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O 6 -Methylguanine-DNA-methyltransferase (MGMT) is a key DNA repair enzyme involved in chemoresistance to DNA-alkylating anti-cancer drugs such as Temozolomide (TMZ) through direct repair of drug-induced O 6 -methylguanine residues in DNA. MGMT substrate analogues, such as O 6 -benzylguanine (BG), efficiently inactivate MGMT in vitro and in cells; however, these drugs failed to reach the clinic due to adverse side effects. Here, we designed hybrid drugs combining a BG residue covalently linked to a DNA-interacting moiety (6-chloro-2-methoxy-9-aminoacridine). Specifically, two series of hybrids, encompassing three compounds each, were obtained by varying the position of the attachment point of BG (N 9 of guanine vs. the benzyl group) and the length and nature of the linker. UV/vis absorption and fluorescence data indicate that all six hybrids adopt an intramolecularly stacked conformation in aqueous solutions in a wide range of temperatures. All hybrids interact with double-stranded DNA, as clearly evidenced by spectrophotometric titrations, without intercalation of the acridine ring and do not induce thermal stabilization of the duplex. All hybrids, as well as the reference DNA intercalator (6-chloro-2-methoxy-9-aminoacridine 8), irreversibly inhibit MGMT in vitro with variable efficiency, comparable to that of BG. In a multidrug-resistant glioblastoma cell line T98G, benzyl-linked hybrids 7a-c and the N 9 -linked hybrid 19b are moderately cytotoxic (GI 50  ≥ 15 μM after 96 h), while N 9 -linked hybrids 19a and 19c are strongly cytotoxic (GI 50  = 1-2 μM), similarly to acridine 8 (GI 50  = 0.6 μM). Among all compounds, hybrids 19a and 19c, similarly to BG, display synergic cytotoxic effect upon co-treatment with subtoxic doses of TMZ, with combination index (CI) values as low as 0.2-0.3. In agreement with in vitro results, compound 19a inactivates cellular MGMT but, unlike BG, does not induce significant levels of DNA damage, either alone or in combination with TMZ, as indicated by the results of γH2AX immunostaining experiments. Instead, and unlike BG, compound 19a alone induces significant apoptosis of T98G cells, which is not further increased in a combination with TMZ. These results indicate that molecular mechanisms underlying the cytotoxicity of 19a and its combination with TMZ are distinct from that of BG. The strongly synergic properties of this combination represent an interesting therapeutic opportunity in treating TMZ-resistant cancers., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Masson SAS. All rights reserved.)

Cancer multi-drug resistance (MDR) caused by P-glycoprotein (P-gp) efflux is a critical unresolved clinical concern. The present study analyzed the effect of cinnamophilin on P-gp inhibition and MDR reversion. The effect of cinnamophilin on P-gp was investigated through drug efflux assay, ATPase assay, MDR1 shift assay, and molecular docking. The cancer MDR-reversing ability and mechanisms were analyzed through cytotoxicity and combination index (CI), cell cycle, and apoptosis experiments. P-gp efflux function was significantly inhibited by cinnamophilin without influencing the drug's expression or conformation. Cinnamophilin uncompetitively inhibited the efflux of doxorubicin and rhodamine 123 and exhibited a distinct binding behavior compared with verapamil, the P-gp standard inhibitor. The half maximal inhibitory concentration of cinnamophilin for doxorubicin and rhodamine 123 efflux was 12.47 and 11.59 μM, respectively. In regard to P-gp energy consumption, verapamil-stimulated ATPase activity was further enhanced by cinnamophilin at concentrations of 0.1, 1, 10, and 20 μM. In terms of MDR reversion, cinnamophilin demonstrated synergistic cytotoxic effects when combined with docetaxel, vincristine, or paclitaxel. The CI was < 0.7 in all experimental combination treatments. The present study showed that cinnamophilin possesses P-gp-modulating effects and cancer MDR resensitizing ability., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Ricin toxin A subunit (RTA) is the catalytic subunit of ricin, which depurinates an adenine from the sarcin/ricin loop in eukaryotic ribosomes. There are no approved inhibitors against ricin. We used a new strategy to disrupt RTA-ribosome interactions by fragment screening using surface plasmon resonance. Here, using a structure-guided approach, we improved the affinity and inhibitory activity of small-molecular-weight lead compounds and obtained improved compounds with over an order of magnitude higher efficiency. Four advanced compounds were characterized by X-ray crystallography. They bind at the RTA-ribosome binding site as the original compound but in a distinctive manner. These inhibitors bind remotely from the catalytic site and cause local conformational changes with no alteration of the catalytic site geometry. Yet they inhibit depurination by ricin holotoxin and inhibit the cytotoxicity of ricin in mammalian cells. They are the first agents that protect against ricin holotoxin by acting directly on RTA.

Cation-π interaction is a type of noncovalent interaction formed between the π-electron system and the positively charged ion or moieties. In this study, we designed a series of novel NQO1 substrates by introducing aliphatic nitrogen-containing side chains to fit with the L-shaped pocket of NQO1 by the formation of cation-π interactions. Molecular dynamics (MD) simulation indicated that the basic N atom in the side chain of NQO1 substrates, which is prone to be protonated under physiological conditions, can form cation-π interactions with the Phe232 and Phe236 residues of the NQO1 enzyme. Compound 4 with a methylpiperazinyl substituent was identified as the most efficient substrate for NQO1 with the reduction rate and catalytic efficiency of 1263 ± 61 μmol NADPH/min/μmol NQO1 and 2.8 ± 0.3 × 10 6  M -1 s -1 , respectively. Notably, compound 4 exhibited increased water solubility (110 μg/mL) compared to that of β-lap (43 μg/mL), especially under acidic condition (pH = 3, solubility > 1000 μg/mL). Compound 4 (IC 50 /A549 = 2.4 ± 0.6 μM) showed potent antitumor activity against NQO1-rich cancer cells through ROS generation via NQO1-mediated redox cycling. These results emphasized that the application of cation-π interactions by introducing basic aliphatic amine moiety is beneficial for both the water solubility and the NQO1-substrate binding, leading to promising NQO1-targeting antitumor candidates with improved druglike properties., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Masson SAS. All rights reserved.)

Since its discovery at the beginning of the past century, the essential nutrient l-Tryptophan (l-Trp) and its catabolic pathways have acquired an increasing interest in an ever wider scientific community for their pivotal roles in underlying many important physiological functions and associated pathological conditions. As a consequence, enzymes catalyzing rate limiting steps along l-Trp catabolic pathways - including IDO1, TDO, TPH1 and TPH2 - have turned to be interesting drug targets for the design and development of novel therapeutic agents for different disorders such as carcinoid syndrome, cancer and autoimmune diseases. This article provides a fresh comparative overview on the most recent advancements that crystallographic studies, biophysical and computational works have brought on structural aspects and molecular recognition patterns of these enzymes toward l-Trp. Finally, a conformational analysis of l-Trp is also discussed as part of the molecular recognition process governing the binding of a substrate to its cognate enzymes., (© 2021 The Authors. ChemMedChem published by Wiley-VCH GmbH.)

