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The glycogen synthase kinase 3β (GSK-3β) is a ubiquitous enzyme that is a validated target for the development of potential therapeutics useful in several diseases including retinal degeneration. Aiming at developing an innovative class of allosteric inhibitors of GSK-3β potentially useful for retinal degeneration, we explored the class of squaramides. The developed compounds (6 a-l) were obtained through a nontoxic one-pot synthetic protocol, which employs low-cost goods and avoids any purification step. Ethanol was used as the reaction solvent, simultaneously allowing the pure reaction products' recovery (by precipitation). Out of this set of squaramides, 6 j stood out, from computational and enzymatic converging data, as an ATP non-competitive inhibitor of GSK-3β of micromolar potency. When engaged in cellular studies using retinal pigment epithelial cells (ARPE-19) transfected with a luciferase reporter gene under the control of T-cell factor/lymphoid enhancer factor (TCF/LEF) binding sites, 6 j was able to dose-dependently induce β-catenin nuclear accumulation, as shown by the increased luciferase activity at a concentration of 2.5 μM.

The neuroprotective performance against neuroinflammation of the endocannabinoid system (ECS) can be remarkably improved by indirect stimulation mediated by the pharmacological inhibition of the key ECS catabolic enzyme fatty acid amide hydrolase (FAAH). Based on our previous works and aiming to discover new selective FAAH inhibitors , we herein reported a new series of carbamate-based FAAH inhibitors (4a-t) which showed improved drug disposition properties compared to the previously reported analogues 2a-b. The introduction of ionizable functions allowed us to obtain new FAAH inhibitors of nanomolar potency characterized by good water solubility and chemical stability at physiological pH. Interesting structure-activity relationships (SARs), deeply analyzed by molecular docking and molecular dynamic (MD) simulations, were obtained. All the newly developed inhibitors showed an excellent selectivity profile evaluated against monoacylglycerol lipase and cannabinoid receptors. The reversible mechanism of action was determined by a rapid dilution assay. Absence of toxicity was confirmed in mouse fibroblasts NIH3T3 (for compounds 4e, 4g, 4n-o, and 4s) and in human astrocytes cell line 1321N1 (for compounds 4e, 4n, and 4s). The absence of undesired cardiac effects was also confirmed for compound 4n. Selected analogues (compounds 4e, 4g, 4n, and 4s) were able to reduce oxidative stress in 1321N1 astrocytes and exhibited notable neuroprotective effects when tested in an ex vivo model of neuroinflammation.

Inherited retinal diseases (IRDs) are a group of retinopathies generally caused by genetic mutations. Retinitis pigmentosa (RP) represents one of the most studied IRDs. RP leads to intense vision loss or blindness resulting from the degeneration of photoreceptor cells. To date, RP is mainly treated with palliative supplementation of vitamin A and retinoids, gene therapies, or surgical interventions. Therefore, a pharmacologically based therapy is an urgent need requiring a medicinal chemistry approach, to validate molecular targets able to deal with retinal degeneration. This Review aims at outlining the recent research efforts in identifying new drug targets for RP, especially focusing on the neuroprotective role of the Wnt/β-catenin/GSK3β pathway and apoptosis modulators (in particular PARP-1) but also on growth factors such as VEGF and BDNF. Furthermore, the role of spatiotemporally expressed G protein-coupled receptors (GPR124) in the retina and the emerging function of histone deacetylase inhibitors in promoting retinal neuroprotection will be discussed.

(S)-2-Amino-3-(5-methyl-3-hydroxyisoxazol-4-yl)propanoic acid (AMPA) receptors comprise an important class of ionotropic glutamate receptors activated by glutamate in the central nervous system. Th...

