Synthesis and in vitro Evaluation of West Nile Virus Protease Inhibitors Based on the 2‐{6‐[2‐(5‐Phenyl‐4H‐{1,2,4]triazol‐3‐ylsulfanyl)acetylamino]benzothiazol‐2‐ylsulfanyl}acetamide Scaffold

In recent years, clinical symptoms resulting from West Nile virus (WNV) infection have worsened in severity, with an increased frequency in neuroinvasive diseases among the elderly. As there are presently no successful therapies against WNV for use in humans, continual efforts to develop new chemotherapeutics against this virus are highly desired. The viral NS2B‐NS3 protease is a promising target for viral inhibition due to its importance in viral replication and its unique substrate preference. In this study, a WNV NS2B‐NS3 protease inhibitor with a 2‐{6‐[2‐(5‐phenyl‐4H‐[1,2,4]triazol‐3‐ylsulfanyl)acetylamino]benzothiazol‐2‐ylsulfanyl}acetamide scaffold was identified during screening. Optimization of this initial hit by synthesis and screening of a focused compound library with this scaffold led to the identification of a novel uncompetitive inhibitor ( 1 a24 , IC₅₀=3.4±0.2 μM ) of the WNV NS2B‐NS3 protease. Molecular docking of 1 a24 into the WNV protease showed that the compound interferes with productive interactions of the NS2B cofactor with the NS3 protease and is an allosteric inhibitor of the WNV NS3 protease.     

Discovery, synthesis, and in vitro evaluation of West Nile virus protease inhibitors based on the 9,10-dihydro-3H,4aH-1,3,9,10a-tetraazaphenanthren-4-one scaffold

West Nile virus (WNV), a member of the Flaviviridae family, is a mosquito‐borne pathogen that causes a great number of human infections each year. Neither vaccines nor antiviral therapies are currently available for human use. In this study, a WNV NS2B–NS3 protease inhibitor with a 9,10‐dihydro‐3H,4aH‐1,3,9,10a‐tetraazaphenanthren‐4‐one scaffold was identified by screening a small library of non‐peptidic compounds. This initial hit was optimized by solution‐phase synthesis and screening of a focused library of compounds bearing this scaffold. This led to the identification of a novel, uncompetitive inhibitor ( 1a40 , IC₅₀=5.41±0.45 μM ) of WNV NS2B–NS3 protease. Molecular docking of this chiral compound onto the WNV protease indicates that the S enantiomer of 1a40 appears to interfere with the productive interactions between the NS2B cofactor and the NS3 protease domain; (S)‐ 1a40 is a preferred isomer for inhibition of WNV NS3 protease.     

Development and Characterization of New Peptidomimetic Inhibitors of the West Nile Virus NS2B–NS3 Protease

A series of new substrate analogue inhibitors of the WNV NS2B–NS3 protease containing decarboxylated arginine mimetics at the P1 position was developed. Among the various analogues, trans‐(4‐guanidino)cyclohexylmethylamide (GCMA) was identified as the most suitable P1 residue. In combination with dichloro‐substituted phenylacetyl groups at the P4 position, three inhibitors with inhibition constants of <0.2 μM were obtained. These GCMA inhibitors have a better selectivity profile than the previously described agmatine analogues, and possess negligible affinity for the trypsin‐like serine proteases thrombin, factor Xa, and matriptase. A crystal structure in complex with the WNV protease was determined for one of the most potent inhibitors, 3,4‐dichlorophenylacetyl‐Lys‐Lys‐GCMA (K_i=0.13 μM ). The inhibitor adopts a horseshoe‐like conformation, most likely due to a hydrophobic contact between the P4 phenyl ring and the P1 cyclohexyl group, which is further stabilized by an intramolecular hydrogen bond between the P1 guanidino group and the P4 carbonyl oxygen atom. These inhibitors are stable, readily accessible, and have a noncovalent binding mode. Therefore, they may serve as suitable lead structures for further development.     

