Exploring the interaction of N-(benzothiazol-2-yl)pyrrolamide DNA gyrase inhibitors with the GyrB ATP-binding site lipophilic floor: A medicinal chemistry and QTAIM study.

N-(Benzothiazole-2-yl)pyrrolamide DNA gyrase inhibitors with benzyl or phenethyl substituents attached to position 3 of the benzothiazole ring or to the carboxamide nitrogen atom were prepared and studied for their inhibition of Escherichia coli DNA gyrase by supercoiling assay. Compared to inhibitors bearing the substituents at position 4 of the benzothiazole ring, the inhibition was attenuated by moving the substituent to position 3 and further to the carboxamide nitrogen atom. A co-crystal structure of (Z)-3-benzyl-2-((4,5-dibromo-1H-pyrrole-2-carbonyl)imino)-2,3-dihydrobenzo[d]-thiazole-6-carboxylic acid (I) in complex with E. coli GyrB24 (ATPase subdomain) was solved, revealing the binding mode of this type of inhibitor to the ATP-binding pocket of the E. coli GyrB subunit. The key binding interactions were identified and their contribution to binding was rationalised by quantum theory of atoms in molecules (QTAIM) analysis. Our study shows that the benzyl or phenethyl substituents bound to the benzothiazole core interact with the lipophilic floor of the active site, which consists mainly of residues Gly101, Gly102, Lys103 and Ser108. Compounds with substituents at position 3 of the benzothiazole core were up to two orders of magnitude more effective than compounds with substituents at the carboxamide nitrogen. In addition, the 6-oxalylamino compounds were more potent inhibitors of E. coli DNA gyrase than the corresponding 6-acetamido analogues.

Synthesis of a new 2-prenylated quinoline as potential drug for metabolic syndrome with pan-PPAR activity and anti-inflammatory effects.

We have previously reported the total synthesis and structure-activity relationships (SAR) of 2-prenylated benzopyrans with PPAR agonist activity. Herein, we have described the synthesis and PPAR activity of 2-prenylated benzopyrans and 2-prenylated quinolines. The benzopyran nucleus was generated via enamine-catalyzed Kabbe condensation, and the quinoline nucleus via Friedländer condensation. Results demonstrated that both benzopyran (5a) and quinoline (4b) derivatives bearing a γ,δ-unsaturated ester displayed a pan-PPAR agonism. They were full PPARα agonists, but showed different preferences for PPARγ and PPARß/δ activation. It was noteworthy that quinoline 4b displayed full hPPARα activation (2-fold than WY-14,643), weak PPARß/δ and partial PPARγ activation. In addition, quinoline 4b showed anti-inflammatory effects on macrophages by reducing LPS-induced expression of both MCP-1 and IL-6. Therefore, 4b emerges as a first-in-class promising hit compound for the development of potential therapeutics aimed at treating metabolic syndrome, metabolic dysfunction-associated fatty liver disease (MAFLD), and its associated cardiovascular comorbidities.

Benzopyran hydrazones with dual PPARα/γ or PPARα/δ agonism and an anti-inflammatory effect on human THP-1 macrophages.

Peroxisome proliferator-activated receptors (PPARs) play a major role in regulating inflammatory processes, and dual or pan-PPAR agonists with PPARγ partial activation have been recognised to be useful to manage both metabolic syndrome and metabolic dysfunction-associated fatty liver disease (MAFLD). Previous works have demonstrated the capacity of 2-prenylated benzopyrans as PPAR ligands. Herein, we have replaced the isoprenoid bond by hydrazone, a highly attractive functional group in medicinal chemistry. In an attempt to discover novel and safety PPAR activators, we efficiently prepared benzopyran hydrazone/hydrazine derivatives containing benzothiazole (series 1) or 5-chloro-3-(trifluoromethyl)-2-pyridine moiety (series 2) with a 3- or 7-carbon side chain at the 2-position of the benzopyran nucleus. Benzopyran hydrazones 4 and 5 showed dual hPPARα/γ agonism, while hydrazone 14 exerted dual hPPARα/δ agonism. These three hydrazones greatly attenuated inflammatory markers such as IL-6 and MCP-1 on the THP-1 macrophages via NF-κB activation. Therefore, we have discovered novel hits (4, 5 and 14), containing a hydrazone framework with dual PPARα/γ or PPARα/δ partial agonism, depending on the length of the side chain. Benzopyran hydrazones emerge as potential lead compounds which could be useful for treating metabolic diseases.

