Indole-3-Glycerol Phosphate Synthase From Mycobacterium tuberculosis: A Potential New Drug Target

Tuberculosis (TB), caused by the pathogen Mycobacterium tuberculosis, affects millions of people worldwide. Several TB drugs have lost efficacy due to emerging drug resistance and new anti-TB targets are needed. Recent research suggests that indole-3-glycerol phosphate synthase (IGPS) in M. tuberculosis (MtIGPS) could be such a target. IGPS is a (β/α)₈-barrel enzyme that catalyzes the conversion of 1-(o-carboxyphenylamino)-1-deoxyribulose 5’-phosphate (CdRP) into indole-glycerol-phosphate (IGP) in the bacterial tryptophan biosynthetic pathway. M. tuberculosis over expresses the tryptophan pathway genes during an immune response and inhibition of MtIGPS allows CD4 T-cells to more effectively fight against M. tuberculosis. Here we review the published data on MtIGPS expression, kinetics, mechanism, and inhibition. We also discuss MtIGPS crystal structures and compare them to other IGPS structures to reveal potential structure-function relationships of interest for the purposes of drug design and biocatalyst engineering.     

Development of Potent Inhibitors of the Mycobacterium tuberculosis Virulence Factor Zmp1 and Evaluation of Their Effect on Mycobacterial Survival inside Macrophages

The enzyme Zmp1 is a zinc‐containing peptidase that plays a critical role in the pathogenicity of Mycobacterium tuberculosis. Herein we describe the identification of a small set of Zmp1 inhibitors based on a novel 8‐hydroxyquinoline‐2‐hydroxamate scaffold. Among the synthesized compounds, N‐(benzyloxy)‐8‐hydroxyquinoline‐2‐carboxamide ( 1 c ) was found to be the most potent Zmp1 inhibitor known to date, and its binding mode was analyzed both by kinetics studies and molecular modeling, identifying critical interactions of 1 c with the zinc ion and residues in the active site. The effect of 1 c on intracellular Mycobacterium survival was assayed in J774 murine macrophages infected with M. tuberculosis H37Rv or M. bovis BCG and human monocyte‐derived macrophages infected with M. tuberculosis H37Rv. Cytotoxicity and genotoxicity were also assessed. Overall, inhibitor 1 c displays interesting in vitro antitubercular properties worthy of further investigation.     

Development of Novel Selective Peptidomimetics Containing a Boronic Acid Moiety,Targeting the 20S Proteasome as Anticancer Agents

This paper describes the design, synthesis, and biological evaluation of peptidomimetic boronates as inhibitors of the 20S proteasome, a validated target in the treatment of multiple myeloma. The synthesized compounds showed a good inhibitory profile against the ChT‐L activity of 20S proteasome. Compounds bearing a β‐alanine residue at the P2 position were the most active, that is, 3‐ethylphenylamino and 4‐methoxyphenylamino (R)‐1‐{3‐[4‐(substituted)‐2‐oxopyridin‐1(2H)‐yl]propanamido}‐3‐methylbutylboronic acids ( 3 c and 3 d , respectively), and these derivatives showed inhibition constants (K_i) of 17 and 20 nM , respectively. In addition, they co‐inhibited post glutamyl peptide hydrolase activity ( 3 c , K_i=2.57 μM ; 3 d , K_i=3.81 μM ). No inhibition was recorded against the bovine pancreatic α‐chymotrypsin, which thus confirms the selectivity towards the target enzyme. Docking studies of 3 c and related inhibitors into the yeast proteasome revealed the structural basis for specificity. The evaluation of growth inhibitory effects against 60 human tumor cell lines was performed at the US National Cancer Institute. Among the selected compounds, 3 c showed 50 % growth inhibition (GI₅₀) values at the sub‐micromolar level on all cell lines.     