In the field of drug discovery, the nitrile group is well represented among drugs and biologically active compounds. It can form both non-covalent and covalent interactions with diverse biological targets, and it is amenable as an electrophilic warhead for covalent inhibition. The main advantage of the nitrile group as a warhead is mainly due to its milder electrophilic character relative to other more reactive groups (e.g., -CHO), reducing the possibility of unwanted reactions that would hinder the development of safe drugs, coupled to the ease of installation through different synthetic approaches. The covalent inhibition is a well-assessed design approach for serine, threonine, and cysteine protease inhibitors. The mechanism of hydrolysis of these enzymes involves the formation of a covalent acyl intermediate, and this mechanism can be exploited by introducing electrophilic warheads in order to mimic this covalent intermediate. Due to the relevant role played by the cysteine protease in the survival and replication of infective agents, spanning from viruses to protozoan parasites, we will review the most relevant and recent examples of protease inhibitors presenting a nitrile group that have been introduced to form or to facilitate the formation of a covalent bond with the catalytic cysteine active site residue.

96 p.-20 fig.-7 tab., Autophagy is a lysosome dependent cell survival mechanism and is central to the maintenance of organismal homeostasis in both physiological and pathological situations. Targeting autophagy in cancer therapy attracted considerable attention in the past as stress-induced autophagy has been demonstrated to contribute to both drug resistance and malignant progression and recently interest in this area has re-emerged. Unlocking the therapeutic potential of autophagy modulation could be a valuable strategy for designing innovative tools for cancer treatment. Microtubule-targeting agents (MTAs) are some of the most successful anti-cancer drugs used in the clinic to date. Scaling up our efforts to develop new anti-cancer agents, we rationally designed multifunctional agents 5a-l with improved potency and safety that combine tubulin depolymerising efficacy with autophagic flux inhibitory activity. Through a combination of computational, biological, biochemical, pharmacokinetic-safety, metabolic studies and SAR analyses we identified the hits 5i,k. These MTAs were characterised as potent pro-apoptotic agents and also demonstrated autophagy inhibition efficacy. To measure their efficacy at inhibiting autophagy, we investigated their effects on basal and starvation-mediated autophagic flux by quantifying the expression of LC3II/LC3I and p62 proteins in oral squamous cell carcinoma and human leukaemia through western blotting and by immunofluorescence study of LC3 and LAMP1 in a cervical carcinoma cell line. Analogues 5i and 5k, endowed with pro-apoptotic activity on a range of hematological cancer cells (including ex-vivo chronic lymphocytic leukaemia (CLL) cells) and several solid tumor cell lines, also behaved as late-stage autophagy inhibitors by impairing autophagosome-lysosome fusion., This research was funded by the European Union's Horizon 2020 (EU) Research and Innovation Programme under the Marie Sklodowska-Curie grant agreement No.721906-TRACT (TK, SM, MG, NMcC, GO’S, RT, VK, SG, DZ, GC, SB); Tuscany strategic project POR-FSE 2014-2020, ‘Medicina di Precisione e Malattie Rare’(MePreMaRe), (ACE-ESCC) (NR, GC, SB), and by “Epigenetic Hallmarks of Multiple Sclerosis” (acronym Epi-MS) (id:415, Merit Ranking Area ERC LS) in VALERE 2019 Program; V:ALERE 2020 – Progetto competitivo “CIRCE” in risposta al bando D.R. n. 138 del February 17, 2020 Program; Blueprint 282510; EPICHEMBIO CM1406; AIRC-17217; VALERE: Vanvitelli per la Ricerca; Campania Regional Government Technology Platform Lotta alle Patologie Oncologiche: iCURE; Campania Regional Government FASE2: IDEAL. MIUR, Proof of Concept POC01_00043. POR Campania FSE 2014-2020 ASSE III.

The potential of natural and synthetic chalcones as therapeutic leads against different pathological conditions has been investigated for several years, and this class of compounds emerged as a privileged chemotype due to its interesting anti-inflammatory, antimicrobial, antiviral, and anticancer properties. The objective of our study was to contribute to the investigation of this class of natural products as anti-leishmanial agents. We aimed at investigating the structure–activity relationships of the natural chalcone lophirone E, characterized by the presence of benzofuran B-ring, and analogues on anti-leishmania activity. Here we describe an effective synthetic strategy for the preparation of the natural chalcone lophirone E and its application to the synthesis of a small set of chalcones bearing different substitution patterns at both the A and heterocyclic B rings. The resulting compounds were investigated for their activity against Leishmania infantum promastigotes disclosing derivatives 1 and 28a,b as those endowed with the most interesting activities (IC50 = 15.3, 27.2, 15.9 μM, respectively). The synthetic approaches here described and the early SAR investigations highlighted the potential of this class of compounds as antiparasitic hits, making this study worthy of further investigation.