Inhibitory effect of galantamine and donepezil combination against cholinesterase: An in silico and in vitro study.

This study aimed to evaluate the in silico and in vitro inhibitory effect of the combined use of galantamine (GAL) and donepezil (DON) against acetylcholinesterase and butyrylcholinesterase (BuChE) enzymes. In silico and in vitro cholinesterase analysis were carried out for GAL and DON alone and combined. Molecular modeling studies were carried out (docking analysis, molecular dynamics simulation, and quantum theory of atoms in molecules). Cholinesterase's inhibitory activities by modified Ellman's method and the drug combination effect using the Chou-Talalay method were assayed. GAL/DON combination showed the co-occupancy of the ligands in both enzymes through in silico studies. Regarding in vitro BuChE inhibition analyses, three of five combinations showed an interaction between GAL and DON at the threshold of additive affect (0.9 < CI < 1.1), with a tendency toward a synergistic effect for higher concentrations. This is the first report showing the efficacy of the GAL/DON combinations inhibiting BuChE, showing the importance of analyzing the behavior of different ligands when co-occupancy into the active site is possible. These combinations might be a possible therapy to improved efficacy, reduced doses, minor side effects, and high levels of the neurotransmitter in the synaptic space for Alzheimer's disease.

Second generation of pyrimidin-quinolone hybrids obtained from virtual screening acting as sphingosine kinase 1 inhibitors and potential anticancer agents.

We report here the virtual screening design, synthesis and activity of eight new inhibitors of SphK1. For this study we used a pre-trained Graph Convolutional Network (GCN) combined with docking calculations. This exploratory analysis proposed nine compounds from which eight displayed significant inhibitory effect against sphingosine kinase 1 (SphK1) demonstrating a high level of efficacy for this approach. Four of these compounds also displayed anticancer activity against different tumor cell lines, and three of them (5), (6) and (7) have shown a wide inhibitory action against many of the cancer cell line tested, with GI50 below 5 µM, being (5) the most promising with TGI below 10 µM for the half of cell lines. Our results suggest that the three most promising compounds reported here are the pyrimidine-quinolone hybrids (1) and (6) linked by p-aminophenylsulfanyl and o-aminophenol fragments respectively, and (8) without such aryl linker. We also performed an exhaustive study about the molecular interactions that stabilize the different ligands at the binding site of SphK1. This molecular modeling analysis was carried out by using combined techniques: docking calculations, MD simulations and QTAIM analysis. In this study we also included PF543, as reference compound, in order to better understand the molecular behavior of these ligands at the binding site of SphK1.These results provide useful information for the design of new inhibitors of SphK1 possessing these structural scaffolds.

The Mechanism of Antimicrobial Small-Cationic Peptides from Coarse-Grained Simulations.

Antimicrobial cationic peptides (AMPs) are excellent candidates for use as therapeutic antimicrobial agents. Among them, short peptides possessing sequences of 9-11 amino acids have some advantages over long-sequence peptides. However, one of the main limitations of short peptides is that their mechanism of action at the molecular level is not well-known. In this article, we report a model based on multiscale molecular dynamics simulations of short peptides interacting with vesicles containing palmitoyl-oleoyl-phosphatidylglycerol (POPG)/palmitoyl-oleoyl-phosphatidylethanolamine (POPE). Simulations using this approach have allowed us to understand the different behaviors of peptides with antimicrobial activity with respect to those that do not produce this effect. We found remarkable agreement with a series of experimental results directly supporting our model. Moreover, these results allow us to understand the mechanism of action at the molecular level of these short peptides. Our simulations suggest that mechanical inhomogeneities appear in the membrane, promoting membrane rupture when a threshold concentration of peptides adsorbed on the membrane is achieved. These results explain the high structural demand for these peptides to maintain a delicate balance between the affinity for the bilayer surface, a low peptide-peptide repulsion (in order to reach the threshold concentration), and an acceptable tendency to penetrate into the bilayer. This mechanism is different from those proposed for peptides with long amino acid sequences. Such information is very useful from the medicinal chemistry point of view for the design of new small antimicrobial peptides.