Inhibition of Cytokine Release by Mycobacterium tuberculosis Phenolic Glycolipid Analogues

Infection by Mycobacterium tuberculosis causes tuberculosis, a disease characterized by alteration of host innate and adaptive immunity. These processes are mediated by a series of bacterial biomolecules, among which phenolic glycolipids (PGLs) and the related p‐hydroxybenzoic acid derivatives have been suggested to play important roles. To probe the importance of structural features of these glycans on cytokine modulation, we synthesized three M. tuberculosis PGL analogues ( 1 – 3 ), which differ from the native glycoconjugates by possessing a simplified lipid algycone. The ability of 1 – 3 to modulate the release of proinflammatory cytokines (TNF‐α, IL‐1β, IL‐6, MCP‐1) and nitric oxide (NO) was evaluated. None of the compounds stimulated the secretion of these signalling molecules. However, all showed a Toll‐like Receptor 2‐mediated, concentration‐dependent inhibition profile that was related to the methylation pattern on the glycan.     

Synthesis of Novel Derivatives of 5,6,7,8-Tetrahydroquinazolines Using α-Aminoamidines and In Silico Screening of Their Biological Activity

α-Aminoamidines are promising reagents for the synthesis of a diverse family of pyrimidine ring derivatives. Here, we demonstrate the use of α-aminoamidines for the synthesis of a new series of 5,6,7,8-tetrahydroquinazolines by their reaction with bis-benzylidene cyclohexanones. The reaction occurs in mild conditions and is characterized by excellent yields. It has easy workup, as compared to the existing methods of tetrahydroquinazoline preparation. Newly synthesized derivatives of 5,6,7,8-tetrahydroquinazoline bear protecting groups at the C2-tert-butyl moiety of a quinazoline ring, which can be easily cleaved, opening up further opportunities for their functionalization. Moreover, molecular docking studies indicate that the synthesized compounds reveal high binding affinity toward some essential enzymes of Mycobacterial tuberculosis, such as dihydrofolate reductase (DHFR), pantothenate kinase (MtPanK), and FAD-containing oxidoreductase DprE1 (MtDprE1), so that they may be promising candidates for the molecular design and the development of new antitubercular agents against multidrug-resistant strains of the Tubercle bacillus. Finally, the high inhibition activity of the synthesized compounds was also predicted against β-glucosidase, suggesting a novel tetrahydroquinazoline scaffold for the treatment of diabetes.     

Development of New and Selective Trypanosoma cruzi trans‐Sialidase Inhibitors from Sulfonamide Chalcones and Their Derivatives

A series of sulfonamide‐containing hydroxylated chalcone ( 4 – 7 ) and quinolinone ( 8 , 9 ) derivatives was synthesised and tested for inhibition of the trans‐sialidase from Trypanosoma cruzi (TcTS). IC₅₀ values for these inhibitors ranged from 0.6 to 7.3 μM , with the dihydroxylated (catechol) derivatives being the tightest binders. Full kinetic analyses of inhibition were performed for these catechol derivatives, both for the transglycosylation reaction in the presence of lactose and for the hydrolysis reaction in its absence. Competitive inhibition was seen in each case with K_i values for 5 , 7 and 9 of 2.0, 2.2 and 0.2 μM , respectively, in the absence of lactose, and 4.6, 3.7 and 0.4 μM in its presence. None of the compounds tested showed any significant inhibition of the human sialidase Neu2, at concentrations up to 200 μM .     

A Bisbenzamidine Phosphonate as a Janus‐faced Inhibitor for Trypsin‐like Serine Proteases

A hybrid approach was applied for the design of an inhibitor of trypsin‐like serine proteases. Compound 16 [(R,R)‐ and (R,S)‐diphenyl (4‐(1‐(4‐amidinobenzylamino)‐1‐oxo‐3‐phenylpropan‐2‐ylcarbamoyl)phenylamino)(4‐amidinophenyl)methylphosphonate hydrochloride], prepared in a convergent synthetic procedure, possesses a phosphonate warhead prone to react with the active site serine residue in a covalent, irreversible manner. Each of the two benzamidine moieties of 16 can potentially be accommodated in the S1 pocket of the target enzyme, but only the benzamidine close to the phosphonate group would then promote an irreversible interaction. The Janus‐faced inhibitor 16 was evaluated against several serine proteases and caused a pronounced inactivation of human thrombin with a second‐order rate constant (k_inac/K_i) of 59 500 M ^?1 s^?1. With human matriptase, 16 showed preference for a reversible mode of inhibition (IC₅₀=2.6 μM ) as indicated by linear progress curves and enzyme reactivation.     