The Discovery of Potent Nonstructural Protein 5A (NS5A) Inhibitors with a Unique Resistance Profile—Part 1

Nonstructural protein 5A (NS5A) represents a novel target for the treatment of hepatitis C virus (HCV). Daclatasvir, recently reported by Bristol–Myers–Squibb, is a potent NS5A inhibitor currently under investigation in phase 3 clinical trials. While the performance of daclatasvir has been impressive, the emergence of resistance could prove problematic and as such, improved analogues are being sought. By varying the biphenyl‐imidazole unit of daclatasvir, novel inhibitors of HCV NS5A were identified with an improved resistance profile against mutant strains of the virus while retaining the picomolar potency of daclatasvir. One compound in particular, methyl ((S)‐1‐((S)‐2‐(4‐(4‐(6‐(2‐((S)‐1‐((methoxycarbonyl)‐L ‐valyl)pyrrolidin‐2‐yl)‐1H‐imidazol‐5‐yl)quinoxalin‐2‐yl)phenyl)‐1H‐imidazol‐2‐yl)pyrrolidin‐1‐yl)‐3‐methyl‐1‐oxobutan‐2‐yl)carbamate ( 17 ), exhibited very promising activity and showed good absorption and a long predicted human pharmacokinetic half‐life. This compound represents a promising lead that warrants further evaluation.     

Structure-Based Macrocyclization of Substrate Analogue NS2B-NS3 Protease Inhibitors of Zika,West Nile and Dengue viruses

A series of cyclic active-site-directed inhibitors of the NS2B-NS3 proteases from Zika (ZIKV), West Nile (WNV), and dengue-4 (DENV4) viruses has been designed. The most potent compounds contain a reversely incorporated d -lysine residue in the P1 position. Its side chain is connected to the P2 backbone, its α-amino group is converted into a guanidine to interact with the conserved Asp129 side chain in the S1 pocket, and its C terminus is connected to the P3 residue via different linker segments. The most potent compounds inhibit the ZIKV protease with K_i values <5 nM. Crystal structures of seven ZIKV protease inhibitor complexes were determined to support the inhibitor design. All the cyclic compounds possess high selectivity against trypsin-like serine proteases and furin-like proprotein convertases. Both WNV and DENV4 proteases are inhibited less efficiently. Nonetheless, similar structure-activity relationships were observed for these enzymes, thus suggesting their potential application as pan-flaviviral protease inhibitors.     

1‐(1H‐Indol‐3‐yl)ethanamine Derivatives as Potent Staphylococcus aureus NorA Efflux Pump Inhibitors

The synthesis of 37 1‐(1H‐indol‐3‐yl)ethanamine derivatives, including 12 new compounds, was achieved through a series of simple and efficient chemical modifications. These indole derivatives displayed modest or no intrinsic anti‐staphylococcal activity. By contrast, several of the compounds restored, in a concentration‐dependent manner, the antibacterial activity of ciprofloxacin against Staphylococcus aureus strains that were resistant to fluoroquinolones due to overexpression of the NorA efflux pump. Structure–activity relationships studies revealed that the indolic aldonitrones halogenated at position 5 of the indole core were the most efficient inhibitors of the S. aureus NorA efflux pump. Among the compounds, (Z)‐N‐benzylidene‐2‐(tert‐butoxycarbonylamino)‐1‐(5‐iodo‐1H‐indol‐3‐yl)ethanamine oxide led to a fourfold decrease of the ciprofloxacin minimum inhibitory concentration against the SA‐1199B strain when used at a concentration of 0.5 mg L ^?1. To the best of our knowledge, this activity is the highest reported to date for an indolic NorA inhibitor. In addition, a new antibacterial compound, tert‐butyl (2‐(3‐hydroxyureido)‐2‐(1H‐indol‐3‐yl)ethyl)carbamate, which is not toxic for human cells, was also found.     