Despite the progress of therapeutic approaches for treating COVID-19 infection, the interest in developing effective antiviral agents is still high, due to the possibility of the insurgence of viable SARS-CoV-2-resistant strains. Accordingly, in this article, we describe a computational protocol for identifying possible SARS-CoV-2 Mpro covalent inhibitors. Combining several in silico techniques, we evaluated the potential of the peptide-based scaffold with different warheads as a significant alternative to nitriles and aldehyde electrophilic groups. We rationally designed four potential inhibitors containing difluorstatone and a Michael acceptor as warheads. In silico analysis, based on molecular docking, covalent docking, molecular dynamics simulation, and FEP, indicated that the conceived compounds could act as covalent inhibitors of Mpro and that the investigated warheads can be used for designing covalent inhibitors against serine or cysteine proteases such as SARS-CoV-2 Mpro. Our work enriches the knowledge on SARS-CoV-2 Mpro, providing a novel potential strategy for its inhibition, paving the way for the development of effective antivirals.

Pin1 catalyzes the cis-trans isomerization of pThr-Pro or pSer-Pro amide bonds of various proteins involved in several physio/pathological processes. In this framework, recent research activity is directed toward the identification of new selective Pin1 inhibitors. Here, we developed a set of peptide-based Pin1 inhibitors. Direct-binding experiments allowed the identification of the peptide-based inhibitor 5 k (methylacetyl-l-alanyl-l-histidyl-l-prolyl-l-phenylalaninate) as a potent ligand of Pin1. Notably, 5 k binds Pin1 with higher affinity than Pin4. The comparative analysis of molecular models of Pin1 and Pin4 with the selected compound gave a rational explanation of the biochemical activity and pinpointed the chemical elements that, if opportunely modified, may further improve inhibitory potency, pharmacological properties, and selectivity of future peptide-based parvulin inhibitors. Since 5 k showed limited cell penetration and no antiproliferative activity, it was conjugated to a polyarginine stretch (R8), known to promote cell penetration of peptides, to obtain the R8-5 k derivative, which displayed antiproliferative effects on cancer cell lines over non-tumor cells. The effect of R8 on cell proliferation was also investigated. This work warrants caution about applying the R8 strategy in the development of cell-penetrating antiproliferative peptides, as it is not inert.

Parasitic diseases cause significant global morbidity and mortality particularly in the poorest regions of the world. Schistosomiasis, one of the most widespread neglected tropical diseases, affects more than 200 million people worldwide. Histone deacetylase (HDAC) inhibitors are prominent epigenetic drugs that are being investigated in the treatment of several diseases, including cancers and parasitic diseases. Schistosoma mansoni HDAC8 (SmHDAC8) is highly expressed in all life cycle stages of the parasite, and selective inhibition is required in order to avoid undesirable off-target effects in the host. Herein, by X-ray crystal structures of SmHDAC8-inhibitor complexes, biochemical and phenotypic studies, we found two schistosomicidal spiroindoline derivatives binding a novel site, next to Trp198, on the enzyme surface. We determined that by acting on this site, either by mutation of the Trp198 or by compound binding, a decrease in the activity of the enzyme is achieved. Remarkably, this allosteric site differs from the human counterpart; rather, it is conserved in all Schistosoma species, as well as Rhabidoptera and Trematoda classes, thus paving the way for the design of HDAC8-selective allosteric inhibitors with improved properties.

Oral squamous cell carcinomas (OSCC) and esophageal squamous cell carcinomas (ESCC) exhibit a survival rate of less than 60% and 40%, respectively. Late-stage diagnosis and lack of effective treatment strategies make both OSCC and ESCC a significant health burden. Autophagy, a lysosome-dependent catabolic process, involves the degradation of intracellular components to maintain cell homeostasis. Targeting autophagy has been highlighted as a feasible therapeutic strategy with clinical utility in cancer treatment, although its associated regulatory mechanisms remain elusive. The detection of relevant biomarkers in biological fluids has been anticipated to facilitate early diagnosis and/or prognosis for these tumors. In this context, recent studies have indicated the presence of specific proteins and small RNAs, detectable in circulating plasma and serum, as biomarkers. Interestingly, the interplay between biomarkers (eg, exosomal microRNAs) and autophagic processes could be exploited in the quest for targeted and more effective therapies for OSCC and ESCC. In this review, we give an overview of the available biomarkers and innovative targeted therapeutic strategies, including the application of autophagy modulators in OSCC and ESCC. Additionally, we provide a viewpoint on the state of the art and on future therapeutic perspectives combining the early detection of relevant biomarkers with drug discovery for the treatment of OSCC and ESCC.

