Enzyme mediated-transesterification of verbascoside and evaluation of antifungal activity of synthesised compounds

<div><p>Enzymatic acylation of verbascoside, a polyhydroxylated natural product, has been reported in this study using five different commercial lipases and taking <i>p</i>-nitrophenyl alkanoates as acyl donors. Out of these enzymes, the immobilised <i>Candida antarctica</i> lipase B was found as the enzyme of choice. Mono- and di-acylated products were formed, with mono as major product indicating high regioselective nature of such transformations. A series of acyl esters of verbascoside have been synthesised by this enzymatic transesterification methodology. The lipophilicity of the synthesised analogues was also checked. The analogues were further subjected to synergistic antifungal activity with amphotericin B (AmB) against <i>Candida albicans.</i> Fourfold reduction in minimum inhibitory concentration of AmB was observed with few synthesised analogues such as verbascoside 4″-octanoate (<b>3b</b>), verbascoside 4″-palmitate (<b>3d</b>) and verbascoside 4″,4′-dipalmitate (<b>4d</b>) at a concentration of 0.5 μg/mL.</p></div

Computationally Guided Identification of Novel <i>Mycobacterium tuberculosis</i> GlmU Inhibitory Leads, Their Optimization, and in Vitro Validation

<i>Mycobacterium tuberculosis</i> (Mtb) infections are causing serious health concerns worldwide. Antituberculosis drug resistance threatens the current therapies and causes further need to develop effective antituberculosis therapy. GlmU represents an interesting target for developing novel Mtb drug candidates. It is a bifunctional acetyltransferase/uridyltransferase enzyme that catalyzes the biosynthesis of UDP-<i>N</i>-acetyl-glucosamine (UDP-GlcNAc) from glucosamine-1-phosphate (GlcN-1-P). UDP-GlcNAc is a substrate for the biosynthesis of lipopolysaccharide and peptidoglycan that are constituents of the bacterial cell wall. In the current study, structure and ligand based computational models were developed and rationally applied to screen a drug-like compound repository of 20 000 compounds procured from ChemBridge DIVERSet database for the identification of probable inhibitors of Mtb GlmU. The in vitro evaluation of the in silico identified inhibitor candidates resulted in the identification of 15 inhibitory leads of this target. Literature search of these leads through SciFinder and their similarity analysis with the PubChem training data set (AID 1376) revealed the structural novelty of these hits with respect to Mtb GlmU. IC₅₀ of the most potent identified inhibitory lead (5810599) was found to be 9.018 ± 0.04 μM. Molecular dynamics (MD) simulation of this inhibitory lead (5810599) in complex with protein affirms the stability of the lead within the binding pocket and also emphasizes on the key interactive residues for further designing. Binding site analysis of the acetyltransferase pocket with respect to the identified structural moieties provides a thorough analysis for carrying out the lead optimization studies

Benzothiazole Derivative as a Novel <i>Mycobacterium tuberculosis</i> Shikimate Kinase Inhibitor: Identification and Elucidation of Its Allosteric Mode of Inhibition

<i>Mycobacterium tuberculosis</i> shikimate kinase (Mtb-SK) is a key enzyme involved in the biosynthesis of aromatic amino acids through the shikimate pathway. Since it is proven to be essential for the survival of the microbe and is absent from mammals, it is a promising target for anti-TB drug discovery. In this study, a combined approach of <i>in silico</i> similarity search and pharmacophore building using already reported inhibitors was used to screen a procured library of 20,000 compounds of the commercially available ChemBridge database. From the <i>in silico</i> screening, 15 hits were identified, and these hits were evaluated <i>in vitro</i> for Mtb-SK enzyme inhibition. Two compounds presented significant enzyme inhibition with IC₅₀ values of 10.69 ± 0.9 and 46.22 ± 1.2 μM. The best hit was then evaluated for the <i>in vitro</i> mode of inhibition where it came out to be an uncompetitive and noncompetitive inhibitor with respect to shikimate (SKM) and ATP, respectively, suggesting its binding at an allosteric site. Potential binding sites of Mtb-SK were identified which confirmed the presence of an allosteric binding pocket apart from the ATP and SKM binding sites. The docking simulations were performed at this pocket in order to find the mode of binding of the best hit in the presence of substrates and the products of the enzymatic reaction. Molecular dynamics (MD) simulations elucidated the probability of inhibitor binding at the allosteric site in the presence of ADP and shikimate-3-phosphate (S-3-P), that is, after the formation of products of the reaction. The inhibitor binding may prevent the release of the product from Mtb-SK, thereby inhibiting its activity. The binding stability and the key residue interactions of the inhibitor to this product complex were also revealed by the MD simulations. Residues ARG43, ILE45, and PHE57 were identified as crucial that were involved in interactions with the best hit. This is the first report of an allosteric binding site of Mtb-SK, which could largely address the selectivity issue associated with kinase inhibitors

Nitrofuranyl Methyl Piperazines as New Anti-TB Agents: Identification, Validation, Medicinal Chemistry, and PK Studies

Whole-cell screening of 20,000 drug-like small molecules led to the identification of nitrofuranyl methylpiperazines as potent anti-TB agents. In the present study, validation followed by medicinal chemistry has been used to explore the structure–activity relationship. Ten compounds demonstrated potent MIC in the range of 0.17–0.0072 μM against H₃₇Rv <i>Mycobacterium tuberculosis</i> (MTB) and were further investigated against nonreplicating and resistant (Rif^R and MDR) strains of MTB. These compounds were also tested for cytotoxicity. Among the 10 tested compounds, five showed submicromolar to nanomolar potency against nonreplicating and resistant (Rif^R and MDR) strains of MTB along with a good safety index. Based on their overall <i>in vitro</i> profiles, the solubility and pharmacokinetic properties of five potent compounds were studied, and two analogues, <b>14f</b> and <b>16g</b>, were found to have comparatively better solubility than others tested and acceptable pharmacokinetic properties. This study presents the rediscovery of a nitrofuranyl class of compounds with improved aqueous solubility and acceptable oral PK properties, opening a new direction for further development

Synthesis and Biological Evaluation of Polar Functionalities Containing Nitrodihydroimidazooxazoles as Anti-TB Agents

Novel polar functionalities containing 6-nitro-2,3-dihydroimidazooxazole (NHIO) analogues were synthesized to produce a compound with enhanced solubility. Polar functionalities including sulfonyl, uridyl, and thiouridyl-bearing NHIO analogues were synthesized and evaluated against <i>Mycobacterium tuberculosis</i> (MTB) H₃₇Rv. The aqueous solubility of compounds with MIC values ≤0.5 μg/mL were tested, and six compounds showed enhanced aqueous solubility. The best six compounds were further tested against resistant (Rif^R and MDR) and dormant strains of MTB and tested for cytotoxicity in HepG2 cell line. Based on its overall <i>in vitro</i> characteristics and solubility profile, compound <b>6d</b> was further shown to possess high microsomal stability, solubility under all tested biological conditions (PBS, SGF and SIF), and favorable oral <i>in vivo</i> pharmacokinetics and <i>in vivo</i> efficacy