Structural Determinants of the Selectivity of 3‐Benzyluracil‐1‐acetic Acids toward Human Enzymes Aldose Reductase and AKR1B10

The human enzymes aldose reductase (AR) and AKR1B10 have been thoroughly explored in terms of their roles in diabetes, inflammatory disorders, and cancer. In this study we identified two new lead compounds, 2‐(3‐(4‐chloro‐3‐nitrobenzyl)‐2,4‐dioxo‐3,4‐dihydropyrimidin‐1(2H)‐yl)acetic acid (JF0048, 3 ) and 2‐(2,4‐dioxo‐3‐(2,3,4,5‐tetrabromo‐6‐methoxybenzyl)‐3,4‐dihydropyrimidin‐1(2H)‐yl)acetic acid (JF0049, 4 ), which selectively target these enzymes. Although 3 and 4 share the 3‐benzyluracil‐1‐acetic acid scaffold, they have different substituents in their aryl moieties. Inhibition studies along with thermodynamic and structural characterizations of both enzymes revealed that the chloronitrobenzyl moiety of compound 3 can open the AR specificity pocket but not that of the AKR1B10 cognate. In contrast, the larger atoms at the ortho and/or meta positions of compound 4 prevent the AR specificity pocket from opening due to steric hindrance and provide a tighter fit to the AKR1B10 inhibitor binding pocket, probably enhanced by the displacement of a disordered water molecule trapped in a hydrophobic subpocket, creating an enthalpic signature. Furthermore, this selectivity also occurs in the cell, which enables the development of a more efficient drug design strategy: compound 3 prevents sorbitol accumulation in human retinal ARPE‐19 cells, whereas 4 stops proliferation in human lung cancer NCI‐H460 cells.     

Development of 3‐Phenyl‐N‐(2‐(3‐phenylureido)ethyl)‐thiophene‐2‐sulfonamide Compounds as Inhibitors of Antiapoptotic Bcl‐2 Family Proteins

Antiapoptotic Bcl‐2 family proteins, such as Bcl‐x_L, Bcl‐2, and Mcl‐1, are often overexpressed in tumor cells, which contributes to tumor cell resistance to chemotherapies and radiotherapies. Inhibitors of these proteins thus have potential applications in cancer treatment. We discovered, through structure‐based virtual screening, a lead compound with micromolar binding affinity to Mcl‐1 (inhibition constant (K_i)=3 μM ). It contains a phenyltetrazole and a hydrazinecarbothioamide moiety, and it represents a structural scaffold not observed among known Bcl‐2 inhibitors. This work presents the structural optimization of this lead compound. By following the scaffold‐hopping strategy, we have designed and synthesized a total of 82 compounds in three sets. All of the compounds were evaluated in a fluorescence‐polarization binding assay to measure their binding affinities to Bcl‐x_L, Bcl‐2, and Mcl‐1. Some of the compounds with a 3‐phenylthiophene‐2‐sulfonamide core moiety showed sub‐micromolar binding affinities to Mcl‐1 (K_i=0.3–0.4 μM ) or Bcl‐2 (K_i≈1 μM ). They also showed obvious cytotoxicity on tumor cells (IC₅₀<10 μM ). Two‐dimensional heteronuclear single quantum coherence NMR spectra of three selected compounds, that is, YCW‐E5, YCW‐E10, and YCW‐E11, indicated that they bind to the BH3‐binding groove on Bcl‐x_L in a similar mode to ABT‐737. Several apoptotic assays conducted on HL‐60 cells demonstrated that these compounds are able to induce cell apoptosis through the mitochondrial pathway. We propose that the compounds with the 3‐phenylthiophene‐2‐sulfonamide core moiety are worth further optimization as effective apoptosis inducers with an interesting selectivity towards Mcl‐1 and Bcl‐2.     

Alzheimer’s Disease: Identification and Development of β‐Secretase (BACE‐1) Binding Fragments and Inhibitors by Dynamic Ligation Screening (DLS)