The enzyme 2-methylerythritol 2,4-cyclodiphosphate synthase, IspF, is essential for the biosynthesis of isoprenoids in most bacteria, some eukaryotic parasites, and the plastids of plant cells. The development of inhibitors that target IspF may lead to novel classes of anti-infective agents or herbicides. Enantiomers of tryptophan hydroxamate were synthesized and evaluated for binding to Burkholderia pseudomallei (Bp) IspF. The L-isomer possessed the highest potency, binding BpIspF with a K D of 36 µM and inhibited BpIspF activity 55% at 120 µM. The high-resolution crystal structure of the L-tryptophan hydroxamate (3)/BpIspF complex revealed a non-traditional mode of hydroxamate binding where the ligand interacts with the active site zinc ion through the primary amine. In addition, two hydrogen bonds are formed with active site groups, and the indole group is buried within the hydrophobic pocket composed of side chains from the 60 s/70 s loop. Along with the co-crystal structure, STD NMR studies suggest the methylene group and indole ring are potential positions for optimization to enhance binding potency., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Type 2 diabetes mellitus (DM) is a complex chronic disorder and a major global health problem. Insulin resistance is the primary detectable abnormality and the main characteristic feature in individuals with type 2 DM. Protein tyrosine phosphatase 1B (PTP1B) is a key negative regulator of the insulin signaling pathway, which dephosphorylates insulin receptor and insulin receptor substrates, suppressing the insulin signaling cascade. Therefore, the inhibition of PTP1B has become a potential strategy in the management of type 2 DM. In this study, a library of 22 pyrazoles was evaluated here for the first time against human PTP1B activity, using a microanalysis screening system. The results showed that 5-(2-hydroxyphenyl)-3-{2-[3-(4-nitrophenyl)-1,2,3,4-tetrahydronaphthyl]}-1-phenylpyrazole 20 and 3-(2-hydroxyphenyl)-5-{2-[3-(4-methoxyphenyl)]naphthyl}pyrazole 22 excelled as the most potent inhibitors of PTP1B, through noncompetitive inhibition mechanism. These findings suggest that the presence of additional benzene rings as functional groups in the pyrazole moiety increases the ability of pyrazoles to inhibit PTP1B. The most active compounds showed selectivity over the homologous T-cell protein tyrosine phosphatase (TCPTP). Molecular docking analyses were performed and revealed a particular contact signature involving residues like TYR46, ASP48, PHE182, TYR46, ALA217 and ILE219. This study represents a significant beginning for the design of novel PTP1B inhibitors., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2021 Elsevier B.V. All rights reserved.)

MutT Homolog 1 (MTH1) has been proven to hydrolyze oxidized nucleotide triphosphates during DNA repair. It can prevent the incorporation of wrong nucleotides during DNA replication and mitigate cell apoptosis. In a cancer cell, abundant reactive oxygen species can lead to substantial DNA damage and DNA mutations by base-pairing mismatch. MTH1 could eliminate oxidized dNTP and prevent cancer cells from entering cell death. Therefore, inhibition of MTH1 activity is considered to be an anti-cancer therapeutic target. In this study, high-throughput screening techniques were combined with a fragment-based library containing 2,313 compounds, which were used to screen for lead compounds with MTH1 inhibitor activity. Four compounds with MTH1 inhibitor ability were selected, and compound MI0639 was found to have the highest effective inhibition. To discover the selectivity and specificity of this action, several derivatives based on the MTH1 and MI0639 complex structure were synthesized. We compared 14 complex structures of MTH1 and the various compounds in combination with enzymatic inhibition and thermodynamic analysis. Nanomolar-range IC 50 inhibition abilities by enzyme kinetics and K d values by thermodynamic analysis were obtained for two compounds, named MI1020 and MI1024. Based on structural information and compound optimization, we aim to provide a strategy for the development of MTH1 inhibitors with high selectivity and specificity., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Present in all organisms, DNA ligases catalyse the formation of a phosphodiester bond between a 3' hydroxyl and a 5' phosphate, a reaction that is essential for maintaining genome integrity during replication and repair. Eubacterial DNA ligases use NAD + as a cofactor and possess low sequence and structural homology relative to eukaryotic DNA ligases which use ATP as a cofactor. These key differences enable specific targeting of bacterial DNA ligases as an antibacterial strategy. In this study, four small molecule accessible sites within functionally important regions of Escherichia coli ligase (EC-LigA) were identified using in silico methods. Molecular docking was then used to screen for small molecules predicted to bind to these sites. Eight candidate inhibitors were then screened for inhibitory activity in an in vitro ligase assay. Five of these (geneticin, chlorhexidine, glutathione (reduced), imidazolidinyl urea and 2-(aminomethyl)imidazole) showed dose-dependent inhibition of EC-LigA with half maximal inhibitory concentrations (IC 50 ) in the micromolar to millimolar range (11-2600 µM). Two (geneticin and chlorhexidine) were predicted to bind to a region of EC-LigA that has not been directly investigated previously, raising the possibility that there may be amino acids within this region that are important for EC-LigA activity or that the function of essential residues proximal to this region are impacted by inhibitor interactions with this region. We anticipate that the identified small molecule binding sites and inhibitors could be pursued as part of an antibacterial strategy targeting bacterial DNA ligases.

Over 50 tetrahydroindazoles were synthesized after 7-bromo-3,6,6-trimethyl-1-(pyridin-2-yl)-5,6,7,7a-tetrahydro-1H-indazol-4(3aH)-one (3) was identified as a hit compound in a high throughput screen for inhibition of CDK2 in complex with cyclin A. The activity of the most promising analogues was evaluated by inhibition of CDK2 enzyme complexes with various cyclins. Analogues 53 and 59 showed 3-fold better binding affinity for CDK2 and 2- to 10-fold improved inhibitory activity against CDK2/cyclin A1, E, and O compared to screening hit 3. The data from the enzyme and binding assays indicate that the binding of the analogues to a CDK2/cyclin complex is favored over binding to free CDK2. Computational analysis was used to predict a potential binding site at the CDK2/cyclin E1 interface., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Masson SAS. All rights reserved.)

Aminoacyl-tRNA synthetases (aaRSs) catalyze the esterification of tRNA with a cognate amino acid and are essential enzymes in all three kingdoms of life. Due to their important role in the translation of the genetic code, aaRSs have been recognized as suitable targets for the development of small molecule anti-infectives. In this review, following a concise discussion of aaRS catalytic and proof-reading activities, the various inhibitory mechanisms of reported natural and synthetic aaRS inhibitors are discussed. Using the expanding repository of ligand-bound X-ray crystal structures, we classified these compounds based on their binding sites, focusing on their ability to compete with the association of one, or more of the canonical aaRS substrates. In parallel, we examined the determinants of species-selectivity and discuss potential resistance mechanisms of some of the inhibitor classes. Combined, this structural perspective highlights the opportunities for further exploration of the aaRS enzyme family as antimicrobial targets.

