocyclization. A following hydrogenolysis provides, with full diastereocontrol, the desired bicyclic amine derivatives whereby the aromatic substitution pattern has been translated into the one of the three-dimensional heterocycle. These molecules can be considered rigid pyrrolidine analogues with a well-defined orientation of their substituents. Furthermore, unsupervised clustering of an expansive virtual database of saturated N-heterocycles revealed these derivatives as effective isosteres of rigidified piperidines. Overall, this platform enables the conversion of nitroarene feedstocks into complex sp3-rich heterocycles of potential interest to drug development.

Dynamic Kinetic Resolution of Indole-Based Sulfenylated Heterobiaryls by Rhodium-Catalyzed Atroposelective Reductive Aldol Reaction.

A highly enantio- and diastereoselective dynamic kinetic resolution (DKR) of configurationally labile 3-aryl indole-2-carbaldehydes is described. The DKR proceeds via a Rh-catalyzed intermolecular asymmetric reductive aldol reaction with acrylate esters, with simultaneous generation of three stereogenic elements. The strategy relies on the labilization of the stereogenic axis that takes place thanks to a transient Lewis acid-base interaction (LABI) between the formyl group and a thioether moiety strategically located at the ortho' position. The atropisomeric indole products present a high degree of functionalization and can be further converted to a series of axially chiral derivatives, thereby expanding their potential application in drug discovery and asymmetric catalysis.

N-Terminal Cysteine Bioconjugation with (2-Cyanamidophenyl)boronic Acids Enables the Direct Formation of Benzodiazaborines on Peptides.

Benzodiazaborines (BDABs) have emerged as a valuable tool to produce stable and functional bioconjugates via a click-type transformation. However, the current available methods to install them on peptides lack bioorthogonality, limiting their applications. Here, we report a strategy to install BDABs directly on peptide chains using (2-cyanamidophenyl)boronic acids (2CyPBAs). The resulting BDAB is stabilized through the formation of a key intramolecular B-N bond. This technology was applied in the selective modification of N-terminal cysteine-containing functional peptides.

Computational studies of Brønsted acid-catalyzed transannular cycloadditions of cycloalkenone hydrazones.

The contribution to the energy barrier of a series of tethers in transannular cycloadditions of cycloalkenes with hydrazones has been computationally studied by using DFT. The Houk's distortion model has been employed to evaluate the influence of the tether in the cycloaddition reaction. That model has been extended to determine the contribution of each tether and, more importantly, the effect exerted between them. In addition to the distortion induced by the tethers, the entropy effects caused by them has also been studied. The analysis of the evolution of the electron localization function along the reaction revealed the highly concerted character of the reaction.

Organocatalytic Enantioselective Vinylcyclopropane-Cyclopentene (VCP-CP) Rearrangement.

We have demonstrated that the catalytic and enantioselective vinylcyclopropane-cyclopentene rearrangement can be carried out on (vinylcyclopropyl)acetaldehydes through activation via enamine intermediates. The reaction makes use of racemic starting materials that, upon ring opening facilitated by the catalytic generation of a donor-acceptor cyclopropane, deliver an acyclic iminium ion/dienolate intermediate in which all stereochemical information has been deleted. The final cyclization step forms the rearrangement product, showing that chirality transfer from the catalyst to the final compound is highly effective and leads to the stereocontrolled formation of a variety of structurally different cyclopentenes.

Molecular Recognition of GalNAc in Mucin-Type O-Glycosylation.