The application of dynamic ligation screening (DLS), a methodology for fragment‐based drug discovery (FBDD), to the aspartic protease β‐secretase (BACE‐1) is reported. For this purpose, three new fluorescence resonance energy transfer (FRET) substrates were designed and synthesized. Their kinetic parameters (V_max, K_M, and k_cat) were determined and compared with a commercial substrate. Secondly, a peptide aldehyde was designed as a chemically reactive inhibitor (CRI) based on the Swedish mutation substrate sequence. Incubation of this CRI with the protease, a FRET substrate, and one amine per well taken from an amine library, which was assembled by a maximum common substructure (MCS) approach, revealed the fragment 3‐(3‐aminophenyl)‐2H‐chromen‐2‐one ( 1 ) to be a competitive BACE‐1 inhibitor that enhanced the activity of the CRI. Irreversibly formed fragment combination products of 1 with the initial peptide sequence were active and confirmed the targeting of the active site through the ethane‐1,2‐diamine isostere. Finally, structure‐assisted combination of fragment 1 with secondary fragments that target the S1 site in hit optimization yielded novel, entirely fragment‐based BACE‐1 inhibitors with up to 30‐fold improved binding affinity. Interactions with the protein were explained by molecular modeling studies, which indicate that the new fragment combinations interact with the catalytic aspartic acid dyad, as well as with the adjacent binding sites required for potency.     

Glucocerebrosidase Enhancers for Selected Gaucher Disease Genotypes by Modification of α‐1‐C‐Substituted Imino‐D‐xylitols (DIXs) by Click Chemistry

A series of hybrid analogues was designed by combination of the iminoxylitol scaffold of parent 1C9‐DIX with triazolylalkyl side chains. The resulting compounds were considered potential pharmacological chaperones in Gaucher disease. The DIX analogues reported here were synthesized by CuAAC click chemistry from scaffold 1 (α‐1‐C‐propargyl‐1,5‐dideoxy‐1,5‐imino‐D ‐xylitol) and screened as imiglucerase inhibitors. A set of selected compounds were tested as β‐glucocerebrosidase (GBA1) enhancers in fibroblasts from Gaucher patients bearing different genotypes. A number of these DIX compounds were revealed as potent GBA1 enhancers in genotypes containing the G202R mutation, particularly compound DIX‐28 (α‐1‐C‐[(1‐(3‐trimethylsilyl)propyl)‐1H‐1,2,3‐triazol‐4‐yl)methyl]‐1,5‐dideoxy‐1,5‐imino‐D ‐xylitol), bearing the 3‐trimethylsilylpropyl group as a new surrogate of a long alkyl chain, with approximately threefold activity enhancement at 10 nM . Despite their structural similarities with isofagomine and with our previously reported aminocyclitols, the present DIX compounds behaved as non‐competitive inhibitors, with the exception of the mixed‐type inhibitor DIX‐28.     

Virtual Screening against p50 NF‐κB Transcription Factor for the Identification of Inhibitors of the NF‐κB–DNA Interaction and Expression of NF‐κB Upregulated Genes

Virtual screening against NF‐κB p50 using docking simulations was applied by starting from a three‐dimensional (3D) database containing more than 4.6 million commercially available structures. This database was filtered by specifying a subset of commercially available compounds sharing a (2E,Z)‐3‐(2‐hydroxyphenyl)‐2‐propenoate substructure and relevant druglike properties. Docking to p50 NF‐κB was performed with a test set of six known inhibitors of NF‐κB–DNA interactions. In agreement with docking results, the highest‐scored compound displayed a high level of inhibitory activity in electrophoretic mobility shift assay (EMSA) experiments (inhibition of NF‐κB–DNA interactions) and on biological functions dependent on NF‐κB activity (inhibition of IL‐8 gene expression in cystic fibrosis IB3‐1 cells). We found that this in silico screening approach is suitable for the identification of low‐molecular‐weight compounds that inhibit NF‐κB–DNA interactions and NF‐κB‐dependent functions. Information deduced from the discovery of the new lead compound and its binding mode could result in further lead optimization resulting in more potent NF‐κB inhibitors.     

Optimization of Thienopyrrole‐Based Finger‐Loop Inhibitors of the Hepatitis C Virus NS5B Polymerase

Infections caused by the hepatitis C virus (HCV) are a significant world health problem for which novel therapies are in urgent demand. The NS5B polymerase of HCV is responsible for the replication of viral RNA and has been a prime target in the search for novel treatment options. We had discovered allosteric finger‐loop inhibitors based on a thieno[3,2‐b]pyrrole scaffold as an alternative to the related indole inhibitors. Optimization of the thienopyrrole series led to several N‐acetamides with submicromolar potency in the cell‐based replicon assay, but they lacked oral bioavailability in rats. By linking the N4‐position to the ortho‐position of the C5‐aryl group, we were able to identify the tetracyclic thienopyrrole 40 , which displayed a favorable pharmacokinetic profile in rats and dogs and is equipotent with recently disclosed finger‐loop inhibitors based on an indole scaffold.     

