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Cannabinoid type 2 receptor (CB2R), belonging to the endocannabinoid system, is overexpressed in pathologies characterized by inflammation, and its activation counteracts inflammatory states. Fatty acid amide hydrolase (FAAH) is an enzyme responsible for the degradation of the main endocannabinoid anandamide; thus, the simultaneous CB2R activation and FAAH inhibition may be a synergistic anti-inflammatory strategy. Encouraged by principal component analysis (PCA) data identifying a wide chemical space shared by CB2R and FAAH ligands, we designed a small library of adamantyl-benzamides, as potential dual agents, CB2R agonists, and FAAH inhibitors. The new compounds were tested for their CB2R affinity/selectivity and CB2R and FAAH activity. Derivatives 13 , 26 , and 27 , displaying the best pharmacodynamic profile as CB2R full agonists and FAAH inhibitors, decreased pro-inflammatory and increased anti-inflammatory cytokines production. Molecular docking simulations complemented the experimental findings by providing a molecular rationale behind the observed activities. These multitarget ligands constitute promising anti-inflammatory agents.

Herein, a robust and reproducible eXplainable Artificial Intelligence (XAI) approach is presented, which allows prediction of developmental toxicity, a challenging human-health endpoint in toxicology. The application of XAI as an alternative method is of the utmost importance with developmental toxicity being one of the most animal-intensive areas of regulatory toxicology. In this work, the established CAESAR (Computer Assisted Evaluation of industrial chemical Substances According to Regulations) training set made of 234 chemicals for model learning is employed. Two test sets, including as a whole 585 chemicals, were instead used for validation and generalization purposes. The proposed framework favorably compares with the state-of-the-art approaches in terms of accuracy, sensitivity, and specificity, thus resulting in a reliable support system for developmental toxicity ensuring informativeness, uncertainty estimation, generalization, and transparency. Based on the eXtreme Gradient Boosting (XGB) algorithm, our predictive model provides easy interpretative keys based on specific molecular descriptors and structural alerts enabling one to distinguish toxic and nontoxic chemicals. Inspired by the Organisation for Economic Co-operation and Development (OECD) principles for the validation of Quantitative Structure-Activity Relationships (QSARs) for regulatory purposes, the results are summarized in a standard report in portable document format, enclosing also details concerned with a density-based model applicability domain and SHAP (SHapley Additive exPlanations) explainability, the latter particularly useful to better understand the effective roles played by molecular features. Notably, our model has been implemented in TIRESIA (Toxicology Intelligence and Regulatory Evaluations for Scientific and Industry Applications), a free of charge web platform available at http://tiresia.uniba.it.

Thiolated self-assembled monolayers (SAMs) are typically used to anchor on a gold surface biomolecules serving as recognition elements for biosensor applications. Here, the design and synthesis of N -(2-hydroxyethyl)-3-mercaptopropanamide (NMPA) in biotinylated mixed SAMs is proposed as an alternative strategy with respect to on-site multistep functionalization of SAMs prepared from solutions of commercially available thiols. In this study, the mixed SAM deposited from a 10:1 solution of 3-mercaptopropionic acid (3MPA) and 11-mercaptoundecanoic acid (11MUA) is compared to that resulting from a 10:1 solution of NMPA:11MUA. To this end, surface plasmon resonance (SPR) and attenuated total reflectance infrared (ATR-IR) experiments have been carried out on both mixed SAMs after biotinylation. The study demonstrated how the fine tuning of the SAM features impacts directly on both the biofunctionalization steps, i.e., the biotin anchoring, and the biorecognition properties evaluated upon exposure to streptavidin analyte. Higher affinity for the target analyte with reduced nonspecific binding and lower detection limit has been demonstrated when NMPA is chosen as the more abundant starting thiol. Molecular dynamics simulations complemented the experimental findings providing a molecular rationale behind the performance of the biotinylated mixed SAMs. The present study confirms the importance of the functionalization design for the development of a highly performing biosensor., Competing Interests: The authors declare no competing financial interest., (Copyright © 2020 American Chemical Society.)