Acetylgalactosamine (GalNAc)-type O-glycosylation is an essential posttranslational modification (PTM) that plays fundamental roles in biology. Malfunction of this PTM is exemplified by the presence of truncated O-glycans in cancer. For instance, the glycoprotein MUC1 is overexpressed in many tumor tissues and tends to carry simple oligosaccharides that allow for the presentation of different tumor-associated antigens, such as the Tn or sTn antigens (GalNAc-α-1-O-Thr/Ser and Neu5Acα2-6GalNAcα1-O-Ser/Thr, respectively). In other cases, such as tumoral calcinosis associated with O-glycosylation of the fibroblast growth factor 23, O-glycans are absent or less abundant. Significant progress has been made in determining the three-dimensional structures of biomolecules that recognize GalNAc, such as antibodies, lectins, mucinases, GalNAc-transferases, and other glycosyltransferases. Analysis of the complexes between these entities and GalNAc-containing glycopeptides, in most cases derived from crystallographic or NMR analysis, provides an understanding of the key structural elements that control molecular recognition of these glycopeptides. Here, we describe and compare the binding sites of these proteins in detail, focusing on how the GalNAc moieties interact selectively with them. We also summarize the differences and similarities in GalNAc recognition. In general, the recognition of GalNAc-containing glycopeptides is determined by hydrogen bonds between hydroxyl groups and the N-acetyl group of GalNAc with proteins, as well as CH-π contacts in which the hydrophobic α-face of the sugar and the methyl group of NHAc can be involved. The latter interaction usually provides the basis for selectivity. It is worth noting that binding of these glycopeptides depends primarily on recognition of the sugar moiety, with some exceptions such as a few anti-MUC1 antibodies that primarily recognize the peptide backbone and use the sugar to facilitate shape complementarity or to establish a limited number of interactions with the protein. Focusing specifically on the GalNAc moiety, we can observe that there is some degeneracy of interactions within the same protein families, likely due to substrate flexibility. However, when all studied proteins are considered together, despite the commonalities within each protein family, no pattern can be discerned between the different families, apart from the presence of common residues such as Tyr, His, or Asp, which are responsible for hydrogen bonds. The lack of a pattern can be anticipated, given the diverse functions of mucinases, glycosyltransferases, antibodies, and lectins. Finally, it is important to point out that the conformational differences observed in solution in glycopeptides bearing GalNAc-α-1-O-Ser or GalNAc-α-1-O-Thr also can be found in the bound state. This unique characteristic is exploited, for instance, by the enzyme C1GalT1 to broadly glycosylate both acceptor substrates. The findings summarized in this review may contribute to the rational structure-guided development of therapeutic vaccines, novel diagnostic tools for early cancer detection, and new cancer treatments for cancer with tailored anti-Tn or anti-STn antibodies or new drugs to inhibit GalNAc-T isoenzymes.

Light-Triggered Control of Glucocerebrosidase Inhibitors: Towards Photoswitchable Pharmacological Chaperones.

Piperidine-based photoswitchable derivatives have been developed as putative pharmacological chaperones for glucocerebrosidase (GCase), the defective enzyme in Gaucher disease (GD). The structure-activity study revealed that both the iminosugar and the light-sensitive azobenzene are essential features to exert inhibitory activity towards human GCase and a system with the correct inhibition trend (IC50 of the light-activated form lower than IC50 of the dark form) was identified. Kinetic analyses showed that all compounds are non-competitive inhibitors (mixed or pure) of GCase and the enzyme allosteric site involved in the interaction was identified by means of MD simulations. A moderate activity enhancement of mutant GCase assessed in GD patients' fibroblasts (ex vivo experiments) carrying the most common mutation was recorded. This promising observation paves the way for further studies to improve the benefit of the light-to-dark thermal conversion for chaperoning activity.

Silver-Free Gold-Catalyzed Heterocyclizations through Intermolecular H-Bonding Activation.

Modulable monosulfonyl squaramides have been shown to exert activation of gold(I) chloride complexes through H-bonding in an intermolecular way. Combinations of (PPh3)AuCl or IPrAuCl complexes and an optimal sulfonyl squaramide cocatalyst bearing two 3,5-bis(trifluoromethyl)phenyl groups efficiently catalyzed diverse heterocyclizations and a cyclopropanation reaction, avoiding in all cases undesired side reactions. Computational studies indicate that the Au-Cl bond breaks by transligation to the triple bond in a ternary complex formed by the actual AuCl···HBD catalyst and the substrate.

Acetyl Group Migration in Xylan and Glucan Model Compounds as Studied by Experimental and Computational Methods.

It was recently demonstrated by us that acetyl groups in oligosaccharides can migrate not only within one saccharide unit but also between two different saccharide units. Kinetics of this phenomenon were previously investigated in both mannan model compounds and a naturally occurring polysaccharide. In addition to mannans, there are also several other naturally acetylated polysaccharides, such as xyloglucans and xylans. Both xyloglucans and xylans are some of the most common acetylated polysaccharides in nature, displaying important roles in the plant cells. Considering the various biological roles of natural polysaccharides, it could be hypothesized that the intramolecular migration of acetyl groups might also be associated with regulation of the biological activity of polysaccharides in nature. Consequently, a better understanding of the overall migration phenomenon across the glycosidic bonds could help to understand the potential role of such migrations in the context of the biological activity of polysaccharides. Here, we present a detailed investigation on acetyl group migration in the synthesized xylan and glucan trisaccharide model compounds by a combination of experimental and computational methods, showing that the migration between the saccharide units proceeds from a secondary hydroxyl group of one saccharide unit toward a primary hydroxyl group of the other unit.