Design,Synthesis, and Biological Evaluation of (S)‐Valine Thiazole‐Derived Cyclic and Noncyclic Peptidomimetic Oligomers as Modulators of Human P‐Glycoprotein (ABCB1)

Multidrug resistance caused by ATP binding cassette transporter P‐glycoprotein (P‐gp) through extrusion of anticancer drugs from the cells is a major cause of failure in cancer chemotherapy. Previously, selenazole‐containing cyclic peptides were reported as P‐gp inhibitors and were also used for co‐crystallization with mouse P‐gp, which has 87 % homology to human P‐gp. It has been reported that human P‐gp can simultaneously accommodate two to three moderately sized molecules at the drug binding pocket. Our in silico analysis, based on the homology model of human P‐gp, spurred our efforts to investigate the optimal size of (S)‐valine‐derived thiazole units that can be accommodated at the drug binding pocket. Towards this goal, we synthesized varying lengths of linear and cyclic derivatives of (S)‐valine‐derived thiazole units to investigate the optimal size, lipophilicity, and structural form (linear or cyclic) of valine‐derived thiazole peptides that can be accommodated in the P‐gp binding pocket and affects its activity, previously an unexplored concept. Among these oligomers, lipophilic linear ( 13 ) and cyclic trimer ( 17 ) derivatives of QZ59S‐SSS were found to be the most and equally potent inhibitors of human P‐gp (IC₅₀=1.5 μM ). As the cyclic trimer and linear trimer compounds are equipotent, future studies should focus on noncyclic counterparts of cyclic peptides maintaining linear trimer length. A binding model of the linear trimer 13 within the drug binding site on the homology model of human P‐gp represents an opportunity for future optimization, specifically replacing valine and thiazole groups in the noncyclic form.     

Triazolophthalazines: Easily Accessible Compounds with Potent Antitubercular Activity

Tuberculosis (TB) remains one of the major causes of death worldwide, in particular because of the emergence of multidrug‐resistant TB. Herein we explored the potential of an alternative class of molecules as anti‐TB agents. Thus, a series of novel 3‐substituted triazolophthalazines was quickly and easily prepared from commercial hydralazine hydrochloride as starting material and were further evaluated for their antimycobacterial activities and cytotoxicities. Four of the synthesized compounds were found to effectively inhibit the Mycobacterium tuberculosis (M.tb) H₃₇Rv strain with minimum inhibitory concentration (MIC) values <10 μg mL^?1, whereas no compounds displayed cytotoxicity against HCT116 human cell lines (IC₅₀>100 μm ). More remarkably, the most potent compounds proved to be active to a similar extent against various multidrug‐resistant M.tb strains, thus uncovering a mode of action distinct from that of standard antitubercular agents. Overall, their ease of preparation, combined with their attractive antimycobacterial activities, make such triazolophthalazine‐based derivatives promising leads for further development.     

Synthesis,Biological Evaluation of 1,1‐Diarylethylenes as a Novel Class of Antimitotic Agents

The cytotoxic activities of 23 new isocombretastatin A derivatives with modifications on the B‐ring were investigated. Several compounds exhibited excellent antiproliferative activity at nanomolar concentrations against a panel of human cancer cell lines. Compounds isoFCA‐4 ( 2 e ), isoCA‐4 ( 2 k ) and isoNH₂CA‐4 ( 2 s ) were the most cytotoxic, and strongly inhibited tubulin polymerization with IC₅₀ values of 4, 2 and 1.5 μM , respectively. These derivatives were found to be 10‐fold more active than phenstatin and colchicine with respect to growth inhibition but displayed similar activities as tubulin polymerization inhibitors. In addition, cell cycle arrest in the G₂/M phase and subsequent apoptosis was observed in three cancer cell lines when treated with these compounds. The disruptive effect of 2 e , 2 k and 2 s on the vessel‐like structures formed by human umbilical vein endothelial cells (HUVEC) suggest that these compounds may act as vascular disrupting agents. Both compounds 2 k and 2 s have the potential for further prodrug modification and development as vascular disrupting agents for treatment of solid tumors.     