We present MuSSeL, a multifingerprint similarity search algorithm, able to predict putative drug targets for a given query small molecule as well as to return a quantitative assessment of its bioactivity in terms of K i or IC 50 values. Predictions are automatically made exploiting a large collection of high quality experimental bioactivity data available from ChEMBL (version 22.1) combining, in a consensus-like approach, predictions resulting from a similarity search performed using 13 different fingerprint definitions. Importantly, the herein proposed algorithm is also effective in detecting and handling activity cliffs. A calibration set including small molecules present in the last updated version of ChEMBL (version 23) was employed to properly tune the algorithm parameters. Three randomly built external sets were instead challenged for model performances. The potential use of MuSSeL was also challenged by a prospective exercise for the prediction of five bioactive compounds taken from articles published in the Journal of Medicinal Chemistry just few months ago. The paper emphasizes the importance of implementing multifingerprint consensus strategies to increase the confidence in prediction of similarity search algorithms and provides a fast and easy-to-run tool for drug target and bioactivity prediction.

P-Glycoprotein is a well-known membrane transporter responsible for the efflux of an ample spectrum of anticancer drugs. Its relevance in the management of cancer chemotherapy is increased in view of its high expression in cancer stem cells, a population of cancer cells with strong tumor-promoting ability. In the present study, a series of compounds were synthesized through structure modulation of [4'-(6,7-dimethoxy-3,4-dihydro-1 H-isoquinolin-2-ylmethyl)biphenyl-4-ol] (MC70), modifying the phenolic group of the lead compound. Among them, compound 5b emerged for its activity against the transporter (EC 50 = 15 nM) and was capable of restoring doxorubicin antiproliferative activity at nontoxic concentration. Its behavior was rationalized through a molecular modeling study consisting of a well-tempered metadynamics simulation, which allowed one to identify the most favorable binding pose, and of a subsequent molecular dynamics run, which indicated a peculiar effect of the compound on the motion pattern of the transporter.

One of the principal reasons for the chemotherapy failure is the overexpression of drug efflux pumps, ABCB1 (also known as MDR1 or P-gp) and ABCC1 (also known as MRP1), whose inhibition remains a priority to circumvent drug resistance. We have recently shown a clear trend between lipophilicity and P-glycoprotein inhibitory activity for a class of galloyl-based modulators targeting P-glycoprotein and MRP1. Herein we report a new series of polymethoxy benzamides, whose lipophilicity was modulated through the establishment of an intramolecular hydrogen bond (IMHB) which allows reaching of P-gp inhibitory activity at the submicromolar IC50 level. The present study provides a strong rationale for candidates in the presence of IMHB as a key element for a high P-gp inhibitory activity.

The multifactorial nature of chemotherapy failure in controlling cancer is often associated with the occurrence of multidrug resistance (MDR), a phenomenon likely related to the increased expression of members of the ATP binding cassette (ABC) transporter superfamily. In this respect, the most extensively characterized MDR transporters include ABCB1 (also known as MDR1 or P-glycoprotein) and ABCC1 (also known as MRP1) whose inhibition remains a priority to circumvent drug resistance. Herein, we report how the simple galloyl benzamide scaffold can be easily and properly decorated for the preparation of either MRP1 or P-gp highly selective inhibitors. In particular, some gallamides and pyrogallol-1-monomethyl ethers showed remarkable affinity and selectivity toward MRP1. On the other hand, trimethyl ether galloyl anilides, with few exceptions, exhibited moderate to very high and selective P-gp inhibition.