GluN2B‐Selective N‐Methyl‐d‐aspartate (NMDA) Receptor Antagonists Derived from 3‐Benzazepines: Synthesis and Pharmacological Evaluation of Benzo[7]annulen‐7‐amines

Given their high neuroprotective potential, ligands that block GluN2B‐containing N‐methyl‐D ‐aspartate (NMDA) receptors by interacting with the ifenprodil binding site located on the GluN2B subunit are of great interest for the treatment of various neuronal disorders. In this study, a novel class of GluN2B‐selective NMDA receptor antagonists with the benzo[7]annulene scaffold was prepared and pharmacologically evaluated. The key intermediate, N‐(2‐methoxy‐5‐oxo‐6,7,8,9‐tetrahydro‐5H‐benzo[7]annulen‐7‐yl)acetamide ( 11 ), was obtained by cyclization of 3‐acetamido‐5‐(3‐methoxyphenyl)pentanoic acid ( 10 b ). The final reaction steps comprise hydrolysis of the amide, reduction of the ketone, and reductive alkylation, leading to cis‐ and trans‐configured 7‐(ω‐phenylalkylamino)benzo[7]annulen‐5‐ols. High GluN2B affinity was observed with cis‐configured γ‐amino alcohols substituted with a 3‐phenylpropyl moiety at the amino group. Removal of the benzylic hydroxy moiety led to the most potent GluN2B antagonists of this series: 2‐methoxy‐N‐(3‐phenylpropyl)‐6,7,8,9‐tetrahydro‐5H‐benzo[7]annulen‐7‐amine ( 20 a , K_i=10 nM ) and 2‐methoxy‐N‐methyl‐N‐(3‐phenylpropyl)‐6,7,8,9‐tetrahydro‐5H‐benzo[7]annulen‐7‐amine ( 23 a , K_i=7.9 nM ). The selectivity over related receptors (phencyclidine binding site of the NMDA receptor, σ₁ and σ₂ receptors) was recorded. In a functional assay measuring the cytoprotective activity of the benzo[7]annulenamines, all tested compounds showed potent NMDA receptor antagonistic activity. Cytotoxicity induced via GluN2A subunit‐containing NMDA receptors was not inhibited by the new ligands.     

8‐Benzyltetrahydropyrazino[2,1‐f]purinediones: Water‐Soluble Tricyclic Xanthine Derivatives as Multitarget Drugs for Neurodegenerative Diseases

8‐Benzyl‐substituted tetrahydropyrazino[2,1‐f]purinediones were designed as tricyclic xanthine derivatives containing a basic nitrogen atom in the tetrahydropyrazine ring to improve water solubility. A library of 69 derivatives was prepared and evaluated in radioligand binding studies at adenosine receptor (AR) subtypes and for their ability to inhibit monoamine oxidases (MAO). Potent dual‐target‐directed A₁/A_2A adenosine receptor antagonists were identified. Several compounds showed triple‐target inhibition; one of the best compounds was 8‐(2,4‐dichloro‐5‐fluorobenzyl)‐1,3‐dimethyl‐6,7,8,9‐tetrahydropyrazino[2,1‐f]purine‐2,4(1H,3H)‐dione ( 72 ) (human AR: K_i A₁ 217 nM , A_2A 233 nM ; IC₅₀ MAO‐B: 508 nM ). Dichlorinated compound 36 [8‐(3,4‐dichlorobenzyl)‐1,3‐dimethyl‐6,7,8,9‐tetrahydropyrazino[2,1‐f]purine‐2,4(1H,3H)‐dione] was found to be the best triple‐target drug in rat (K_i A₁ 351 nM , A_2A 322 nm; IC₅₀ MAO‐B: 260 nM ), and may serve as a useful tool for preclinical proof‐of‐principle studies. Compounds that act at multiple targets relevant for symptomatic as well as disease‐modifying treatment of neurodegenerative diseases are expected to show advantages over single‐target therapeutics.     