Tuberculosis, caused by Mycobacterium tuberculosis (M. tuberculosis), is still responsible for a large number of fatal cases, especially in developing countries with alarming rates of incidence and prevalence worldwide. Mycobacterium tuberculosis has a remarkable ability to develop new resistance mechanisms to the conventional antimicrobials treatment. Because of this, there is an urgent need for novel bioactive compounds for its treatment. The dehydroquinate dehydratase II (DHQase II) is considered a key enzyme of shikimate pathway, and it can be used as a promising target for the design of new bioactive compounds with antibacterial action. The aim of this work was the construction of QSAR models to aid the design of new potential DHQase II inhibitors. For that purpose, various molecular modeling approaches, such as activity cliff, QSAR models and computer-aided ligand design were utilized. A predictive in silico 4D-QSAR model was built using a database comprising 86 inhibitors of DHQase II, and the model was used to predict the activity of the designed ligands. The obtained model proved to predict well the DHQase II inhibition for an external validation dataset ([Formula: see text] = 0.72). Also, the Activity Cliff analysis shed light on important structural features applied to the ligand design.

The spread of antibiotic resistance within the ESKAPE group of human pathogenic bacteria poses severe challenges in the treatment of infections and maintenance of safe hospital environments. This motivates efforts to validate novel target proteins within these species that could be pursued as potential targets for antibiotic development. Genetic data suggest that the enzyme FabG, which is part of the bacterial fatty acid biosynthetic system FAS-II, is essential in several ESKAPE pathogens. FabG catalyzes the NADPH dependent reduction of 3-keto-acyl-ACP during fatty acid elongation, thus enabling lipid supply for production and maintenance of the cell envelope. Here we report on small-molecule screening on the FabG enzymes from A. baumannii and S. typhimurium to identify a set of µM inhibitors, with the most potent representative (1) demonstrating activity against six FabG-orthologues. A co-crystal structure with FabG from A. baumannii (PDB:6T65) confirms inhibitor binding at an allosteric site located in the subunit interface, as previously demonstrated for other sub-µM inhibitors of FabG from P. aeruginosa. We show that inhibitor binding distorts the oligomerization interface in the FabG tetramer and displaces crucial residues involved in the interaction with the co-substrate NADPH. These observations suggest a conserved allosteric site across the FabG family, which can be potentially targeted for interference with fatty acid biosynthesis in clinically relevant ESKAPE pathogens., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

SNM1A is a nuclease that is implicated in DNA interstrand crosslink repair and, as such, its inhibition is of interest for overcoming resistance to chemotherapeutic crosslinking agents. However, the number and identity of the metal ion(s) in the active site of SNM1A are still unconfirmed, and only a limited number of inhibitors have been reported to date. Herein, we report the synthesis and evaluation of a family of malonate-based modified nucleosides to investigate the optimal positioning of metal-binding groups in nucleoside-derived inhibitors for SNM1A. These compounds include ester, carboxylate and hydroxamic acid malonate derivatives which were installed in the 5'-position or 3'-position of thymidine or as a linkage between two nucleosides. Evaluation as inhibitors of recombinant SNM1A showed that nine of the twelve compounds tested had an inhibitory effect at 1 mM concentration. The most potent compound contains a hydroxamic acid malonate group at the 5'-position. Overall, our studies advance the understanding of requirements for nucleoside-derived inhibitors for SNM1A and indicate that groups containing a negatively charged group in close proximity to a metal chelator, such as hydroxamic acid malonates, are promising structures in the design of inhibitors.

6-Hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) is a key enzyme in the folate biosynthesis pathway. It catalyzes pyrophosphoryl transfer from ATP to 6-hydroxymethyl-7,8-dihydropterin (HP). HPPK is essential for microorganisms but absent in mammals; therefore, it is an attractive target for developing novel antimicrobial agents. Previously, based on our studies of the structure and mechanism of HPPK, we created first-generation bisubstrate inhibitors by linking 6-hydroxymethylpterin to adenosine through phosphate groups, and developed second-generation inhibitors by replacing the phosphate bridge with a linkage that contains a piperidine moiety. Here, we report third-generation inhibitors designed based on the piperidine-containing inhibitor, mimicking the transition state. We synthesized two such inhibitors, characterized their protein-binding and enzyme inhibition properties, and determined their crystal structures in complex with HPPK, advancing the development of such bisubstrate analog inhibitors., (Published by Elsevier Ltd.)

Inosine-5'-monophosphate dehydrogenase (IMPDH) is a potential target for microorganisms. However, identifying inhibitor design determinants for IMPDH orthologs continues to evolve. Herein, a series of mycophenolic anilide inhibitors of Cryptosporidium parvum and human IMPDHs are reported. Furthermore, molecular docking of 12 (e.g. SH-19; CpIMPDH K i,app  = 0.042 ± 0.015 µM, HsIMPDH2 K i,app  = 0.13 ± 0.05 µM) supports different binding modes with the two enzymes. For CpIMPDH the inhibitor extends into a pocket in an adjacent subunit. In contrast, docking suggests the inhibitor interacts with Ser276 in the NAD binding site in HsIMPDH2, as well as an adjacent pocket within the same subunit. These results provide further guidance for generating IMPDH inhibitors for enzymes found in an array of pathogenic microorganisms, including Mycobacterium tuberculosis., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Trypanosoma brucei is the causative agent of human African trypanosomiasis (HAT). Nitrogen-containing bisphosphonates, a current treatment for bone diseases, have been shown to block the growth of the T. brucei parasites by inhibiting farnesyl pyrophosphate synthase (FPPS); however, due to their poor pharmacokinetic properties, they are not well suited for antiparasitic therapy. Recently, an allosteric binding pocket was discovered on human FPPS, but its existence on trypanosomal FPPS was unclear. We applied NMR and X-ray fragment screening to T. brucei FPPS and report herein on four fragments bound to this previously unknown allosteric site. Surprisingly, non-bisphosphonate active-site binders were also identified. Moreover, fragment screening revealed a number of additional binding sites. In an early structure-activity relationship (SAR) study, an analogue of an active-site binder was unexpectedly shown to bind to the allosteric site. Overlaying identified fragment binders of a parallel T. cruzi FPPS fragment screen with the T. brucei FPPS structure, and medicinal chemistry optimisation based on two binders revealed another example of fragment "pocket hopping". The discovery of binders with new chemotypes sets the framework for developing advanced compounds with pharmacokinetic properties suitable for the treatment of parasitic infections by inhibition of FPPS in T. brucei parasites., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Andrographolide and its benzylidene derivatives, SRJ09 and SRJ23, potentially bind oncogenic K-Ras to exert anticancer activity. Their molecular interactions with K-Ras oncoproteins that lead to effective biological activity are of major interest. Methods & results: In silico docking and molecular dynamics simulation were performed using Glide and Desmond, respectively; while saturation transfer difference NMR was performed using GDP-bound K-RasG12V. SRJ23 was found to bind strongly and selectively to K-RasG12V, by anchoring to a binding pocket (namely p2) principally via hydrogen bond and hydrophobic interactions. The saturation transfer difference NMR analysis revealed the proximity of protons of functional moieties in SRJ23 to K-RasG12V, suggesting positive binding. Conclusion: SRJ23 binds strongly and interacts stably with K-RasG12V to exhibit its inhibitory activity.