Due to the surge in resistance to common therapies, malaria remains a significant concern to human health worldwide. In chloroquine (CQ)-resistant (CQ-R) strains of Plasmodium falciparum, CQ and related drugs are effluxed from the parasite’s digestive vacuole (DV). This process is mediated by mutant isoforms of a protein called CQ resistance transporter (PfCRT). CQ-R strains can be partially re-sensitized to CQ by verapamil (VP), primaquine (PQ) and other compounds, and this has been shown to be due to the ability of these molecules to inhibit drug transport via PfCRT. We have previously developed a series of clotrimazole (CLT)-based antimalarial agents that possess inhibitory activity against PfCRT (4a,b). In our endeavor to develop novel PfCRT inhibitors, and to perform a structure-activity relationship analysis, we synthesized a new library of analogues. When the benzhydryl system was linked to a 4-aminoquinoline group (5a-f) the resulting compounds exhibited good cytotoxicity against both CQ-R and CQ-S strains of P. falciparum. The most potent inhibitory activity against the PfCRT-mediated transport of CQ was obtained with compound 5k. When compared to the reference compound, benzhydryl analogues of PQ (5i,j) showed a similar activity against blood-stage parasites, and a stronger in vitro potency against liver-stage parasites. Unfortunately, in the in vivo transmission blocking assays, 5i,j were inactive against gametocytes.

Temporal lobe epilepsy is the most common form of epilepsy, and current antiepileptic drugs are ineffective in many patients. The endocannabinoid system has been associated with an on-demand protective response to seizures. Blocking endocannabinoid catabolism would elicit antiepileptic effects, devoid of psychotropic effects. We herein report the discovery of selective anandamide catabolic enzyme fatty acid amide hydrolase (FAAH) inhibitors with promising antiepileptic efficacy, starting from a further investigation of our prototypical inhibitor2a. When tested in two rodent models of epilepsy,2areduced the severity of the pilocarpine-induced status epilepticus and the elongation of the hippocampal maximal dentate activation. Notably,2adid not affect hippocampal dentate gyrus long-term synaptic plasticity. These data prompted our further endeavor aiming at discovering new antiepileptic agents, developing a new set of FAAH inhibitors (3a-m). Biological studies highlighted3hand3mas the best performing analogues to be further investigated. In cell-based studies, using a neuroblastoma cell line,3hand3mcould reduce the oxinflammation state by decreasing DNA-binding activity of NF-kB p65, devoid of cytotoxic effect. Unwanted cardiac effects were excluded for3h(Langendorff perfused rat heart). Finally, the new analogue3hreduced the severity of the pilocarpine-induced status epilepticus as observed for2a.

[Image: see text] Histone deacetylase inhibitors (HDACi) have emerged as promising therapeutics for the treatment of neurodegeneration, cancer, and rare disorders. Herein, we report the development of a series of spiroindoline-based HDAC6 isoform-selective inhibitors based on the X-ray crystal studies of the hit 6a. We identified compound 6j as the most potent and selective hHDAC6 inhibitor of the series. Biological investigation of compounds 6b, 6h, and 6j demonstrated their antiproliferative activity against several cancer cell lines. Western blotting studies indicated that they were able to increase tubulin acetylation, without significant variation in histone acetylation state, and induced PARP cleavage indicating their apoptotic potential at the molecular level. 6j induced HDAC6-dependent pSTAT3 inhibition.