The Essential Role of Water Molecules in the Reaction Mechanism of Protein O-Fucosyltransferase 2.

Protein O-fucosyltransferase 2 (PoFUT2) is an inverting glycosyltransferase (GT) that fucosylates thrombospondin repeats (TSRs) from group 1 and 2. PoFUT2 recognizes a large and diverse number of TSRs through a dynamic network of water-mediated interactions. By X-ray structural studies of C. elegans PoFUT2 complexed to a TSR of group 2, we demonstrate that this GT recognizes similarly the 3D structure of TSRs from both groups 1 and 2. Its active site is highly exposed to the solvent, suggesting that water molecules might also play an essential role in the fucosylation mechanism. We applied QM/MM methods using human PoFUT2 as a model, and found that HsPoFUT2 follows a classical SN 2 reaction mechanism in which water molecules contribute to a great extent in facilitating the release of the leaving pyrophosphate unit, causing the H transfer from the acceptor nucleophile (Thr/Ser) to the catalytic base, which is the last event in the reaction. This demonstrates the importance of water molecules not only in recognition of the ligands but also in catalysis.

Brønsted Acid versus Phase-Transfer Catalysis in the Enantioselective Transannular Aminohalogenation of Enesultams.

We have studied the enantioselective transannular aminohalogenation reaction of unsaturated medium-sized cyclic benzosulfonamides by using both chiral Brønsted acid and phase-transfer catalysis. Under optimized conditions, a variety of bicyclic adducts can be obtained with good yields and high enantioselectivities. The mechanism of the reaction was also studied by using computational tools; we observed that the reaction involves the participation of a conformer of the nine-membered cyclic substrate with planar chirality in which the stereochemical outcome is controlled by the relative reactivity of the two pseudorotational enantiomers when interacting with the chiral catalyst.

Acyl Group Migration in Pyranosides as Studied by Experimental and Computational Methods.

Acyl group migration affects the synthesis, isolation, manipulation and purification of all acylated organic compounds containing free hydroxyl groups, in particular carbohydrates. While several isolated studies on the migration phenomenon in different buffers have been reported, comprehensive insights into the overall migration process in different monosaccharides under similar conditions have been lacking. Here, we have studied the acyl migration in different monosaccharides using five different acyl groups by a combination of experimental, kinetic and theoretical tools. The results show that the anomeric configuration in the monosaccharide has a major influence on the migration rate, together with the relative configurations of the other hydroxyl groups and the nature of the migrating acyl group. Full mechanistic model, based on computations, demonstrates that the acyl migration proceeds through an anionic stepwise mechanism with linear dependence on the [OH- ] and the pKa of the hydroxyl group toward which the acyl group is migrating.

Absence of Intermediates in the BINOL-Derived Mg(II)/Phosphate-Catalyzed Desymmetrizative Ring Expansion of 1-Vinylcyclobutanols.

The catalyzed desymmetrizative ring expansion of alkenylcyclobutanols promoted by halofunctionalization of the alkene moiety with N-bromosuccinimide has been experimentally and computationally studied. The reaction yields highly enantioenriched cyclopentanones bearing two all-carbon quaternary stereocenters, one of them being generated in the rearrangement of the cyclobutane ring and the other by enantioselective desymmetrization. The reaction is competitive with the formation of a spiroepoxide, but it turns completely selective toward the cyclopentanone when a chiral bisphosphonium magnesium salt is employed as a catalyst. Mechanistic studies support the formation of an ion pair leading to a complex with only a unit of phosphoric acid, which is the resting state of the catalytic cycle. Calculations reproduce in an excellent way the observed reactivity and predict the effect exerted by the substituents of the aromatic ring linked to the double bond. The computational studies also revealed the reaction as a highly asynchronous concerted process taking place as one kinetic step but in two stages: (i) halogenation of the double bond and (ii) rearrangement of the cyclobutane. No intermediates are present in the reaction as energy minima. The experimental scope of the reaction further confirms the predictions for the observed reactivity and selectivity.

A cost-effective IoT learning environment for the training and assessment of surgical technical skills with visual learning analytics.