Discovery of MK‐8742: An HCV NS5A Inhibitor with Broad Genotype Activity

The NS5A protein plays a critical role in the replication of HCV and has been the focus of numerous research efforts over the past few years. NS5A inhibitors have shown impressive in vitro potency profiles in HCV replicon assays, making them attractive components for inclusion in all oral combination regimens. Early work in the NS5A arena led to the discovery of our first clinical candidate, MK‐4882 [2‐((S)‐pyrrolidin‐2‐yl)‐5‐(2‐(4‐(5‐((S)‐pyrrolidin‐2‐yl)‐1H‐imidazol‐2‐yl)phenyl)benzofuran‐5‐yl)‐1H‐imidazole]. While preclinical proof‐of‐concept studies in HCV‐infected chimpanzees harboring chronic genotype 1 infections resulted in significant decreases in viral load after both single‐ and multiple‐dose treatments, viral breakthrough proved to be a concern, thus necessitating the development of compounds with increased potency against a number of genotypes and NS5A resistance mutations. Modification of the MK‐4882 core scaffold by introduction of a cyclic constraint afforded a series of tetracyclic inhibitors, which showed improved virologic profiles. Herein we describe the research efforts that led to the discovery of MK‐8742, a tetracyclic indole‐based NS5A inhibitor, which is currently in phase 2b clinical trials as part of an all‐oral, interferon‐free regimen for the treatment of HCV infection.     

Synthesis and Biological Evaluation of 9‐Oxo‐9H‐indeno[1,2‐b]pyrazine‐2,3‐dicarbonitrile Analogues as Potential Inhibitors of Deubiquitinating Enzymes

High‐throughput screening highlighted 9‐oxo‐9H‐indeno[1,2‐b]pyrazine‐2,3‐dicarbonitrile ( 1 ) as an active inhibitor of ubiquitin‐specific proteases (USPs), a family of hydrolytic enzymes involved in the removal of ubiquitin from protein substrates. The chemical behavior of compound 1 was examined. Moreover, the synthesis and in vitro evaluation of new compounds, analogues of 1 , led to the identification of potent and selective inhibitors of the deubiquitinating enzyme USP8.     

Structural Studies on 4,5‐Disubstituted 2‐Aminoimidazole‐Based Biofilm Modulators that Suppress Bacterial Resistance to β‐Lactams

A library of 4,5‐disubstituted 2‐aminoimidazole triazole amide (2‐AITA) conjugates has been successfully assembled. Upon biological screening, this class of small molecules was discovered as enhanced biofilm regulators through non‐microbicidal mechanisms against methicillin‐resistant Staphylococcus aureus (MRSA) and multidrug‐resistant Acinetobacter baumannii (MDRAB), with active concentrations in the low micromolar range. The library was also subjected to synergism and resensitization studies with β‐lactam antibiotics against MRSA. Lead compounds were identified that suppress the antibiotic resistance of MRSA by working synergistically with oxacillin, a β‐lactam antibiotic resistant to penicillinase. A further structure–activity relationship (SAR) study on the parent 2‐AITA compound delivered a 2‐aminoimidazole diamide (2‐AIDA) conjugate with significantly increased synergistic activity with oxacillin against MRSA, decreasing the MIC value of the β‐lactam antibiotic by 64‐fold. Increased anti‐biofilm activity did not necessarily lead to increased suppression of antibiotic resistance, which indicates that biofilm inhibition and resensitization are most likely occurring via distinct mechanisms.     

para‐Substituted 2‐Phenyl‐3,4‐dihydroquinazolin‐4‐ones As Potent and Selective Tankyrase Inhibitors

Human tankyrases are attractive drug targets, especially for the treatment of cancer. We identified a set of highly potent tankyrase inhibitors based on a 2‐phenyl‐3,4‐dihydroquinazolin‐4‐one scaffold. Substitutions at the para position of the scaffold′s phenyl group were evaluated as a strategy to increase potency and improve selectivity. The best compounds displayed single‐digit nanomolar potencies, and profiling against several human diphtheria‐toxin‐like ADP‐ribosyltransferases revealed that a subset of these compounds are highly selective tankyrase inhibitors. The compounds also effectively inhibit Wnt signaling in HEK293 cells. The binding mode of all inhibitors was studied by protein X‐ray crystallography. This allowed us to establish a structural basis for the development of highly potent and selective tankyrase inhibitors based on the 2‐phenyl‐3,4‐dihydroquinazolin‐4‐one scaffold and outline a rational approach to the modification of other inhibitor scaffolds that bind to the nicotinamide site of the catalytic domain.     