The enoyl-acyl carrier protein reductase enzyme FabI is essential for fatty acid biosynthesis in Staphylococcus aureus and represents a promising target for the development of novel, urgently needed anti-staphylococcal agents. Here, we elucidate the mode of action of the kalimantacin antibiotics, a novel class of FabI inhibitors with clinically-relevant activity against multidrug-resistant S. aureus. By combining X-ray crystallography with molecular dynamics simulations, in vitro kinetic studies and chemical derivatization experiments, we characterize the interaction between the antibiotics and their target, and we demonstrate that the kalimantacins bind in a unique conformation that differs significantly from the binding mode of other known FabI inhibitors. We also investigate mechanisms of acquired resistance in S. aureus and identify key residues in FabI that stabilize the binding of the antibiotics. Our findings provide intriguing insights into the mode of action of a novel class of FabI inhibitors that will inspire future anti-staphylococcal drug development., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Blood coagulation factor XIII (FXIII, F13) is considered to be a promising target for anticoagulants with reduced bleeding risk because of its unique position in the coagulation cascade downstream of thrombin. However, until now, no potent drug addressing FXIII has been available, indeed no compound has even entered clinical trials yet. In 2013, we published the co-crystal structure of FXIII in the active state (FXIIIa°), thereby providing a detailed map of the active site for the rational design of potent FXIIIa blockers. Here we report, for the first time, a structure-based approach to improving the affinity of FXIIIa inhibitors. FXIII was crystallized in complex with a methyl thiazole moiety to address a novel transient hydrophobic pocket close to the catalytic center. By subsequent structure-based design to rationalize the introduction of an ethyl ester, the potency of the inhibitor was improved significantly compared to that of the parent lead compound. The occupancy of the hydrophobic pocket described here might turn out to be a key step in the development of a potent reversible and orally available FXIIIa blocker., (© 2020 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.)

A large number of crystallographic structures of IDO1 in different ligand-bound and -unbound states have been disclosed over the last decade. Yet, only a few of them have been exploited for structure-based drug design (SBDD) campaigns. In this study, we analyzed the structural motifs and molecular-recognition properties of three groups of IDO1 structures: 1) structures containing the heme group and inhibitors in the catalytic site; 2) heme-free structures of IDO1; 3) substrate-bound structures of IDO1. The results suggest that unrelated conformations of the enzyme have been solved with different ligand-induced changes of secondary motifs that localize even in regions remote from the catalytic site. Moreover, the study identified an uncharted region of molecular-recognition space covered by IDO1 binding sites that could guide the selection of diverse structures for additional SBDD studies aimed at the identification of novel lead compounds with differentiated chemical scaffolds., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Bedaquiline (BDQ) has demonstrated formidable bactericidal activity towards Mycobacterium tuberculosis (Mtb) in the treatment of multi-drug resistant (MDR) and extensively drug resistant (XDR) tuberculosis (TB). BDQ elicits its therapeutic function by halting the ionic shuttle of Mtb via mycobacterial F 1 F 0 ATP-synthase blockade. However, triple mutations (L59 V, E61D and I66 M) at the ligand-binding cavity characterize emerging BDQ-resistant strains thereby restraining the potentials embedded in this anti-microbial compound, particularly in MDR/XDR-TB therapy. In this report, the effects of these triple mutations on BDQ-Mtb F 1 F 0 ATP-synthase binding were investigated using molecular dynamics, free energy binding and residue interaction network (RIN) analyses. The highlight of our findings is the drastic reduction in BDQ binding affinity (ΔG) in the triple mutant protein, which was caused by a systemic loss in high-affinity interactions primarily mediated by L59, E61 and I66. While wildtype L59 and I66 formed pi-alkyl interactions with BDQ at the F 1 F 0 ATP-synthase binding site, E61 elicited conventional (O--HO) bond. Upon transition, V59 and I66 were devoid of interactions with BDQ while D61 existed in a weaker non-conventional (C--HO) bond. Likewise, these mutations distorted the binding site and overall structural architecture of F 1 F 0 ATP-synthase in the presence of BDQ as revealed by the RIN and conformational analyses. Insights from this study could serve as a starting point for the structure-based design of novel inhibitors that could overcome mutational setbacks posed by BDQ-resistant strains in MDR/XDR-TB treatment., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Mutations in isocitrate dehydrogenase 1 (IDH1) are found in 6% of AML patients. Mutant IDH produces R-2-hydroxyglutarate (R-2HG), which induces histone- and DNA-hypermethylation through the inhibition of epigenetic regulators, thus linking metabolism to tumorigenesis. Here we report the biochemical characterization, in vivo antileukemic effects, structural binding, and molecular mechanism of the inhibitor HMS-101, which inhibits the enzymatic activity of mutant IDH1 (IDH1mut). Treatment of IDH1mut primary AML cells reduced 2-hydroxyglutarate levels (2HG) and induced myeloid differentiation in vitro. Co-crystallization of HMS-101 and mutant IDH1 revealed that HMS-101 binds to the active site of IDH1mut in close proximity to the regulatory segment of the enzyme in contrast to other IDH1 inhibitors. HMS-101 also suppressed 2HG production, induced cellular differentiation and prolonged survival in a syngeneic mutant IDH1 mouse model and a patient-derived human AML xenograft model in vivo. Cells treated with HMS-101 showed a marked upregulation of the differentiation-associated transcription factors CEBPA and PU.1, and a decrease in cell cycle regulator cyclin A2. In addition, the compound attenuated histone hypermethylation. Together, HMS-101 is a unique inhibitor that binds to the active site of IDH1mut directly and is active in IDH1mut preclinical models.

Poly(ADP-ribose) glycohydrolase (PARG) plays an essential role in poly(ADP-ribose) (PAR) turnover, and thereby regulating DNA transactions, such as DNA repair, replication, transcription and recombination. Here, we examined the inhibitory activities of 6-hydroxy-3H-xanthene-3-one (HXO) derivatives and analyzed their binding modes in the active site of PARG by in silico docking study. Among the derivatives, Rose Bengal was found to be the most potent inhibitor of PARG and its halogen groups were revealed to cooperatively potentiate the inhibitory activity. Importantly, the binding mode of Rose Bengal occupied the active site of PARG revealed the presence of unique "Sandwich" residues of Asn869 and Tyr792, which enable the inhibitor to bind tightly with the active pocket. This sandwich interaction could stabilize the π-π interactions of HXO scaffold with Phe902 and Tyr795. In addition, to increase the binding affinity, the iodine and chlorine atoms of this inhibitor could contribute to the inducing of favorable disorders, which promote an entropy boost on the active site of PARG for structural plasticity, and making the stable configuration of HXO scaffold in the active site, respectively, as judged by the analysis of binding free energy. These results provide new insights into the active site of PARG and an additional opportunity for designing selective PARG inhibitors., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Protein tyrosine phosphatase 2 (SHP-2) has long been proposed as a cancer drug target. Several small-molecule compounds with different mechanisms of SHP-2 inhibition have been reported, but none are commercially available. Pool selectivity over protein tyrosine phosphatase 1 (SHP-1) and a lack of cellular activity have hindered the development of selective SHP-2 inhibitors. In this review, we describe the binding modes of existing inhibitors and SHP-2 binding sites, summarize the characteristics of the sites involved in selectivity, and identify the suitable groups for interaction with the binding sites., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