BACKGROUND: Surgeons need to train and certify their technical skills. This is usually done with the intervention of experts who monitor and assess trainees. Nevertheless, this is a time-consuming task that is subject to variations among evaluators. In recent decades, subjectivity has been significantly reduced through 1) the introduction of standard curricula, such as the Fundamentals of Laparoscopic Surgery (FLS) program, which measures students' performance in specific exercises, and 2) rubrics, which are widely accepted in the literature and serve to provide feedback about the overall technical skills of the trainees. Although these two elements reduce subjectivity, they do not, however, eliminate the figure of the expert evaluator, and so the process remains time consuming. OBJECTIVES: The objective of this work is to automate those parts of the work of the expert evaluator that the technology can measure objectively, using sensors to collect evidence, and visualizations to provide feedback. We designed and developed 1) a cost-effective IoT (Internet of Things) learning environment for the training and assessment of surgical technical skills and 2) visualizations supported by the literature on visual learning analytics (VLA) to provide feedback about the exercises (in real time) and overall performance (at the end of the training) of the trainee. METHODS: A hybrid approach was followed based on previous research for the design of the sensor based IoT learning environment. Previous studies were used as the basis for getting best practices on the tracking of surgical instruments and on the detection of the force applied to the tissue, with a focus on reducing the costs of data collection. The monitoring of the specific exercises required the design of sensors and collection mechanisms from scratch as there is little existing research on this subject. Moreover, it was necessary to design the overall architecture to collect, process, synchronize and communicate the data coming from the different sensors to provide high-level information relevant to the end user. The information to be presented was already validated by the literature and the focus was on how to visualize this information and the optimal time for its presentation to end users. The visualizations were validated with 18 VLA experts assessing the technical aspects of the visualizations and 4 medical experts assessing their functional aspects. RESULTS: This IoT learning environment amplifies the evaluation mechanisms already validated by the literature, allowing automatic data collection. First, it uses IoT sensors to automatically correct two of the exercises defined in the FLS (peg transfer and precision cutting), providing real-time visualizations. Second it monitors the movement of the surgical instruments and the force applied to the tissues during the exercise, computing 6 of the high-level indicators used by expert evaluators in their rubrics (efficiency, economy of movement, hand tremor, depth perception, bimanual dexterity, and respect for tissue), providing feedback about the technical skills of the trainee using a radar chart with these six indicators at the end of the training (summative visualizations). CONCLUSIONS: The proposed IoT learning environment is a promising and cost-effective alternative to help in the training and assessment of surgical technical skills. The system shows the trainees' progress and presents new indicators about the correctness of each specific exercise through real-time visualizations, as well as their general technical skills through summative visualizations, aligned with the 6 more frequent indicators in standardized scales. Early results suggest that although both types of visualizations are useful, it is necessary to reduce the cognitive load of the graphs presented in real time during training. Nevertheless, an additional evaluation is needed to confirm these results.

Piperidine Azasugars Bearing Lipophilic Chains: Stereoselective Synthesis and Biological Activity as Inhibitors of Glucocerebrosidase (GCase).

We report a straightforward synthetic strategy for the preparation of trihydroxypiperidine azasugars decorated with lipophilic chains at both the nitrogen and the adjacent carbon as potential inhibitors of the lysosomal enzyme glucocerebrosidase (GCase), which is involved in Gaucher disease. The procedure relies on the preparation of C-erythrosyl N-alkylated nitrones 10 through reaction of aldehyde 8 and primary amines 13 followed by oxidation of the imines formed in situ with the methyltrioxorhenium catalyst and urea hydrogen peroxide. The addition of octylMgBr to nitrone 10e provided access to both epimeric hydroxylamines 21 and 22 with opposite configuration at the newly created stereocenter in a stereodivergent and completely stereoselective way, depending on the absence or presence of BF3·Et2O. Final reductive amination and acetonide deprotection provided compounds 14 and 15 from low-cost d-mannose in remarkable 43 and 32% overall yields, respectively, over eight steps. The C-2 R-configured bis-alkylated trihydroxypiperidine 15 was the best ligand for GCase (IC50 = 15 µM), in agreement with MD simulations that allowed us to identify the chair conformation corresponding to the best binding affinity.

Biomarkers-based personalized follow-up in chronic heart failure improves patient's outcomes and reduces care associate cost.