6,7‐Dimethoxy‐2‐{2‐[4‐(1H‐1,2,3‐triazol‐1‐yl)phenyl]ethyl}‐1,2,3,4‐tetrahydroisoquinolines as Superior Reversal Agents for P‐Glycoprotein‐Mediated Multidrug Resistance

P‐glycoprotein (P‐gp)‐mediated multidrug resistance (MDR) is a major obstacle for successful cancer chemotherapy. Based on our previous study, 17 novel compounds with the 6,7‐dimethoxy‐2‐{2‐[4‐(1H‐1,2,3‐triazol‐1‐yl)phenyl]ethyl}‐1,2,3,4‐tetrahydroisoquinoline scaffold were designed and synthesized. Among them, 2‐[(1‐{4‐[2‐(6,7‐dimethoxy‐3,4‐dihydroisoquinolin‐2(1H)‐yl)ethyl]phenyl}‐1H‐1,2,3‐triazol‐4‐yl)methoxy]‐N‐(p‐tolyl)benzamide (compound 7 h ) was identified as a potent modulator of P‐gp‐mediated MDR, with high potency (EC₅₀=127.5±9.1 nM ), low cytotoxicity (TI>784.3), and long duration (>24 h) in reversing doxorubicin (DOX) resistance in K562/A02 cells. Compound 7 h also enhanced the effects of other MDR‐related cytotoxic agents (paclitaxel, vinblastine, and daunorubicin), increased the accumulation of DOX and blocked P‐gp‐mediated rhodamine 123 efflux function in K562/A02 MDR cells. Moreover, 7 h did not have any effect on cytochrome (CYP3A4) activity. These results indicate that 7 h is a relatively safe modulator of P‐gp‐mediated MDR that has good potential for further development.     

Design,Synthesis, and Evaluation of 2‐Amino‐6‐nitrobenzothiazole‐Derived Hydrazones as MAO Inhibitors: Role of the Methylene Spacer Group

A series of 2‐amino‐6‐nitrobenzothiazole‐derived extended hydrazones were designed, synthesized, and investigated for their ability to inhibit monoamine oxidase A and B (MAO‐A/MAO‐B). The compounds were found to exhibit inhibitory activities in the nanomolar to micromolar range. Some of the compounds showed excellent potency and selectivity against the MAO‐B isoform. N′‐(5‐Chloro‐2‐oxoindolin‐3‐ylidene)‐2‐(6‐nitrobenzothiazol‐2‐ylamino)acetohydrazide (compound 31 ) showed the highest MAO‐B inhibitory activity (IC₅₀=1.8±0.3 nm , selectivity index [SI]=766.67), whereas compound 6 [N′‐(1‐(4‐bromophenyl)ethylidene)‐2‐(6‐nitrobenzothiazol‐2‐ylamino)acetohydrazide] was found to be the most active MAO‐A inhibitor (IC₅₀=0.42±0.003 μm ). Kinetic studies revealed that compounds 6 and 31 exhibit competitive‐type reversible inhibition against both MAO‐A and MAO‐B, respectively. Structure–activity relationship (SAR) studies disclosed several structural aspects significant for potency and the contribution of the methylene spacer toward MAO‐B inhibitory potency, with minimal or no neurotoxicity. Molecular modeling studies yielded a good correlation between experimental and theoretical inhibitory data. Binding pose analysis revealed the significance of cumulative effects of π–π stacking and hydrogen bond interactions for effective stabilization of virtual ligand–protein complexes. Further optimization studies of compound 31 , including co‐crystallization of inhibitor–MAO‐B complexes, are essential to develop these compounds as potential therapeutic agents for MAO‐B‐associated neurodegenerative diseases.