Arginase is a metalloenzyme that plays a central role in Leishmania infection. Previously, rosmarinic and caffeic acids were described as antileishmanial and as a Leishmania amazonensis arginase inhibitor and now, we describe the inhibition of arginase in L. amazonensis by rosmarinic acid analogs (1-7) and new caffeic acids derived amides (8-10). Caffeic acid esters and amides were produced by means of the engineered synthesis in E. coli and tested against L. amazonensis arginase. New amides (8-10) were biosynthesized in E. coli cultured with 2 mM of different combinations of feeding substrates. The most potent arginase inhibitors showed Ki(s) between 2 - 5.7 μM. Compounds 2-4 and 7 inhibited L-ARG through a noncompetitive mechanism, and 9 showed a competitive inhibition. By applying an in silico protocol we determined the binding mode of compound 9. The competitive inhibitor of L-ARG targets key residues within the binding site of the enzyme establishing a metal coordination bond with the metal ions as well as a series of hydrophobic and polar contacts supporting its micromolar inhibition of L-ARG. These results highlight that the dihydroxycinnamic-derived compounds can be used as the basis for the development of new drugs using a powerful tool based on the biosynthesis of arginase inhibitors.

Toll-like receptors (TLRs) are a class of proteins that recognize pathogen-associated molecular patterns (PAMPs) and damaged-associated molecular patterns (DAMPs), and they are involved in the regulation of innate immune system. These transmembrane receptors, localized at the cellular or endosomal membrane, trigger inflammatory processes through either myeloid differentiation primary response 88 (MyD88) or TIR-domain-containing adapter-inducing interferon-β (TRIF) signaling pathways. In the last decades, extensive research has been performed on TLR modulators and their therapeutic implication under several pathological conditions, spanning from infections to cancer, from metabolic disorders to neurodegeneration and autoimmune diseases. This Perspective will highlight the recent discoveries in this field, emphasizing the role of TLRs in different diseases and the therapeutic effect of their natural and synthetic modulators, and it will discuss insights for the future exploitation of TLR modulators in human health.

Schistosomiasis is a neglected tropical disease mainly affecting the poorest tropical and subtropical areas of the world with the impressive number of roughly 200 million infections per year. Schistosomes are blood trematode flukes of the genus Schistosoma causing symptoms in humans and animals. Organ morbidity is caused by the accumulation of parasite eggs and subsequent development of fibrosis. If left untreated, schistosomiasis can result in substantial morbidity and even mortality. Praziquantel (PZQ) is the most effective and widely used compound for the treatment of the disease, in prevention and control programs in the last 30 years. Unfortunately, it has no effect on juvenile immature schistosomes and cannot prevent reinfection or interfere with the schistosome life cycle; moreover drug-resistance represents a serious threat. The search for an alternative or complementary treatment is urgent and drug repurposing could accelerate a solution. The anti-anginal drug perhexiline maleate (PHX) has been previously shown to be effective on larval, juvenile, and adult stages of S. mansoni and to impact egg production in vitro. Since PHX is a racemic mixture of R-(+)- and S-(−)-enantiomers, we designed and realized a stereoselective synthesis of both PHX enantiomers and developed an analytical procedure for the direct quantification of the enantiomeric excess also suitable for semipreparative separation of PHX enantiomers. We next investigated the impact of each enantiomer on viability of newly transformed schistosomula (NTS) and worm pairs of S. mansoni as well as on egg production and vitellarium morphology by in vitro studies. Our results indicate that the R-(+)-PHX is mainly driving the anti-schistosomal activity but that also the S-(−)-PHX possesses a significant activity towards S. mansoni in vitro.

Epigenetic regulation orchestrates many cellular processes and greatly influences key disease mechanisms. Histone deacetylase (HDAC) enzymes play a crucial role either as biomarkers or therapeutic targets owing to their involvement in specific pathophysiological pathways. Beyond their well-characterized role as histone modifiers, HDACs also interact with several nonhistone substrates and their increased expression has been highlighted in specific diseases. The HDAC6 isoform, due to its unique cytoplasmic localization, modulates the acetylation status of tubulin, HSP90, TGF-β, and peroxiredoxins. HDAC6 also exerts noncatalytic activities through its interaction with ubiquitin. Both catalytic and noncatalytic functions of HDACs are being actively studied in the field of specific rare disorders beyond the well-established role in carcinogenesis. This Perspective outlines the application of HDAC(6) inhibitors in rare diseases, such as Rett syndrome, inherited retinal disorders, idiopathic pulmonary fibrosis, and Charcot-Marie-Tooth disease, highlighting their therapeutic potential as innovative and targeted disease-modifying agents.