EDTA is commonly used as an efficient chelator of metal ion enzyme cofactors. It is highly soluble, optically inactive and does not interfere with most chemicals used in standard buffers making EDTA a common choice to generate metal-free conditions for biochemical and biophysical investigations. However, the controversy in the literature on metal-free enzyme activities achieved using EDTA or by other means called our attention to a putative effect of EDTA beyond chelation. Here, we show that EDTA competes for the nucleotide binding site of the nucleotide hydrolase dUTPase by developing an interaction network within the active site similar to that of the substrate. To achieve these findings, we applied kinetics and molecular docking techniques using two different dUTPases. Furthermore, we directly measured the binding of EDTA to dUTPases and to two other dNTPases, the Taq polymerase and MutT using isothermal titration calorimetry. EDTA binding proved to be exothermic and mainly enthalpy driven with a submicromolar dissociation constant considerably lower than that of the enzyme:substrate or the Mg:EDTA complexes. Control proteins, including an ATPase, did not interact with EDTA. Our findings indicate that EDTA may act as a selective inhibitor against dNTP hydrolyzing enzymes and urge the rethinking of the utilization of EDTA in enzymatic experiments., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

A rational approach that may be applied to a broad class of enzyme-catalyzed reactions to design enzyme inhibitors affords a powerful strategy, facilitating the development of drugs and/or molecular probes of enzyme mechanisms. A strategy for the development of substrate-product analogues (SPAs) as inhibitors of racemases and epimerases is elaborated using isoleucine 2-epimerase from Lactobacillus buchneri (LbIleE) as a model enzyme. LbIleE catalyzes the PLP-dependent, reversible, racemization or epimerization of nonpolar amino acids at the C-2 position. The enzyme plays an important role in the biosynthesis of branched-chain d-amino acids and is a potential target for the development of antimicrobial agents. 3-Ethyl-3-methyl-l-norvaline (Ki = 2.9 ± 0.2 mM) and 3-ethyl-3-methyl-d-norvaline (Ki = 1.5 ± 0.2 mM) were designed as SPAs based on the movement of the sec-butyl side chain of the substrate l-Ile during catalysis, and were competitive inhibitors with binding affinities exceeding that of l-Ile by 1.3- and 2.5-fold, respectively. Surprisingly, these compounds were not substrates, but the corresponding compounds lacking the 3-methyl group were substrates. Unlike serine, glutamate, and proline racemases, which exhibit stringent steric requirements at their active sites, the active site of LbIleE was amenable to binding bulky SPAs. Moreover, LbIleE bound the SPA 2,2-di-n-butylglycine (Ki = 11.0 ± 0.2 mM) as a competitive inhibitor, indicating that the hydrophobic binding pocket at the active site was sufficiently plastic to tolerate gem-dialkyl substitution at the α-carbon of an amino acid. Overall, these results reveal that amino acid racemases/epimerases are amenable to inhibition by SPAs provided that they possess a capacious active site.

Among the >120 modified ribonucleosides in the prokaryotic epitranscriptome, many tRNA modifications are critical to bacterial survival, which makes their synthetic enzymes ideal targets for antibiotic development. Here we performed a structure-based design of inhibitors of tRNA-(N 1 G37) methyltransferase, TrmD, which is an essential enzyme in many bacterial pathogens. On the basis of crystal structures of TrmDs from Pseudomonas aeruginosa and Mycobacterium tuberculosis , we synthesized a series of thienopyrimidinone derivatives with nanomolar potency against TrmD in vitro and discovered a novel active site conformational change triggered by inhibitor binding. This tyrosine-flipping mechanism is uniquely found in P. aeruginosa TrmD and renders the enzyme inaccessible to the cofactor S -adenosyl-l-methionine (SAM) and probably to the substrate tRNA. Biophysical and biochemical structure-activity relationship studies provided insights into the mechanisms underlying the potency of thienopyrimidinones as TrmD inhibitors, with several derivatives found to be active against Gram-positive and mycobacterial pathogens. These results lay a foundation for further development of TrmD inhibitors as antimicrobial agents.

The first chemical probe to primarily occupy the co-factor binding site of a Su(var)3-9, enhancer of a zeste, trithorax (SET) domain containing protein lysine methyltransferase (PKMT) is reported. Protein methyltransferases require S -adenosylmethionine (SAM) as a co-factor (methyl donor) for enzymatic activity. However, SAM itself represents a poor medicinal chemistry starting point for a selective, cell-active inhibitor given its extreme physicochemical properties and its role in multiple cellular processes. A previously untested medicinal chemistry strategy of deliberate file enrichment around molecules bearing the hallmarks of SAM, but with improved lead-like properties from the outset, yielded viable hits against SET and MYND domain-containing protein 2 (SMYD2) that were shown to bind in the co-factor site. These leads were optimized to identify a highly biochemically potent, PKMT-selective, and cell-active chemical probe. While substrate-based inhibitors of PKMTs are known, this represents a novel, co-factor-derived strategy for the inhibition of SMYD2 which may also prove applicable to lysine methyltransferase family members previously thought of as intractable.

The glyoxalase-I (GLO-I) enzyme, which is the initial enzyme of the glyoxalase system that is responsible for the detoxification of cytotoxic α-ketoaldehydes, such as methylglyoxal, has been approved as a valid target in cancer therapy. Overexpression of GLO-I has been observed in several types of carcinomas, including breast, colorectal, prostate, and bladder cancer. In this work we aimed to identify potential GLO-I inhibitors via employing different structure-based drug design techniques including structure-based poly-pharmacophore modelling, virtual screening, and molecular docking. Poly-pharmacophore modelling was applied in this study in order to thoroughly explore the binding site of the target enzyme, thereby, revealing hits that could bind in a nonconventional way which can pave the way for designing more potent and selective ligands with novel chemotypes. The modelling phase has resulted in the selection of 31 compounds that were biologically evaluated against human GLO-I enzyme. Among the tested set, seven compounds showed excellent inhibitory activities with IC 50 values ranging from 0.34 to 30.57 μM. The most active compound (ST018515) showed an IC 50 of 0.34 ± 0.03 μM, which, compared to reported GLO-I inhibitors, can be considered a potent inhibitor, making it a good candidate for further optimization towards designing more potent GLO-I inhibitors.