BACKGROUND: Heart failure (HF) is a major and growing medical and economic problem, with high prevalence and incidence rates worldwide. Cardiac Biomarker is emerging as a novel tool for improving management of patients with HF with a reduced left ventricular ejection fraction (HFrEF). METHODS: This is a before and after interventional study, that assesses the impact of a personalized follow-up procedure for HF on patient's outcomes and care associated cost, based on a clinical model of risk stratification and personalized management according to that risk. A total of 192 patients were enrolled and studied before the intervention and again after the intervention. The primary objective was the rate of readmissions, due to a HF. Secondary outcome compared the rate of ED visits and quality of life improvement assessed by the number of patients who had reduced NYHA score. A cost-analysis was also performed on these data. RESULTS: Admission rates significantly decreased by 19.8% after the intervention (from 30.2 to 10.4), the total hospital admissions were reduced by 32 (from 78 to 46) and the total length of stay was reduced by 7 days (from 15 to 9 days). The rate of ED visits was reduced by 44% (from 64 to 20). Thirty-one percent of patients had an improved functional class score after the intervention, whereas only 7.8% got worse. The overall cost saving associated with the intervention was € 72,769 per patient (from € 201,189 to € 128,420) and €139,717.65 for the whole group over 1 year. CONCLUSIONS: A personalized follow-up of HF patients led to important outcome benefits and resulted in cost savings, mainly due to the reduction of patient hospitalization readmissions and a significant reduction of care-associated costs, suggesting that greater attention should be given to this high-risk cohort to minimize the risk of hospitalization readmissions.

Brønsted Acid Catalyzed (4 + 2) Cyclocondensation of 3-Substituted Indoles with Donor-Acceptor Cyclopropanes.

Acylcyclopropanes are employed as useful donor-acceptor cyclopropanes that undergo formal (4 + 2) cyclocondensation with N-unprotected 3-substituted indoles in the presence of a Brønsted acid catalyst. The reaction involves the simultaneous alkylation of both the N and C-2 positions of the indole and provides access to the 8,9-dihydropyrido[1,2-a]indole scaffold that is the central core of several biologically relevant indole alkaloids in excellent yields and good selectivities.

Objective and automated assessment of surgical technical skills with IoT systems: A systematic literature review.

The assessment of surgical technical skills to be acquired by novice surgeons has been traditionally done by an expert surgeon and is therefore of a subjective nature. Nevertheless, the recent advances on IoT (Internet of Things), the possibility of incorporating sensors into objects and environments in order to collect large amounts of data, and the progress on machine learning are facilitating a more objective and automated assessment of surgical technical skills. This paper presents a systematic literature review of papers published after 2013 discussing the objective and automated assessment of surgical technical skills. 101 out of an initial list of 537 papers were analyzed to identify: 1) the sensors used; 2) the data collected by these sensors and the relationship between these data, surgical technical skills and surgeons' levels of expertise; 3) the statistical methods and algorithms used to process these data; and 4) the feedback provided based on the outputs of these statistical methods and algorithms. Particularly, 1) mechanical and electromagnetic sensors are widely used for tool tracking, while inertial measurement units are widely used for body tracking; 2) path length, number of sub-movements, smoothness, fixation, saccade and total time are the main indicators obtained from raw data and serve to assess surgical technical skills such as economy, efficiency, hand tremor, or mind control, and distinguish between two or three levels of expertise (novice/intermediate/advanced surgeons); 3) SVM (Support Vector Machines) and Neural Networks are the preferred statistical methods and algorithms for processing the data collected, while new opportunities are opened up to combine various algorithms and use deep learning; and 4) feedback is provided by matching performance indicators and a lexicon of words and visualizations, although there is considerable room for research in the context of feedback and visualizations, taking, for example, ideas from learning analytics.

Computational evidence of glycosyl cations.

Glycosyl cations are key intermediates in the glycosylation reactions taking place through a SN1-type mechanism. To obtain a reliable description of the glycosylation reaction mechanism a combination of computational studies and experimental data such as kinetic isotopic effects is needed. Computational studies have elucidated SN2-type glycosylation reaction mechanisms, but elucidation of mechanisms in which ion pairs can be formed presents some difficulties because of the recombination of the ions. Recent topological and dynamic studies open the door to the ultimate confirmation of the presence of glycosyl cations in the form of intimate ion pairs during certain glycosylation reactions. This review covers the state-of-the-art tools and applications of computational chemistry mainly developed during the last ten years to understand glycosylation reactions in which an oxocarbenium ion could be involved.