A molecular library of quaternary ammonium salts (QASs), mainly composed of symmetrical bis-quaternary heterocyclic bromides exhibiting choline kinase (ChoK) inhibitory activity, were evaluated for their ability to inhibit acetyl- and butyrylcholinesterase (AChE and BChE, respectively). The molecular framework of QASs consisted of two positively charged heteroaromatic (pyridinium or quinolinium) or sterically hindered aliphatic (quinuclidinium) nitrogen rings kept at an appropriate distance by lipophilic rigid or semirigid linkers. Many homodimeric QASs showed AChE and BChE inhibitory potency in the nanomolar range along with a low enzymatic selectivity. Computational studies on AChE, BChE, and ChoK allowed identification of the key molecular determinants for high affinity and selectivity over either one of the three enzymes and guided the design of a hybrid bis-QAS (56) exhibiting the highest AChE affinity (IC(50) = 15 nM) and selectivity over BChE and ChoK (SI = 50 and 562, respectively) and a promising pharmacological potential in myasthenia gravis and neuromuscular blockade.

The design, synthesis, and biological evaluation of a series of new aromatase (AR, CYP19) inhibitors bearing an imidazole ring linked to a 7-substituted coumarin scaffold at position 4 (or 3) are reported. Many compounds exhibited an aromatase inhibitory potency in the nanomolar range along with a high selectivity over 17-α-hydroxylase/C17-20 lyase (CYP17). The most potent AR inhibitor was the 7-(3,4-difluorophenoxy)-4-imidazolylmethyl coumarin 24 endowed with an IC(50) = 47 nM. Docking simulations on a selected number of coumarin derivatives allowed the identification of the most important interactions driving the binding and clearly indicated the allowed and disallowed regions for appropriate structural modifications of coumarins and closely related heterocyclic molecular scaffolds.

In an effort to discover novel selective monoamine oxidase (MAO) B inhibitors with favorable physicochemical and pharmacokinetic profiles, 7-[(m-halogeno)benzyloxy]coumarins bearing properly selected polar substituents at position 4 were designed, synthesized, and evaluated as MAO inhibitors. Several compounds with MAO-B inhibitory activity in the nanomolar range and excellent MAO-B selectivity (selectivity index SI > 400) were identified. Structure-affinity relationships and docking simulations provided valuable insights into the enzyme-inhibitor binding interactions at position 4, which has been poorly explored. Furthermore, computational and experimental studies led to the identification and biopharmacological characterization of 7-[(3-chlorobenzyl)oxy]-4-[(methylamino)methyl]-2H-chromen-2-one methanesulfonate 22b (NW-1772) as an in vitro and in vivo potent and selective MAO-B inhibitor, with rapid blood-brain barrier penetration, short-acting and reversible inhibitory activity, slight inhibition of selected cytochrome P450s, and low in vitro toxicity. On the basis of this preliminary preclinical profile, inhibitor 22b might be viewed as a promising clinical candidate for the treatment of neurodegenerative diseases.

A novel approach was developed to rationally interface structure- and ligand-based drug design through the rescoring of docking poses and automated generation of molecular alignments for 3D quantitative structure-activity relationship investigations. The procedure was driven by a genetic algorithm optimizing the value of a novel fitness function, accounting simultaneously for best regressions among binding-energy docking scores and affinities and for minimal geometric deviations from properly established crystal-based binding geometry. The GRID/CPCA method, as implemented in GOLPE, was used to feature molecular determinants of ligand binding affinity for each molecular alignment. In addition, unlike standard procedures, a novel multipoint equation was adopted to predict the binding affinity of ligands in the prediction set. Selectivity was investigated through square plots reporting experimental versus recalculated binding affinities on the targets under examination. The application of our approach to the modeling of affinity data of a large series of 3-amidinophenylalanine inhibitors of thrombin, trypsin, and factor Xa generated easily interpretable and independent models with robust statistics. As a further validation study, our approach was successfully applied to a series of 3,4,7-substituted coumarins, acting as selective MAO-B inhibitors.