Antinociceptive effect of cyclic and linear diterpenoids as new atypical agonists of κ-opioid receptors obtained from four species of the Baccharis genus, and vehiculated in nanometric niosomes.

New natural analgesic compounds that act in KORs are important alternatives for potential therapeutical use in medicine. In this work, we report and compare here the antinociceptive activity displayed by cyclic and linear diterpenes, obtained from the genus Baccharis. The antinociceptive activities determined were relatively strong, in comparison whit morphine. The antinociceptive mechanism of action was made through naloxone administration (a non-selective antagonist of opioid receptors). The more active compounds were vehiculized successfully in niosomes at nanometric scale. The observed antinociceptive activity for Bartemidiolide oxide (BARTO), obtain from Baccharis artemisioides, was greater than Flabeloic acid dimer (DACD), the first compound isolated from Baccharis flabellata that was reported possessing antinociceptive effects. We also conducted docking calculations and molecular dynamics simulations, which suggested that the newly identified diterpenes might share the molecular action mechanism reported for Salvinorin A (SalA). Molecular simulations have allowed us to appreciate some subtle differences between molecular interactions of these ligands stabilizing their respective complexes; such information might be useful for designing and searching for new inhibitors of KORs.

Anti-inflammatory effects and improved metabolic derangements in ob/ob mice by a newly synthesized prenylated benzopyran with pan-PPAR activity.

BACKGROUND AND PURPOSE: Selective peroxisome proliferator-activated receptors (PPARs) are widely used to treat metabolic complications; however, the limited effect of PPARα agonists on glucose metabolism and the adverse effects associated with selective PPARγ activators have stimulated the development of novel pan-PPAR agonists to treat metabolic disorders. Here, we synthesized a new prenylated benzopyran (BP-2) and evaluated its PPAR-activating properties, anti-inflammatory effects and impact on metabolic derangements. EXPERIMENTAL APPROACH: BP-2 was used in transactivation assays to evaluate its agonism to PPARα, PPARß/δ and PPARγ. A parallel-plate flow chamber was employed to investigate its effect on TNFα-induced leukocyte-endothelium interactions. Flow cytometry and immunofluorescence were used to determine its effects on the expression of endothelial cell adhesion molecules (CAMs) and chemokines and p38-MAPK/NF-κB activation. PPARs/RXRα interactions were determined using a gene silencing approach. Analysis of its impact on metabolic abnormalities and inflammation was performed in ob/ob mice. KEY RESULTS: BP-2 displayed strong PPARα activity, with moderate and weak activity against PPARß/δ and PPARγ, respectively. In vitro, BP-2 reduced TNFα-induced endothelial ICAM-1, VCAM-1 and fractalkine/CX3CL1 expression, suppressed mononuclear cell arrest via PPARß/δ-RXRα interactions and decreased p38-MAPK/NF-κB activation. In vivo, BP-2 improved the circulating levels of glucose and triglycerides in ob/ob mice, suppressed T-lymphocyte/macrophage infiltration and proinflammatory markers in the liver and white adipose tissue, but increased the expression of the M2-like macrophage marker CD206. CONCLUSION AND IMPLICATIONS: BP-2 emerges as a novel pan-PPAR lead candidate to normalize glycemia/triglyceridemia and minimize inflammation in metabolic disorders, likely preventing the development of further cardiovascular complications.

Evaluating the conformational space of the active site of D2 dopamine receptor. Scope and limitations of the standard docking methods.