Indanones As High‐Potency Reversible Inhibitors of Monoamine Oxidase

Recent reports document that α‐tetralone (3,4‐dihydro‐2H‐naphthalen‐1‐one) is an appropriate scaffold for the design of high‐potency monoamine oxidase (MAO) inhibitors. Based on the structural similarity between α‐tetralone and 1‐indanone, the present study involved synthesis of 34 1‐indanone and related indane derivatives as potential inhibitors of recombinant human MAO‐A and MAO‐B. The results show that C6‐substituted indanones are particularly potent and selective MAO‐B inhibitors, with IC₅₀ values ranging from 0.001 to 0.030 μM . C5‐Substituted indanone and indane derivatives are comparatively weaker MAO‐B inhibitors. Although the 1‐indanone and indane derivatives are selective inhibitors of the MAO‐B isoform, a number of homologues are also potent MAO‐A inhibitors, with three homologues possessing IC₅₀ values <0.1 μM . Dialysis of enzyme–inhibitor mixtures further established a selected 1‐indanone as a reversible MAO inhibitor with a competitive mode of inhibition. It may be concluded that 1‐indanones are promising leads for the design of therapies for neurodegenerative and neuropsychiatric disorders such as Parkinson’s disease and depression.     

Targeting a Targeted Drug: An Approach Toward Hypoxia‐Activatable Tyrosine Kinase Inhibitor Prodrugs

Tyrosine kinase inhibitors (TKIs), which have revolutionized cancer therapy over the past 15 years, are limited in their clinical application due to serious side effects. Therefore, we converted two approved TKIs (sunitinib and erlotinib) into 2‐nitroimidazole‐based hypoxia‐activatable prodrugs. Kinetics studies showed very different stabilities over 24 h; however, fast reductive activation via E. coli nitroreductase could be confirmed for both panels. The anticancer activity and signaling inhibition of the compounds against various human cancer cell lines were evaluated in cell culture. These data, together with molecular docking simulations, revealed distinct differences in the impact of structural modifications on drug binding to the enzymes: whereas the catalytic pocket of the epidermal growth factor receptor (EGFR) accepted all new erlotinib derivatives, the vascular endothelial growth factor receptor (VEGFR)‐inhibitory potential in the case of the sunitinib prodrugs was dramatically diminished by derivatization. In line, hypoxia dependency of ERK signaling inhibition was observed with the sunitinib prodrugs, while oxygen levels had no impact on the activity of the erlotinib derivatives. Overall, proof of principle could be shown for this concept, and the results obtained are an important basis for the future development of tyrosine kinase inhibitor prodrugs.     

Benzofuroxan Derivatives as Potent Agents against Multidrug-Resistant Mycobacterium tuberculosis

Tuberculosis (TB) is currently the leading cause of death related to infectious diseases worldwide, as reported by the World Health Organization. Moreover, the increasing number of multidrug-resistant tuberculosis (MDR-TB) cases has alarmed health agencies, warranting extensive efforts to discover novel drugs that are effective and also safe. In this study, 23 new compounds were synthesized and evaluated in vitro against the drug-resistant strains of M. tuberculosis. The compound 6-((3-fluoro-4-thiomorpholinophenyl)carbamoyl)benzo[c][1,2,5]oxadiazole 1-N-oxide ( 5 b ) was particularly remarkable in this regard as it demonstrated MIC₉₀ values below 0.28 μM against all the MDR strains evaluated, thus suggesting that this compound might have a different mechanism of action. Benzofuroxans are an attractive new class of anti-TB agents, exemplified by compound 5 b , with excellent potency against the replicating and drug-resistant strains of M. tuberculosis.     