A total of 142 matrix metalloproteinase (MMP) X-ray crystallographic structures were retrieved from the Protein Data Bank (PDB) and analyzed by an automated and efficient routine, developed in-house, with a series of bioinformatic tools. Highly informative heat maps and hierarchical clusterograms provided a reliable and comprehensive representation of the relationships existing among MMPs, enlarging and complementing the current knowledge in the field. Multiple sequence and structural alignments permitted better location and display of key MMP motifs and quantification of the residue consensus at each amino acid position in the most critical binding subsites of MMPs. The MMP active site consensus sequences, the C-alpha root-mean-square deviation (RMSd) analysis of diverse enzymatic subsites, and the examination of the chemical nature, binding topologies, and zinc binding groups (ZBGs) of ligands extracted from crystallographic complexes provided useful insights on the structural arrangements of the most potent MMP inhibitors.

A number of condensed azines, mostly belonging to the families of indeno-fused pyridazines (1), pyrimidines (2, 3), and 1,2,4-triazines (4, 5), were synthesized and evaluated in vitro as monoamine oxidase (MAO) A and B inhibitors. Most of them showed higher inhibition potency toward MAO-B, the most effective one being 3-(3-nitrophenyl)-9H-indeno[1,2-e] [1,2,4]triazin-9-one (4c), which displayed an IC50 value of 80 nM and proved to be 10-fold more potent than its [2,1-e] fusion isomer 5. Replacing the 3-phenyl group of the known indeno[1,2-c]pyridazin-5-one MAO-B inhibitors with a flexible phenoxymethyl group enhanced the inhibitory potency. The inhibition data highlighted the importance of the aza-heterocyclic scaffold in affecting the MAO isoform selectivity. The 3-phenyl derivatives with type 1, 4, and 5 scaffolds were inhibitors of MAO-B with little or no MAO-A effect, whereas 2- or 3-phenyl derivatives of type 2 and 3 pyrimidine-containing fusion isomers inhibited both isoenzymes with a structure-dependent preference toward MAO-A.

Safinamide, (S)-N2-{4-[(3-fluorobenzyl)oxy]benzyl}alaninamide methanesulfonate, which is in phase III clinical trials as an anti-Parkinson drug, and a library of alkanamidic analogues were prepared through an expeditious solid-phase synthesis and evaluated for their monoamine oxidase B (MAO-B) and monoamine oxidase A (MAO-A) inhibitory activity and selectivity. (S)-3-Chlorobenzyloxyalaninamide (8) and (S)-3-chlorobenzyloxyserinamide (13) derivatives proved to be more potent MAO-B inhibitors than safinamide (IC50 = 33 and 43 nM, respectively, vs 98 nM) but with a lower MAO-B selectivity (SI = 3455 and 1967, respectively, vs 5918). The highest MAO-B inhibitory potency (IC50 = 17 nM) and a good selectivity (SI = 2941) were displayed by (R)-21, a tetrahydroisoquinoline analogue of safinamide. Structure-affinity relationships and docking simulations pointed out strong negative steric effects of alpha-aminoamide side chains and para substituents of the benzyloxy groups and favorable hydrophobic interactions of meta substituents. The significantly diverse MAO-B affinities of a number of R and S alpha-aminoamide enantiomers, including the two rigid analogues (21) of safinamide, indicated likely enantioselective interactions at the enzymatic binding sites.

The impact of species-dependent differences between human and rat MAO B on inhibitor screening was evidenced for two classes of compounds, coumarin and 5H-indeno[1,2-c]pyridazin-5-one derivatives. All examined compounds have shown a greater inhibitor potency toward human MAO B than toward rat MAO B. Moreover, no correlation was found between human and rat pIC(50) values. These divergences have important implications for the design and development of drugs involved in the MAO B metabolic pathway, suggesting that results obtained using rat enzyme cannot be extrapolated to human CNS, a priori. Indeed, the selection of a hit compound for lead generation could be different using human rather than rat enzyme. Moreover, the influence of substituents on the in vitro inhibition of human MAO B was markedly different between homogeneous series of coumarin and 5H-indeno[1,2-c]pyridazin-5-one derivatives, suggesting different binding modes, a hypothesis clearly supported by molecular docking simulations of inhibitors into the active site of human MAO B.