We report here for the first time the potential energy surfaces (PES) of phenyletilamine (PEA) and meta-tyramine (m-OH-PEA) at the D2 dopamine receptor (D2DR) binding site. PESs not only allow us to observe all the critical points of the surface (minimums, maximums, and transition states), but also to note the ease or difficulty that each local minima have for their conformational inter-conversions and therefore know the conformational flexibility that these ligands have in their active sites. Taking advantage of possessing this valuable information, we analyze how accurate a standard docking study is in these cases. Our results indicate that although we have to be careful in how to carry out this type of study and to consider performing some extra-simulations, docking calculations can be satisfactory. In order to analyze in detail the different molecular interactions that are stabilizing the different ligand-receptor (L-R) complexes, we carried out quantum theory of atoms in molecules (QTAIM) computations and NMR shielding calculations. Although some of these techniques are a bit tedious and require more computational time, our results demonstrate the importance of performing computational simulations using different types of combined techniques (docking/MD/hybrid QM-MM/QTAIM and NMR shielding calculations) in order to obtain more accurate results. Our results allow us to understand in details the molecular interactions stabilizing and destabilizing the different L-R complexes reported here. Thus, the different activities observed for dopamine (DA), m-OH-PEA, and PEA can be clearly explained at molecular level.

Arylated analogues of cypronazole: fungicidal effect and activity on human fibroblasts. Docking analysis and molecular dynamics simulations.

Triadimefon (TDM) and cyproconazole (CPZ) are two triazoles widely used as fungicides. Several azoles were synthesised starting from commercial TDM and CPZ. The compounds were evaluated against phytopathogenic filamentous fungi, including Aspergillus fumigatus (AF), A. niger (AN), A. ustus (AU), A. japonicus (AJ), A. terreus (AT), Fusarium oxysporum and Botrytis cinerea isolated from grapevine in the province of San Juan, Argentina. Three of the synthesised compounds (1-(Biphenyl-4-yloxy)-3,3-dimethyl-1-(1H-1,2,4-triazol-1-yl)butan-2-one, 1; 2-(Biphenyl-4-yl)-3-cyclopropyl-1-(1H-1,2,4-triazol-1-yl)butan-2-ol, 3; 3-Cyclopropyl-2-(4'-fluorobiphenyl-4-yl)-1-(1H-1,2,4-triazol1-yl)butan-2-ol, 4) presented remarkable in vitro fungicidal properties, with better effects than TDM and CPZ on some of the target fungi. Cytotoxicity was assessed using human lung fibroblasts MRC5. Derivative 1, with IC50 values of 389.4 µM, was less toxic towards MRC-5 human lung fibroblasts than commercial TDM (248.5 µM) and CPZ (267.4 µM). Docking analysis and molecular dynamics simulations suggest that the compounds present the same interaction in the binding pocket of the CYP51B enzyme and with the same amino acids as CPZ. The derivatives investigated could be considered broad-spectrum but with some selectivity towards imperfect fungi.

Pseudorotaxane formation affected by stereo-electronic effects. A theoretical and experimental study.

We report a theoretical and experimental study on different complexes of pseudorotaxanes possessing pyridine axles. In order to evaluate the stereo-electronic effects of the methyl substituents in the pyridine ring, complexes with different substitution patterns were synthesized. In this way, it was possible to analyze the different behaviors of these complexes according to the positions of their methyl substituents. Combined techniques of molecular dynamics and quantum mechanical calculations with the help of molecular electrostatic potentials for a simpler visualization of the electronic effects were employed. We have sought experimental support of NMR spectroscopy analysis to corroborate the conclusions obtained from the molecular simulations. Our results not only clearly demonstrate that both electronic and steric effects play key roles in the feasibility of the formation of such complexes, but also the simulations reported here might predict the degree of difficulty of their formation. The combination of computational techniques employed here seems to be an excellent approach to be able to predict whether or not a complex can be formed and with what degree of difficulty. In addition, our experimental and theoretical results have allowed us to visualize the formation of external complexes in the rotaxanes reported here. In this case, the use of bolaforms with trimethylammonium groups at both ends was very useful to evaluate in detail the formation of the so-called external complexes in these systems.

Quinazoline-tethered hydrazone: A versatile scaffold toward dual anti-TB and EGFR inhibition activities in NSCLC.