The fatty acid amide hydrolase (FAAH) executes a unique role in terminating endocannabinoid signaling, and is the major regulator of anadamide catabolism in vivo. Inhibition of FAAH elicits indirect agonism on cannabinoid receptors, and therapeutic efficacy, devoid of psychotropic effects. This supports its relevance as an attractive therapeutic target. Great efforts in the medicinal chemistry, biochemistry, and crystallography fields have synergistically provided pivotal information key to the production of a number of inhibitors, characterized by different structures and mechanisms of action, for attaining FAAH inhibition. As the knowledge about the enzyme structure and functioning grew, in the past two decades, the developed compounds became more specific and structurally different from the natural substrates, thus allowing better selectivity and inhibition potency to be attained. This was pivotal for the identification of leads and druggable compounds to be clinically exploited for the treatment of a number of diseases. The lesson learned by the tragic outcome of the clinical studies with BIA 10-2474 have drawn a clear trajectory for pre-clinical assessment of off-target liabilities, while offering a chance to clearly outline the safety of FAAH as a drug target. A number of natural compounds from mammalian and vegetal sources have been identified so far and have been characterized as FAAH inhibitors and modulators. These compounds may inspire the design of further effective FAAH inhibitors that encompass the enzyme physiological functioning and enzyme trafficking. This issue may offer a new view that considers the crucial events that regulate substrates and compounds accessibility to the FAAH catalytic site and that may affect in vivo efficacy.

Telomeres are protective chromosomal ends that shield the chromosomes from DNA damage, exonucleolytic degradation, recombination, and end-to-end fusion. Telomerase is a ribonucleoprotein that adds TTAGGG tandem repeats to the telomeric ends. It has been observed that 85 to 90% of human tumors express high levels of telomerase, playing a crucial role in the development of cancers. Interestingly, the telomerase activity is generally absent in normal somatic cells. This selective telomerase expression has driven scientists to develop novel anti-cancer therapeutics with high specificity and potency. Several advancements have been made in this area, which is reflected by the enormous success of the anticancer agent Imetelstat. Since the discovery of Imetelstat, several research groups have contributed to enrich the therapeutic arsenal against cancer. Such contributions include the application of new classes of small molecules, peptides, and hTERT-based immunotherapeutic agents (p540, GV1001, GRNVAC1 or combinations of these such as Vx-001). Many of these therapeutic tools are under different stages of clinical trials and have shown promising outcomes. In this review, we highlight the current status of telomerase-based cancer therapeutics and the outcome of these investigations.

This study describes the activity of five natural hydroxycinnamic acids and derived compound: caffeic (1), rosmarinic (2), chlorogenic (3), and cryptochlorogenic (4), acids and isoverbascoside (5). All compounds inhibited Leishmania amazonensis arginase with IC50 -in range of 1.5-11 μM. Compounds 2 and 5 also showed activity against promastigotes of L. amazonensis with IC50 = 61 (28-133) μM and IC50 = 14 (9-24) μM, respectively. Further computational studies applying molecular docking simulations were performed on the competitive inhibitors to gain insight into the molecular basis for arginase inhibition and could be exploited to the development of new antileishmanials drug targeting parasite arginase.

Therapies addressing multiple stages of Plasmodium falciparum life cycle are highly desirable for implementing malaria elimination strategies. MMV019918 (1, 1-[5-(4-bromo-2-chlorophenyl)furan-2-yl]-N-[(piperidin-4-yl)methyl]methanamine) was selected from the MMV Malaria Box for its dual activity against both asexual stages and gametocytes. In-depth structure-activity relationship studies and cytotoxicity evaluation led to the selection of 25 for further biological investigation. The potential transmission blocking activity of 25 versus P. falciparum was confirmed through the standard membrane-feeding assay. Both 1 and 25 significantly prolonged atrioventricular conduction time in Langendorff-isolated rat hearts, and showed inhibitory activity of Ba(2+) current through Ca(v)1.2 channels. An in silico target-fishing study suggested the enzyme phosphoethanolamine methyltransferase (PfPMT) as a potential target. However, compound activity against PfPMT did not track with the antiplasmodial activity, suggesting the latter activity relies on a different molecular target. Nevertheless, 25 showed interesting activity against PfPMT, which could be an important starting point for the identification of more potent inhibitors active against both sexual and asexual stages of the parasite.