A new series of 3-, 4-, 7-polysubstituted coumarins have been designed and evaluated for their monoamine oxidase A and monoamine oxidase B (MAO-A and MAO-B) inhibitory potency. Substituents at position 7 consisted of a bridge of different physicochemical nature linking a phenyl ring to the coumarin scaffold. Structure-affinity and structure-selectivity relationships, derived through CoMFA-GOLPE and docking studies, revealed the key physicochemical interactions responsible for the observed MAO-B and MAO-A inhibitory potency and suggested the main structural determinants for high selectivity toward one of the two enzymatic isoforms. The predictive power of our models was proved with the design of a new inhibitor demonstrating an outstanding MAO-B affinity (pIC50 = 8.29) and the highest MAO-B selectivity (DeltapIC50 = 3.39) within the entire series of ligands examined herein.

The design, synthesis, and biological evaluation of a series of new aromatase inhibitors bearing an imidazole or triazole ring linked to a fluorene (A), indenodiazine (B), or coumarin scaffold (C) are reported. Properly substituted coumarin derivatives displayed the highest aromatase inhibitory potency and selectivity over 17-alpha-hydroxylase/17-20 lyase. The modeling of the aromatase inhibition data by Comparative Molecular Field Analysis (CoMFA/GOLPE 3D QSAR approach) led to the development of a PLS model with good fitting and predictive powers (n = 22, ONC = 3, r(2) = 0.949, s = 0.216, and q(2) = 0.715). The relationship between aromatase inhibition and the steric and electrostatic fields generated by the examined azole inhibitors enables a clear understanding of the nature and spatial location of the main interactions modulating the aromatase inhibitory potency.

A highly efficient solid-phase synthesis method for the preparation of fluorogenic protease substrates based upon the bifunctional leaving group 7-amino-4-carbamoylmethylcoumarin (ACC) is reported. Methods for the large-scale preparation of the novel fluorogenic leaving-group ACC are provided (Scheme 1). Detailed procedures are also provided for loading a diverse set of amino acids to support-bound ACC in good yields and with minimal racemization. Finally, procedures are included for the preparative synthesis of optimized ACC substrates for HIV-1 protease and plasmin.

A set of 17 coumarin and 2 chromone derivatives with known inhibitory activity toward monoamine oxidase (MAO) A and B were tested as acetylcholinesterase (AChE) inhibitors. All compounds inhibited AChE with values in the micromolar range (3-100 microM). A kinetic study showed that most compounds acted as noncompetitive AChE inhibitors. This finding may be of interest in the context of Alzheimer's disease because recent observations suggest that MAO and AChE inhibition might decrease beta-amyloid deposition.

A large series of coumarin derivatives (71 compounds) were tested for their monoamine oxidase A and B (MAO-A and MAO-B) inhibitory activity. Most of the compounds acted preferentially on MAO-B with IC(50) values in the micromolar to low-nanomolar range; high inhibitory activities toward MAO-A were also measured for sulfonic acid esters. The most active compound was 7-[(3, 4-difluorobenzyl)oxy]-3,4-dimethylcoumarin, with an IC(50) value toward MAO-B of 1.14 nM. A QSAR study of 7-X-benzyloxy meta-substituted 3,4-dimethylcoumarin derivatives acting on MAO-B yielded good statistical results (q(2)() = 0.72, r(2)() = 0.86), revealing the importance of lipophilic interactions in modulating the inhibition and excluding any dependence on electronic properties. CoMFA was performed on two data sets of MAO-A and MAO-B inhibitors. The GOLPE procedure, with variable selection criteria, was applied to improve the predictivity of the models and to facilitate the graphical interpretation of results.