Globally, lung cancer and tuberculosis are considered to be very serious and complex diseases. Evidence suggests that chronic infection with tuberculosis (TB) can often lead to lung tumors; therefore, developing drugs that target both diseases is of great clinical significance. In our study, we designed and synthesized a suite of 14 new quinazolinones (5a-n) and performed biological investigations of these compounds in Mycobacterium tuberculosis (MTB) and cancer cell lines. In addition, we conducted a molecular modeling study to determine the mechanism of action of these compounds at the molecular level. Compounds that showed anticancer activity in the preliminary screening were further evaluated in three cancer cell lines (A549, Calu-3, and BT-474 cells) and characterized in an epidermal growth factor receptor (EGFR) binding assay. Cytotoxicity in noncancerous lung fibroblast cells was also evaluated to obtain safety data. Our theoretical and experimental studies indicated that our compounds showed a mechanism of action similar to that of erlotinib by inhibiting the EGFR tyrosine kinase. In turn, the antituberculosis activity of these compounds would be produced by the inhibition of enoyl-ACP-reductase. From our findings, we were able to identify two potential lead compounds (5i and 5l) with dual activity and elevated safety toward noncancerous lung fibroblast cells. In addition, our data identified three compounds with excellent anti-TB activities (compounds 5i, 5l, and 5n).

Novel Sulfonamide-Based Carbamates as Selective Inhibitors of BChE.

A series of 14 target benzyl [2-(arylsulfamoyl)-1-substituted-ethyl]carbamates was prepared by multi-step synthesis and characterized. All the final compounds were tested for their ability to inhibit acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) in vitro, and the selectivity index (SI) was determined. Except for three compounds, all compounds showed strong preferential inhibition of BChE, and nine compounds were even more active than the clinically used rivastigmine. Benzyl {(2S)-1-[(2-methoxybenzyl)sulfamoyl]-4-methylpentan-2-yl}carbamate (5k), benzyl {(2S)-1-[(4-chlorobenzyl)sulfamoyl]-4-methylpentan-2-yl}carbamate (5j), and benzyl [(2S)-1-(benzylsulfamoyl)-4-methylpentan-2-yl]carbamate (5c) showed the highest BChE inhibition (IC50 = 4.33, 6.57, and 8.52 µM, respectively), indicating that derivatives 5c and 5j had approximately 5-fold higher inhibitory activity against BChE than rivastigmine, and 5k was even 9-fold more effective than rivastigmine. In addition, the selectivity index of 5c and 5j was approx. 10 and that of 5k was even 34. The process of carbamylation and reactivation of BChE was studied for the most active derivatives 5k, 5j. The detailed information about the mode of binding of these compounds to the active site of both BChE and AChE was obtained in a molecular modeling study. In this study, combined techniques (docking, molecular dynamic simulations, and QTAIM (quantum theory of atoms in molecules) calculations) were employed.

Covalence and π-electron delocalization influence on hydrogen bonds in proton transfer process of o-hydroxy aryl Schiff bases: A combined NMR and QTAIM analysis.

Within the framework of the density functional theory approach, we studied the relationship between the chemical nature of intramolecular hydrogen bonds (HBs) and nuclear magnetic resonance (NMR) parameters, J-couplings and 1H-chemical shifts [δ(1H)], of the atoms involved in such bonds in o-hydroxyaryl Schiff bases during the proton transfer process. For the first time, the shape of the dependence of the degree of covalence in HBs on 1J(N-H), 1J(O-H), 2hJ(O-N), and δ(1H) during the proton transfer process in o-hydroxyaryl Schiff bases was analyzed. Parameters obtained from Bader's theory of atoms in molecules were used to assess the dependence of covalent character in HBs with both the NMR properties. The influence of π-electronic delocalization on 2hJ(N-O) under the proton transfer process was investigated. 2hJ(O-N) in a Mannich base was also studied in order to compare the results with an unsaturated system. In addition, substituent effects on the phenolic ring were investigated. Our results indicate that the covalent character of HBs on both sides of the transition state undergoes a smooth exponential increase as the δ(1H) moves downfield. The degree of covalence of the N⋯H (O⋯H) bond increases linearly as 1J(N-H) (1J(O-H)) becomes more negative, even after reaching the transition state. Non-vanishing values of spin dipolar (SD) and paramagnetic spin orbital terms of 2hJ(O-N) show that π-electronic delocalization has a non-negligible effect on tautomeric equilibrium and gives evidence of the presence of the resonance assisted HB.Variation of the SD term of 2hJ(O-N) follows a similar pattern as the change in the para-delocalization aromaticity index of the chelate ring.