Summary Aims We recently described multifunctional tools (2a–c) as potent inhibitors of human Cholinesterases (ChEs) also able to modulate events correlated with Aβ aggregation. We herein propose a thorough biological and computational analysis aiming at understanding their mechanism of action at the molecular level. Methods We determined the inhibitory potency of 2a–c on Aβ1–42 self-aggregation, the interference of 2a with the toxic Aβ oligomeric species and with the postaggregation states by capillary electrophoresis analysis and transmission electron microscopy. The modulation of Aβ toxicity was assessed for 2a and 2b on human neuroblastoma cells. The key interactions of 2a with Aβ and with the Aβ-preformed fibrils were computationally analyzed. 2a–c toxicity profile was also assessed (human hepatocytes and mouse fibroblasts). Results Our prototypical pluripotent analogue 2a interferes with Aβ oligomerization process thus reducing Aβ oligomers-mediated toxicity in human neuroblastoma cells. 2a also disrupts preformed fibrils. Computational studies highlighted the bases governing the diversified activities of 2a. Conclusion Converging analytical, biological, and in silico data explained the mechanism of action of 2a on Aβ1–42 oligomers formation and against Aβ-preformed fibrils. This evidence, combined with toxicity data, will orient the future design of safer analogues.

Prion diseases are serious, not curable neurodegenerative disorders caused by the accumulation of the misfolded protein PrP(Sc) that represents the pathological variant of the normally folded cellular protein PrP(C). Molecules that bind the cellular isoform PrP(C) preventing its misfolding, could arrest the progression of pathological conditions related to the abnormal PrP protein. In this context, by combining 3D-QSAR model, derived from pharmacophore-based alignment, with molecular docking procedures and physico-chemical properties prediction we have developed a virtual screening protocol to find novel chemicals able to prevent PrP(C) misfolding. We identified different hits characterized by low toxicity and able to inhibit PrP(Sc) accumulation in vitro in prion-infected neuroblastoma cell lines (ScN2a). In this assay, the pyrroloquinoxaline hydrazone 96 showed the higest potency with an IC50 value of 1.6 μM. Pyrroloquinoxaline 96 was demonstrated also to bind PrP(Sc) aggregates in infected ScN2a cells with a fluorescence pattern comparable to that found for Thioflavin-T. In consideration of its satisfactory physico-chemical properties, including predicted blood brain barrier permeability, 96 could represent an interesting prototypic hit for the development of diagnostic and therapeutic probes for prion diseases.

Tuberculosis (TB) is one of the top 10 causes of death worldwide. This scenario is further complicated by the insurgence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) TB. The identification of appropriate drugs with multi-target affinity profiles is considered to be a widely accepted strategy to overcome the rapid development of resistance. The aim of this study was to discover Food and Drug Administration (FDA)-approved drugs possessing antimycobacterial activity, potentially coupled to an effective multi-target profile. An integrated screening platform was implemented based on computational procedures (high-throughput docking techniques on the target enzymes peptide deformylase and Zmp1) and in vitro phenotypic screening assays using two models to evaluate the activity of the selected drugs against Mycobacterium tuberculosis (Mtb), namely, growth of Mtb H37Rv and of two clinical isolates in axenic media, and infection of peripheral blood mononuclear cells with Mtb. Starting from over 3000 FDA-approved drugs, we selected 29 marketed drugs for submission to biological evaluation. Out of 29 drugs selected, 20 showed antimycobacterial activity. Further characterization suggested that five drugs possessed promising profiles for further studies. Following a repurposing strategy, by combining computational and biological efforts, we identified marketed drugs with relevant antimycobacterial profiles.