Falcipain-2 (FP-2) is a Plasmodium falciparum cysteine protease that has been extensively targeted to identify novel antimalarials. Remarkably, previous reports have shown that FP-2 can be allosterically modulated and, for a particular noncompetitive chalcone inhibitor, the existing lines of experimental evidence can guide the prediction of its unknown binding mode to the enzyme in a reliable fashion. In this work, we propose a structure of FP-2 in complex with the aforementioned compound that fulfills all of the experimental data, by employing a combination of molecular modeling tools, such as pocket volume measurements, docking, molecular dynamics (MD) simulations, and free energy calculations. Our results show that the studied inhibitor binds a transient pocket occluded in all of the available FP-2 crystal structures and lying in a region previously characterized as a potential allosteric site in related cysteine proteases. In addition, we detected in silico the occurrence of significant community reorganization in FP-2, increased signal transmission between the allosteric pocket and the active site, and change in loop motions and residue p K a values upon the compound binding, thus providing insight into the uncharacterized allosteric mechanism. Overall, this study yields valuable predictions for the design of novel allosteric inhibitors against FP-2 and other cysteine proteases.

Giardia lamblia is the causal agent of giardiasis, one of the most prevalent parasitosis in the world. Even though effective pharmacotherapies against this parasite are available, the disadvantages associated with its use call for the development of new antigiardial compounds. Based on the Giardia dependence on glycolysis as a main energy source, glycolytic enzymes appear to be attractive targets with antiparasitic potential. Among these, fructose 1,6-biphosphate aldolase (GlFBPA) has been highlighted as a promising target for drug design. Current efforts are based on the design of competitive inhibitors of GlFBPA; however, in the kinetic context of metabolic pathways, competitive inhibitors seem to have low potential as therapeutic agents. In this work, we performed an experimental and in silico structure-based approach to propose a non-catalytic binding site which could be used as a hot spot for antigardial drug design. The druggability of the selected binding site was experimentally tested; the alteration of the selected region by site directed mutagenesis disturbs the catalytic properties and the stability of the enzyme. A computational automated search of binding sites supported the potential of this region as functionally relevant. A preliminary docking study was performed, in order to explore the feasibility and type of molecules to be able to accommodate in the proposed binding region. Altogether, the results validate the proposed region as a specific molecular binding site with pharmacological potential.

Herein, we describe the development of a novel staple with an electrophilic warhead to enable the generation of stapled peptide covalent inhibitors of the p53-MDM2 protein-protein interaction (PPI). The peptide developed showed complete and selective covalent binding resulting in potent inhibition of p53-MDM2 PPI.

Tyrosinase enzyme plays a crucial role in melanin biosynthesis and enzymatic browning process of vegetables and fruits. A series of veratric acid derivatives containing benzylidene-hydrazine moieties with different substitutions were synthesized and their inhibitory effect on mushroom tyrosinase and free radical scavenging activity were evaluated. The results indicated that N'-(4-chlorobenzylidene)-3,4-dimethoxybenzohydrazide (D5) and N'-(2,3-dihydroxybenzylidene)-3,4-dimethoxybenzohydrazide (D12) showed the highest tyrosinase inhibitory activity with IC 50 values of 19.72 ± 1.84 and 20.63 ± 0.79 μM, respectively, that were comparable with the IC 50 value of kojic acid (19.08 ± 1.21 μM). D12 was also a potent radical scavenger with EC 50 value of 0.0097 ± 0.0011 mM. The free radical scavenging activity of D12 was comparable with the standard quercetin. The inhibition kinetic analyzed by Lineweaver-Burk plots revealed that compound D5 was a competitive tyrosinase inhibitor. Molecular docking study was carried out for the derivatives demonstrating tyrosinase inhibitory activity. D5 and D12 possessed the most negative estimated free energies of binding in mushroom tyrosinase active site. Therefore, D5 and D12 could be introduced as potent tyrosinase inhibitors that might be promising leads in medicine, cosmetics and food industry., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

For drug design projects it is essential to rationally induce and explain selectivity. In this context shape complementarity as well as protein and ligand flexibility represent important factors. Currently available tools for the analysis of protein-ligand interactions focus mainly on electrostatic complementarity and/or static structures. Here we address the shortcomings of available methods by presenting two new tools: The first one can be used to assess steric complementarity in flexible protein-ligand complexes in order to explain selectivity of known ligands. It further allows to determine ligand atoms with especially good or bad shape-fit which can be of use in lead optimization projects. The second tool was designed to detect differences in protein flexibility in similar proteins along with their exploitation for virtual screening. Both tools yield interesting results when applied to data of protein tyrosine phosphatase 1B (PTP1B): The case of PTP1B has proven especially difficult in terms of selectivity, due to a closely related phosphatase connected to severe undesired effects. With our tool for steric complementarity assessment we were able to explain previously undisclosed causes of moderate selectivity of selected PTP1B ligands. The second tool allowed us to find differences of flexibility in the two highly similar proteins and give directions for exploitation in virtual screening., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Glutathione S-transferase π (GSTP 1-1 ) is overexpressed in many types of cancer and is involved in drug resistance. Therefore, GSTP 1-1 is an important target in cancer therapy, and many GST inhibitors have been reported. We had previously developed an irreversible inhibitor, GS-ESF, as an effective GST inhibitor; however, its cellular permeability was too low for it to be used in inhibiting intracellular GST. We have now developed new irreversible inhibitors by introducing sulfonyl fluoride (SF) into chloronitrobenzene (CNB). The mechanism of action was revealed to be that CNBSF first reacts with glutathione (GSH) through an aromatic substitution in the cell, then the sulfonyl group on the GSH conjugate with CNBSF reacts with Tyr108 of GST to form a sulfonyl ester bond. Our new inhibitor irreversible inhibited GSTP 1-1 both in vitro and in cellulo with a long duration of action., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

ATP-citrate lyase (ACLY) is a central metabolic enzyme and catalyses the ATP-dependent conversion of citrate and coenzyme A (CoA) to oxaloacetate and acetyl-CoA 1-5 . The acetyl-CoA product is crucial for the metabolism of fatty acids 6,7 , the biosynthesis of cholesterol 8 , and the acetylation and prenylation of proteins 9,10 . There has been considerable interest in ACLY as a target for anti-cancer drugs, because many cancer cells depend on its activity for proliferation 2,5,11 . ACLY is also a target against dyslipidaemia and hepatic steatosis, with a compound currently in phase 3 clinical trials 4,5 . Many inhibitors of ACLY have been reported, but most of them have weak activity 5 . Here we report the development of a series of low nanomolar, small-molecule inhibitors of human ACLY. We have also determined the structure of the full-length human ACLY homo-tetramer in complex with one of these inhibitors (NDI-091143) by cryo-electron microscopy, which reveals an unexpected mechanism of inhibition. The compound is located in an allosteric, mostly hydrophobic cavity next to the citrate-binding site, and requires extensive conformational changes in the enzyme that indirectly disrupt citrate binding. The observed binding mode is supported by and explains the structure-activity relationships of these compounds. This allosteric site greatly enhances the 'druggability' of ACLY and represents an attractive target for the development of new ACLY inhibitors.