New short cationic antibacterial peptides. Synthesis, biological activity and mechanism of action.

We report a theoretical and experimental study on a new series of small-sized antibacterial peptides. Synthesis and bioassays for these peptides are reported here. In addition, we evaluated different physicochemical parameters that modulate antimicrobial activity (charge, secondary structure, amphipathicity, hydrophobicity and polarity). We also performed molecular dynamic simulations to assess the interaction between these peptides and their molecular target (the membrane). Biophysical characterization of the peptides was carried out with different techniques, such as circular dichroism (CD), linear dichroism (LD), infrared spectroscopy (IR), dynamic light scattering (DLS), fluorescence spectroscopy and TEM studies using model systems (liposomes) for mammalian and bacterial membranes. The results of this study allow us to draw important conclusions on three different aspects. Theoretical and experimental results indicate that small-sized peptides have a particular mechanism of action that is different to that of large peptides. These results provide additional support for a previously proposed four-step mechanism of action. The possible pharmacophoric requirement for these small-sized peptides is discussed. Furthermore, our results indicate that a net +4 charge is the adequate for 9 amino acid long peptides to produce antibacterial activity. The information reported here is very important for designing new antibacterial peptides with these structural characteristics.

A Potent N-(piperidin-4-yl)-1H-pyrrole-2-carboxamide Inhibitor of Adenylyl Cyclase of G. lamblia: Biological Evaluation and Molecular Modelling Studies.

In this work, we report a derivative of N-(piperidin-4-yl)-1H-pyrrole-2-carboxamide as a new inhibitor for adenylyl cyclase of Giardia lamblia which was obtained from a study using structural data of the nucleotidyl cyclase 1 (gNC1) of this parasite. For such a study, we developed a model for this specific enzyme by using homology techniques, which is the first model reported for gNC1 of G. lamblia. Our studies show that the new inhibitor has a competitive mechanism of action against this enzyme. 2-Hydroxyestradiol was used as the reference compound for comparative studies. Results in this work are important from two points of view. on the one hand, an experimentally corroborated model for gNC1 of G. lamblia obtained by molecular modelling is presented; on the other hand, the new inhibitor obtained is an undoubtedly excellent starting structure for the development of new metabolic inhibitors for G. lamblia.

Hydroxynaphthalenecarboxamides and substituted piperazinylpropandiols, two new series of BRAF inhibitors. A theoretical and experimental study.

The oncogenic mutated kinase BRAFV600E is an attractive molecular target because it is expressed in several human cancers, including melanoma. To present, only three BRAF small inhibitors are approved by the FDA for the treatment of patients with metastatic melanoma: Vemurafenib, Dabrafenib and Encorafenib. Although many protocol treatments have been probed in clinical trials, BRAF inhibition has a limited effectiveness because patients invariably develop resistance and secondary toxic effects associated with the therapy. These limitations highlight the importance of designing new and better inhibitors with different structures that could establish different interactions in the active site of the enzyme and therefore decrease resistance progress. Considering the data from our previous report, here we studied two series of derivatives of structural scaffolds as potential BRAF inhibitors: hydroxynaphthalenecarboxamides and substituted piperazinylpropandiols. Our results indicate that structural analogues of substituted piperazinylpropandiols do not show significantly better activities to that previously reported. In contrast, the hydroxynaphthalenecarboxamides derivatives significantly inhibited cell viability and ERK phosphorylation, a measure of BRAF activity, in Lu1205 BRAFV600E melanoma cells. In order to better understand these experimental results, we carried out a molecular modeling study using different combined techniques: docking, MD simulations and quantum theory of atoms in molecules (QTAIM) calculations. Thus, by using this approach we determined that the molecular interactions that stabilize the different molecular complexes are closely related to Vemurafenib, a well-documented BRAF inhibitor. Furthermore, we found that bi-substituted compounds may interact more strongly respect to the mono-substituted analogues, by establishing additional interactions with the DFG-loop at the BRAF-active site. On the bases of these results we synthesized and tested a new series of hydroxynaphthalenecarboxamides bi-substituted. Remarkably, all these compounds displayed significant inhibitory effects on the bioassays performed. Thus, the structural information reported here is important for the design of new BRAFV600E inhibitors possessing this type of structural scaffold.