Schistosomiasis (also known as bilharzia) is a neglected tropical disease caused by platyhelminths of the genus

We describe herein the development and experimental validation of a computational protocol for optimizing a series of 3-hydroxy-pyran-4-one derivatives as HIV integrase inhibitors (HIV INIs). Starting from a previously developed micromolar inhibitors of HIV integrase (HIV IN), we performed an in-depth investigation based on an in silico structure-based combinatorial library designing approach. This method allowed us to combine a combinatorial library design and side chain hopping with Quantum Polarized Ligand Docking (QPLD) studies and Molecular Dynamics (MD) simulation. The combinatorial library design allowed the identification of the best decorations for our promising scaffold. The resulting compounds were assessed by the mentioned QPLD methodology using a homology model of full-length binary HIV IN/DNA for retrieving the best performing compounds acting as HIV INIs. Along with the prediction of physico-chemical properties, we were able to select a limited number of drug-like compounds potentially displaying potent HIV IN inhibition. From this final set, based on the synthetic accessibility, we further shortlisted three representative compounds for the synthesis. The compounds were experimentally assessed in vitro for evaluating overall HIV-1 IN inhibition, HIV-1 IN strand transfer activity inhibition, HIV-1 activity inhibition and cellular toxicity. Gratifyingly, all of them showed relevant inhibitory activity in the in vitro tests along with no toxicity. Among them HPCAR-28 represents the most promising compound as potential anti-HIV agent, showing inhibitory activity against HIV IN in the low nanomolar range, comparable to that found for Raltegravir, and relevant potency in inhibiting HIV-1 replication and HIV-1 IN strand transfer activity. In summary, our results outline HPCAR-28 as a useful optimized hit for the potential treatment of HIV-1 infection by targeting HIV IN. ispartof: FRONTIERS IN CHEMISTRY vol:7 ispartof: location:Switzerland status: published

Despite recent advancements in its control, malaria is still a deadly parasitic disease killing millions of people each year. Progresses in combating the infection have been made by using the so-called artemisinin combination therapies (ACTs). Natural and synthetic peroxides are an important class of antimalarials. Here we describe a new series of peroxides synthesized through a new elaboration of the scaffold of bicyclic-fused/bridged synthetic endoperoxides previously developed by us. These peroxides are produced by a straightforward synthetic protocol and are characterized by submicromolar potency when tested against both chloroquine-sensitive and chloroquine-resistant Plasmodium falciparum strains. To investigate their mode of action, the biomimetic reaction of the representative compound 6w with Fe(II) was studied by EPR and the reaction products were characterized by NMR. Rationalization of the observed structure-activity relationship studies was performed by molecular docking. Taken together, our data robustly support the hypothesized mode of activation of peroxides 6a-cc and led to the definition of the key structural requirements responsible for the antiplasmodial potency. These data will pave the way in future to the rational design of novel optimized antimalarials suitable for in vivo investigation.

Telomerase is a ribonucleic reverse transcriptase enzyme that uses an integral RNA component as a template to add tandem telomeric DNA repeats, TTAGGG, at the 3' end of the chromosomes. 85-90% of human tumors and their derived cell lines predominantly express high levels of telomerase, therefore contributing to cancer cell development. However, in normal cells, telomerase activity is almost always absent except in germ cells and stem cells. This differential expression has been exploited to develop highly specific and potent cancer therapeutics. In this review, we outline recent advances in the development of telomerase inhibitors as anticancer agents.

Quercetin related compounds were tested against Leishmania amazonensis arginase, a potential target for the development of new approaches in treating leishmaniasis. The IC50 and kinetic analysis were performed to determine the dissociation constant Ki and the inhibition mechanism of the parasite's arginase enzyme. The best arginase inhibition was obtained from taxifolin (dihydroquercetin) with IC50 = 1.6 ± 0.1 μM. This study showed for the first time that rutin (IC50 = 10.4 ± 0.8 μM), and human metabolite quercetin-3-O-glucuronide (IC50 = 8.2 ± 0.4 μM), target L. amazonensis arginase. In addition, computational studies applying molecular docking simulations were performed to gain insight into the molecular basis for arginase inhibition by the competitive inhibitors. Our results suggest that these compounds could be exploited to develop new approaches for treating leishmaniasis through molecular nutrition supplement in a drug-based therapy.