Generation and screening of oxime libraries by competitive MS Binding Assays represents a powerful tool for the identification of new compounds, with affinity to mGAT1, the most abundant plasma membrane bound GABA transporter in the CNS. By screening a guvacine derived oxime library, new potent inhibitors of mGAT1 had been revealed. In the present study, oxime libraries generated by reaction of a large excess of a rac-nipecotic acid derivative displaying a hydroxylamine functionality in which various aldehydes under suitable conditions, were examined for new potent inhibitors of mGAT1. The pK i values obtained of the best hits were compared with those of related compounds displaying a guvacine instead of a nipecotic acid subunit as hydrophilic moiety. Amongst the new compounds one of the most affine ligands of mGAT1 known so far (pK i  = 8.55 ± 0.04) was found., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Xanthohumol (XN), a prenylated chalcone unique to hops (Humulus lupulus) and two derived prenylflavanones, isoxanthohumol (IX) and 8-prenylnaringenin (8-PN) gained increasing attention as potential anti-diabetic and cancer preventive compounds. Two enzymes of the aldo-keto reductase (AKR) superfamily are notable pharmacological targets in cancer therapy (AKR1B10) and in the treatment of diabetic complications (AKR1B1). Our results show that XN, IX and 8-PN are potent uncompetitive, tight-binding inhibitors of human aldose reductase AKR1B1 (K i  = 15.08 μM, 0.34 μM, 0.71 μM) and of human AKR1B10 (K i  = 20.11 μM, 2.25 μM, 1.95 μM). The activity of the related enzyme AKR1A1 was left unaffected by all three compounds. This is the first time these three substances have been tested on AKRs. The results of this study may provide a basis for further quantitative structure?activity relationship models and promising scaffolds for future anti-diabetic or carcinopreventive drugs.

In this study, two novel phthalocyanine complexes were synthesized using their corresponding metal salts and 4-(4-(3-(2,4,5-trimethoxyphenyl)acryloyl)phenoxy)phthalo-nitrile as chalcone ligand (4), which was prepared from the reaction of 4-nitrophthalonitrile with 4-hydroxyphenyl-3-(2,4,5-trimethoxyphenyl)prop-2-en-1-one (3). These metallophthalocyanines showed good solubility in organic solvents such as CDCl 3 , DCM, THF, DMF, and DMSO. The novel phthalocyanine compounds 4a (Pc-Zn) and 4b (Pc-Co) were characterized using their UV-vis, FT-IR, 1 H NMR, 13 C NMR, and MALDI-TOF mass spectra and elemental analysis. Then the DNA-binding and xanthine oxidase and carbonic anhydrase-I inhibition properties of compounds 4a and 4b were investigated. Photochemical properties (such as singlet oxygen generation and photodegradation) of this novel chalcone phthalocyanine (4a) were determined in dimethyl sulfoxide (DMSO)., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Human indoleamine 2,3-dioxygenase 1 (hIDO1) is an important heme-containing enzyme that is a key drug target for cancer immunotherapy. Several hIDO1 inhibitors have entered clinical trials, among which BMS-986205 (BMS) stands out as the only suicide inhibitor. Despite its "best-in-class" activity, the action mechanism of BMS remains elusive. Here, we report three crystal structures of hIDO1-BMS complexes that define the complete binding trajectory of the inhibitor. BMS first binds in a solvent exposed surface cleft near the active site in an extended conformation. The initial binding partially unfolds the active site, which triggers heme release, thereby exposing a new binding pocket. The inhibitor then undergoes a large scale movement to this new binding pocket, where it binds by adopting a high energy kinked conformation. Finally, the inhibitor relaxes to a bent conformation, via an additional large scale rearrangement, culminating in the energy minimum state. The structural data offer a molecular explanation for the remarkable efficacy and suicide inhibition activity of the inhibitor. They also suggest a novel strategy that can be applied for drug development targeting hIDO1 and related enzymes.

Aristolochic acid I (AAI) is a phytocompound that is linked to the progressive renal disease and development of human urothelial carcinoma. The bladder cancer-associated protein (BLCAP) gene exhibits a tumor suppressor function in various tumors, including bladder carcinoma. This study evaluated the effect of AAI on BLCAP expression and its associated mechanism in human cells. Administering AAI to human embryonic kidney cells (HEK293), human proximal tubule epithelial cells (HK-2) and urinary bladder cancer cells (HT-1376) significantly reduced the expression of BLCAP mRNA and protein. AAI also effectively suppressed the luciferase activities driven by BLCAP promoters of various lengths in HEK293 cells. AAI significantly reduced both activator protein 1 (AP-1) and nuclear factor-κB (NF-κB) activities in reporter assays, but further point mutations revealed that Ap-1 and NF-κB binding sites on the BLCAP promoter were not AAI-responsive elements. Application of the DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine (5-aza-dC), reversed the decline of BLCAP expression that had been induced by AAI. However, AAI exposure did not alter hypermethylation of the BLCAP promoter, determined by methyl-specific polymerase chain reaction (PCR) and bisulfate sequencing. Knocking down BLCAP in HEK293 cell line enhanced the potential for cellular migration, invasion, and proliferation, along with the induction of a capacity for anchorage-independent growth. In conclusion, AAI down-regulated the expression of BLCAP gene and the deficiency in BLCAP expression contributed to the malignant transformation of human cells, implying that BLCAP may have a role in mediating AAI-associated carcinogenesis., (Copyright © 2018 Elsevier B.V. All rights reserved.)

We have previously reported benzimidazole-based compounds to be potent inhibitors of FabI for Francisella tularensis (FtFabI), making them promising antimicrobial hits. Optically active enantiomers exhibit markedly differing affinities toward FtFabI. The IC 50 of benzimidazole (-)-1 is ∼100× lower than the (+)-enantiomer, with similar results for the 2 enantiomers. Determining the absolute configuration for these optical compounds and elucidating their binding modes is important for further design. Electronic circular dichroism (ECD) quantum calculations have become important in determining absolute configurations of optical compounds. We determined the absolute configuration of (-)/(+)-1 and (-)/(+)-2 by comparing experimental spectra and theoretical density functional theory (DFT) simulations of ECD spectra at the B3LYP/6-311+G(2d, p) level using Gaussian09. Comparison of experimental and calculated ECD spectra indicates that the S configuration corresponds to the (-)-rotation for both compounds 1 and 2, while the R configuration corresponds to the (+)-rotation. Further, molecular dynamics simulations and MM-GBSA binding energy calculations for these two pairs of enantiomers with FtFabI show much tighter binding MM-GBSA free energies for S-1 and S-2 than for their enantiomers, R-1 and R-2, consistent with the S configuration being the more active one, and with the ECD determination of the S configuration corresponding to (-) and the R configuration corresponding to (+). Thus, our computational studies allow us to assign (-) to (S)- and (+) to (R)- for compounds 1 and 2, and to further evaluate structural changes to improve efficacy., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Methyl gallate was purified, by lipoxygenase (LOX) inhibitory activity-guided method since its alleged anti-inflammatory property, from Bergenia ligulata (Wall), a plant used in the traditional, Ayurvedic system of medicine extensively. The LOX inhibitory property of methyl gallate was studied by enzyme kinetics, isothermal titration calorimetry and molecular docking followed by molecular simulation studies. The wet-laboratory experiments and in silico studies showed complete agreement, and promise of methyl gallate as a drug-lead molecular scaffold for anti-inflammatory therapy, based on LOX inhibition. The expressed work shows the need of nonactive site binding parameters to be considered while designing of inhibitors based on the specificities toward active sites of enzymes.