Probing the Peptidylglycine α‐Hydroxylating Monooxygenase Active Site with Novel 4‐Phenyl‐3‐butenoic Acid Based Inhibitors

Specific inhibition of the copper‐containing peptidylglycine α‐hydroxylating monooxygenase (PHM), which catalyzes the post‐translational modification of peptides involved in carcinogenesis and tumor progression, constitutes a new approach for combating cancer. We carried out a structure–activity study of new compounds derived from a well‐known PHM substrate analogue, the olefinic compound 4‐phenyl‐3‐butenoic acid (PBA). We designed, synthesized, and tested various PBA derivatives both in vitro and in silico. We show that it is possible to increase PBA affinity for PHM by appropriate functionalization of its aromatic nucleus. Compound 2 d , for example, bears a meta‐benzyloxy substituent, and exhibits better inhibition features (K_i=3.9 μM , k_inact/K_i=427 M ^?1 s^?1) than the parent PBA (K_i=19 μM , k_inact/K_i=82 M ^?1 s^?1). Docking calculations also suggest two different binding modes for PBA derivatives; these results will aid in the development of further PHM inhibitors with improved features.     

Biological Evaluation of Potent Triclosan‐Derived Inhibitors of the Enoyl–Acyl Carrier Protein Reductase InhA in Drug‐Sensitive and Drug‐Resistant Strains of Mycobacterium tuberculosis

New triclosan (TRC) analogues were evaluated for their activity against the enoyl–acyl carrier protein reductase InhA in Mycobacterium tuberculosis (Mtb). TRC is a well‐known inhibitor of InhA, and specific modifications to its positions 5 and 4′ afforded 27 derivatives; of these compounds, seven derivatives showed improved potency over that of TRC. These analogues were active against both drug‐susceptible and drug‐resistant Mtb strains. The most active compound in this series, 4‐(n‐butyl)‐1,2,3‐triazolyl TRC derivative 3 , had an MIC value of 0.6 μg mL^?1 (1.5 μM ) against wild‐type Mtb. At a concentration equal to its MIC, this compound inhibited purified InhA by 98 %, and showed an IC₅₀ value of 90 nM . Compound 3 and the 5‐methylisoxazole‐modified TRC 14 were able to inhibit the biosynthesis of mycolic acids. Furthermore, mc²4914, an Mtb strain overexpressing inhA, was found to be less susceptible to compounds 3 and 14 , supporting the notion that InhA is the likely molecular target of the TRC derivatives presented herein.     

3-substituted 2-phenylimidazo[2,1-b]benzothiazoles: synthesis, anticancer activity, and inhibition of tubulin polymerization

A new series of 3‐substituted 2‐phenylimidazo[2,1‐b]benzothiazoles ( 3 a – h ) were synthesized by C‐arylation of 2‐arylimidazo[2,1‐b]benzothiazoles using palladium acetate as catalyst, and the resulting compounds were evaluated for their anticancer activity. Compounds 3 a , 3 e , and 3 h exhibited good antiproliferative activity, with GI₅₀ values in the range of 0.19–83.1 μM . Compound 3 h showed potent anticancer efficacy against 60 human cancer cell lines, with a mean GI₅₀ value of 0.88 μM . This compound also induced cell‐cycle arrest in the G₂/M phase and inhibited tubulin polymerization followed by activation of caspase‐3 and apoptosis. A high‐throughput tubulin polymerization assay showed that the level of inhibition for compound 3 h is similar to that of combretastatin A‐4. Molecular modeling studies provided a molecular basis for the favorable binding of compounds 3 a , 3 e , and 3 h to the colchicine binding pocket of tubulin.     