Conformational and electronic study of dopamine interacting with the D2 dopamine receptor.

We report an exhaustive conformational and electronic study on dopamine (DA) interacting with the D2 dopamine receptor (D2 DR). For the first time, the complete surface of the conformational potential energy of the complex DA/D2 DR is reported. Such a surface was obtained through the use of QM/MM calculations. A detailed study of the molecular interactions that stabilize and destabilize the different molecular complexes was carried out using two techniques: Quantum Theory of Atoms in Molecules computations and nuclear magnetic shielding constants calculations. A comparative study of the behavior of DA in the gas phase, aqueous solution, and in the active site of D2 DR has allowed us to evaluate the degree of deformation suffered by the ligand and, therefore, analyze how rustic are the lock-key model and the induced fit theory in this case. Our results allow us to propose one of the conformations obtained as the "biologically relevant" conformation of DA when it is interacting with the D2 DR.

Targeting defective sphingosine kinase 1 in Niemann-Pick type C disease with an activator mitigates cholesterol accumulation.

Niemann-Pick type C (NPC) disease is a lysosomal storage disorder arising from mutations in the cholesterol-trafficking protein NPC1 (95%) or NPC2 (5%). These mutations result in accumulation of low-density lipoprotein-derived cholesterol in late endosomes/lysosomes, disruption of endocytic trafficking, and stalled autophagic flux. Additionally, NPC disease results in sphingolipid accumulation, yet it is unique among the sphingolipidoses because of the absence of mutations in the enzymes responsible for sphingolipid degradation. In this work, we examined the cause for sphingosine and sphingolipid accumulation in multiple cellular models of NPC disease and observed that the activity of sphingosine kinase 1 (SphK1), one of the two isoenzymes that phosphorylate sphingoid bases, was markedly reduced in both NPC1 mutant and NPC1 knockout cells. Conversely, SphK1 inhibition with the isotype-specific inhibitor SK1-I in WT cells induced accumulation of cholesterol and reduced cholesterol esterification. Of note, a novel SphK1 activator (SK1-A) that we have characterized decreased sphingoid base and complex sphingolipid accumulation and ameliorated autophagic defects in both NPC1 mutant and NPC1 knockout cells. Remarkably, in these cells, SK1-A also reduced cholesterol accumulation and increased cholesterol ester formation. Our results indicate that a SphK1 activator rescues aberrant cholesterol and sphingolipid storage and trafficking in NPC1 mutant cells. These observations highlight a previously unknown link between SphK1 activity, NPC1, and cholesterol trafficking and metabolism.

Second-generation 4,5,6,7-tetrahydrobenzo[d]thiazoles as novel DNA gyrase inhibitors.

Aim: DNA gyrase and topoisomerase IV are essential bacterial enzymes, and in the fight against bacterial resistance, they are important targets for the development of novel antibacterial drugs. Results: Building from our first generation of 4,5,6,7-tetrahydrobenzo[d]thiazole-based DNA gyrase inhibitors, we designed and prepared an optimized series of analogs that show improved inhibition of DNA gyrase and topoisomerase IV from Staphylococcus aureus and Escherichia coli, with IC50 values in the nanomolar range. Importantly, these inhibitors also show improved antibacterial activity against Gram-positive strains. Conclusion: The most promising inhibitor, 29, is active against Enterococcus faecalis, Enterococcus faecium and S. aureus wild-type and resistant strains, with minimum inhibitory concentrations between 4 and 8 µg/ml, which represents good starting point for development of novel antibacterials.