Bacterial RNA polymerase (RNAP) is an effective target for antibacterial treatment. In order to search new potential targets in RNAP of Mycobacterium, we detected adaptive selections of RNAP related genes in 13 strains of Mycobacterium by phylogenetic analysis. We first collected sequences of 17 genes including rpoA, rpoB, rpoC, rpoZ, and sigma factor A-M. Then maximum likelihood trees were constructed, followed by positive selection detection. We found that sigG shows positive selection along the clade (M. tuberculosis, M. bovis), suggesting its important evolutionary role and its potential to be a new antibacterial target. Moreover, the regions near 933Cys and 935His on the rpoB subunit of M. tuberculosis showed significant positive selection, which could also be a new attractive target for anti-tuberculosis drugs., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The aminopeptidase N (APN) plays a critical role in angiogenesis and is over-expressed in tumor cells. In this paper, we report the synthesis and enzyme inhibition assay of furoic peptidomimetic compounds. These new compounds exhibit potent inhibitory ability toward APN with IC50 values lying in the micromolar level. The binding mode of inhibitors in APN active site was explained by a molecular simulation study. These data reveal that ligand coordinating with the catalytic Zn-ion is very important for inhibitory activities.

The bifunctional farnesyl/geranylgeranyl diphosphate synthase (FPPS/GGPPS) is a key branchpoint enzyme in isoprenoid biosynthesis in Plasmodium falciparum (malaria) parasites. PfFPPS/GGPPS is a validated, high-priority antimalarial drug target. Unfortunately, current bisphosphonate drugs that inhibit FPPS and GGPPS enzymes by acting as a diphosphate substrate analog show poor bioavailability and selectivity for PfFPPS/GGPPS. We identified a new non-bisphosphonate compound, MMV019313, which is highly selective for PfFPPS/GGPPS and showed no activity against human FPPS or GGPPS. Inhibition of PfFPPS/GGPPS by MMV019313, but not bisphosphonates, was disrupted in an S228T variant, demonstrating that MMV019313 and bisphosphonates have distinct modes of inhibition. Molecular docking indicated that MMV019313 did not bind previously characterized substrate sites in PfFPPS/GGPPS. Our finding uncovers a new, selective small-molecule binding site in this important antimalarial drug target with superior druggability compared with the known inhibitor site and sets the stage for the development of Plasmodium-specific FPPS/GGPPS inhibitors., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Synthesized cyclic perylene diimide, cPDI, showed the binding constant of 6.3 × 10 6  M -1 with binding number of n = 2 with TA-core as a tetraplex DNA in 50 mM Tris-HCl buffer (pH = 7.4) containing 100 mM KCl using Schatchard analysis and showed a higher preference for tetraplex DNA than for double stranded DNA with over 10 3 times. CD spectra showed that TA-core induced its antiparallel conformation upon addition of cPDI in the absence or presence of K + or Na + ions. The cPDI inhibits the telomerase activity with IC 50 of 0.3 µM using TRAP assay which is potential anti-cancer drug with low side effect., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

The synthesis of the O-3 triazole-linked galactosyl arylsulfonamides 1-7 as potential inhibitors of Trypanosoma cruzi cell invasion is described. These target compounds were synthesized by Cu(I)-catalysed azide-alkyne cycloaddition reaction ('click chemistry') between different azide arylsulfonamides and the alkyne-based sugar 3-O-propynyl-βGalOMe. Inhibition assays of T. cruzi cell invasion with compounds 1-7 showed reduced values of infection index (∼20) for compounds 3 and 5, bearing the corresponding 5-methylisoxazole and 2,4-dimethoxypyrimidine groups, which also presented higher binding affinities to galectin-3 (EC 50 17-18 μM) in Corning Epic label-free assays. In agreement with experimental results, the assessment of the theoretical binding of compounds 1-7 to galectin-3 by MM/PBSA method displayed higher affinities for compounds 3 (-9.7 kcal/mol) and 5 (-11.1 kcal/mol). Overall, these achievements highlight compounds 3 and 5 as potential T. cruzi cell invasion blockers by means of a galectin-3 binding-related mechanism, revealing galectin-3 as an important host target for design of novel anti-trypanosomal agents., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Viruses of the Caliciviridae cause significant and sometimes lethal diseases, however despite substantial research efforts, specific antivirals are lacking. Broad-spectrum antivirals could combat multiple viral pathogens, offering a rapid solution when no therapies exist. The RNA-dependent RNA polymerase (RdRp) is an attractive antiviral target as it is essential for viral replication and lacks mammalian homologs. To focus the search for pan-Caliciviridae antivirals, the RdRp was probed with non-nucleoside inhibitors (NNIs) developed against hepatitis C virus (HCV) to reveal both allosteric ligands for structure-activity relationship enhancement, and highly-conserved RdRp pockets for antiviral targeting. The ability of HCV NNIs to inhibit calicivirus RdRp activities was assessed using in vitro enzyme and murine norovirus cell culture assays. Results revealed that three NNIs which bound the HCV RdRp Thumb I (TI) site also inhibited transcriptional activities of six RdRps spanning the Norovirus, Sapovirus and Lagovirus genera of the Caliciviridae. These NNIs included JTK-109 (RdRp inhibition range: IC 50 4.3-16.6 μM), TMC-647055 (IC 50 range: 18.8-45.4 μM) and Beclabuvir (IC 50 range: 23.8->100 μM). In silico studies and site-directed mutagenesis indicated the JTK-109 binding site was within the calicivirus RdRp thumb domain, in a pocket termed Site-B, which is highly-conserved within all calicivirus RdRps. Additionally, RdRp inhibition assays revealed that JTK-109 was antagonistic with the previously reported RdRp inhibitor pyridoxal-5'-phosphate-6-(2'-naphthylazo-6'-nitro-4',8'-disulfonate) tetrasodium salt (PPNDS), that also binds to Site-B. Moreover, like JTK-109, PPNDS was also a potent inhibitor of polymerases from six viruses spanning the three Caliciviridae genera tested (IC 50 range: 0.1-2.3 μM). Together, this study demonstrates the potential for de novo development of broad-spectrum antivirals that target the highly-conserved RdRp thumb pocket, Site-B. We also revealed three broad-spectrum HCV NNIs that could be used as antiviral scaffolds for further development against caliciviruses and other viruses., (Copyright © 2017 Elsevier B.V. All rights reserved.)