Chemoenzymatic Synthesis of GDP‐L‐Fucose Derivatives as Potent and Selective α‐1,3‐Fucosyltransferase Inhibitors

Fucosyltransferases (FucTs) usually catalyze the final step of glycosylation and are critical to many biological processes. High levels of specific FucT activities are often associated with various cancers. Here we report the development of a chemoenzymatic method for synthesizing a library of guanosine diphosphate β‐L ‐fucose (GDP‐Fuc) derivatives, followed by in situ screening for inhibitory activity against bacterial and human α‐1,3‐FucTs. Several compounds incorporating appropriate hydrophobic moieties were identified from the initial screening. These were individually synthesized, purified and characterized in detail for their inhibition kinetics. Compound 5 had a K_i of 29 nM for human FucT‐VI, and is 269 and 11 times more selective than for Helicobacter pylori FucT (K_i=7.8 μM) and for human FucT‐V (K_i=0.31 μM).     

Inhibitors of the salicylate synthase (MbtI) from Mycobacterium tuberculosis discovered by high-throughput screening

A simple steady‐state kinetic high‐throughput assay was developed for the salicylate synthase MbtI from Mycobacterium tuberculosis, which catalyzes the first committed step of mycobactin biosynthesis. The mycobactins are small‐molecule iron chelators produced by M. tuberculosis, and their biosynthesis has been identified as a promising target for the development of new antitubercular agents. The assay was miniaturized to a 384‐well plate format and high‐throughput screening was performed at the National Screening Laboratory for the Regional Centers of Excellence in Biodefense and Emerging Infectious Diseases (NSRB). Three classes of compounds were identified comprising the benzisothiazolones (class I), diarylsulfones (class II), and benzimidazole‐2‐thiones (class III). Each of these compound series was further pursued to investigate their biochemical mechanism and structure–activity relationships. Benzimidazole‐2‐thione 4 emerged as the most promising inhibitor owing to its potent reversible inhibition.     

Comparative Binding Energy (COMBINE) Analysis for Understanding the Binding Determinants of Type II Dehydroquinase Inhibitors

Herein we report comparative binding energy (COMBINE) analyses to derive quantitative structure–activity relationship (QSAR) models that help rationalize the determinants of binding affinity for inhibitors of type II dehydroquinase (DHQ2), the third enzyme of the shikimic acid pathway. Independent COMBINE models were derived for Helicobacter pylori and Mycobacterium tuberculosis DHQ2, which is an essential enzyme in both these pathogenic bacteria that has no counterpart in human cells. These studies quantify the importance of the hydrogen bonding interactions between the ligands and the water molecule involved in the DHQ2 reaction mechanism. They also highlight important differences in the ligand interactions with the interface pocket close to the active site that could provide guides for future inhibitor design.     

Hybrids from Farnesylthiosalicylic Acid and Hydroxamic Acid as Dual Ras‐Related Signaling and Histone Deacetylase (HDAC) Inhibitors: Design,Synthesis and Biological Evaluation

A novel series of hybrids was designed and synthesized by combining key elements from farnesylthiosalicylic acid (FTS) and hydroxamic acid. Several 3,7,11‐trimethyldodeca‐2,6,10‐trien‐1‐yl) thio)benzamide derivatives, particularly those with branched and linear aliphatic linkers between the hydroxamic zinc binding group (ZBG) and the benzamide core, not only displayed significant antitumor activities against six human cancer cells but also exhibited histone deacetylase (HDAC) inhibitory effects in vitro. Among them, N‐(4‐(hydroxyamino)‐4‐oxobutyl)‐2‐(((2E,6E)‐3,7,11‐trimethyldodeca‐2,6, 10‐trien‐1‐yl)thio)benzamide ( 8 d ) was the most potent, with IC₅₀ values of 4.9–7.6 μM ; these activities are eight‐ to sixteen‐fold more potent than FTS and comparable to that of suberoylanilide hydroxamic acid (SAHA). Derivative 8 d induced cell cycle arrest in the G0/G1 phase, inhibited the acetylation of histone H3 and α‐tubulin, and blocked Ras‐related signaling pathways in a dose‐dependent manner. The improved tumor growth inhibition and cell‐cycle arrest in vitro might result from the dual inhibition. These findings suggest dual inhibitors of Ras‐related signaling pathway and HDAC hold promise as therapeutic agents for the treatment of cancer.