N-Pyrazinylhydroxybenzamides as biologically active compounds: a hit-expansion study and antimicrobial evaluation.

Background: The development of novel antimicrobial drugs is an essential part of combatting the uprising of antimicrobial resistance. Proper hit-to-lead development is crucially needed. Methods & results: We present a hit-expansion study of N-pyrazinyl- and N-pyridyl-hydroxybenzamides with a comprehensive determination of structure-activity relationships. The antimicrobial screening revealed high selectivity to staphylococci along with antimycobacterial activity with the best value of 6.25 µg/ml against Mycobacterium tuberculosis H37Rv. We proved an inhibition of proteosynthesis and a membrane depolarization of methicillin-resistant Staphylococcus aureus. Conclusion: Our results are a good starting point for further development of new antimicrobial compounds, where the next step would be tuning the potential between relatively nonspecific membrane depolarization effect and specific inhibition of proteosynthesis.

Design, Synthesis and Antimicrobial Evaluation of New N-(1-Hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-6-yl)(hetero)aryl-2-carboxamides as Potential Inhibitors of Mycobacterial Leucyl-tRNA Synthetase.

Tuberculosis remains a serious killer among infectious diseases due to its incidence, mortality, and occurrence of resistant mycobacterial strains. The challenge to discover new antimycobacterial agents forced us to prepare a series of N-(1-hydroxy-1,3-dihydrobenzo[c][1,2]oxaborol-6-yl)(hetero)aryl-2-carboxamides 1-19 via the acylation of 6-aminobenzo[c][1,2]oxaborol-1(3H)-ol with various activated (hetero)arylcarboxylic acids. These novel compounds have been tested in vitro against a panel of clinically important fungi and bacteria, including mycobacteria. Some of the compounds inhibited the growth of mycobacteria in the range of micromolar concentrations and retained this activity also against multidrug-resistant clinical isolates. Half the maximal inhibitory concentrations against the HepG2 cell line indicated an acceptable toxicological profile. No growth inhibition of other bacteria and fungi demonstrated selectivity of the compounds against mycobacteria. The structure-activity relationships have been derived and supported with a molecular docking study, which confirmed a selectivity toward the potential target leucyl-tRNA synthetase without an impact on the human enzyme. The presented compounds can become important materials in antimycobacterial research.

Hybridization Approach Toward Novel Antituberculars: Design, Synthesis, and Biological Evaluation of Compounds Combining Pyrazinamide and 4-Aminosalicylic Acid.

Apart from the SARS-CoV-2 virus, tuberculosis remains the leading cause of death from a single infectious agent according to the World Health Organization. As part of our long-term research, we prepared a series of hybrid compounds combining pyrazinamide, a first-line antitubercular agent, and 4-aminosalicylic acid (PAS), a second-line agent. Compound 11 was found to be the most potent, with a broad spectrum of antimycobacterial activity and selectivity toward mycobacterial strains over other pathogens. It also retained its in vitro activity against multiple-drug-resistant mycobacterial strains. Several structural modifications were attempted to improve the in vitro antimycobacterial activity. The δ-lactone form of compound 11 (11') had more potent in vitro antimycobacterial activity against Mycobacterium tuberculosis H37Rv. Compound 11 was advanced for in vivo studies, where it was proved to be nontoxic in Galleria mellonella and zebrafish models, and it reduced the number of colony-forming units in spleens in the murine model of tuberculosis. Biochemical studies showed that compound 11 targets mycobacterial dihydrofolate reductases (DHFR). An in silico docking study combined with molecular dynamics identified a viable binding mode of compound 11 in mycobacterial DHFR. The lactone 11' opens in human plasma to its parent compound 11 (t1/2 = 21.4 min). Compound 11 was metabolized by human liver fraction by slow hydrolysis of the amidic bond (t1/2 = 187 min) to yield PAS and its starting 6-chloropyrazinoic acid. The long t1/2 of compound 11 overcomes the main drawback of PAS (short t1/2 necessitating frequent administration of high doses of PAS).

Comprehensive insight into anti-staphylococcal and anti-enterococcal action of brominated and chlorinated pyrazine-based chalcones.

The greatest threat and medicinal impact within gram-positive pathogens are posed by two bacterial genera, Staphylococcus and Enterococcus. Chalcones have a wide range of biological activities and are recognized as effective templates in medicinal chemistry. This study provides comprehensive insight into the anti-staphylococcal and anti-enterococcal activities of two recently published brominated and chlorinated pyrazine-based chalcones, CH-0y and CH-0w. Their effects against 4 reference and 12 staphylococcal and enterococcal clinical isolates were evaluated. Bactericidal action, the activity in combination with selected conventional antibiotics, the study of post-antimicrobial effect (PAE, PAE/SME), and in vitro and in vivo toxicity, were included. In CH-0y, anti-staphylococcal activity ranging from MIC = 15.625 to 62.5 µM, and activity against E. faecium from 31.25 to 62.5 µM was determined. In CH-0w, anti-staphylococcal activity ranging from 31.25 to 125 µM, and activity against E. faecium and E. faecalis (62.5 µM) was revealed. Both CH-0y and CH-0w showed bactericidal action, beneficial impact on bacterial growth delay within PAE and PAE/SME studies, and non/low toxicity in vivo. Compared to CH-0w, CH-0y seems to have higher anti-staphylococcal and less toxic potential. In conclusion, chalcones CH-0y and CH-0w could be considered as structural pattern for future adjuvants to selected antibiotic drugs.

Enantioselective Construction of Spirooxindole-Fused Cyclopentanes.

The present study reports an asymmetric organocatalytic cascade reaction of oxindole derivates with α,ß-unsaturated aldehydes efficiently catalyzed by simple chiral secondary amine. Spirooxindole-fused cyclopentanes were produced in excellent isolated yields (up to 98%) with excellent enantiopurities (up to 99% ee) and moderate to high diastereoselectivities. The synthetic utility of the protocol was exemplified on a set of additional transformations of the corresponding spiro compounds. In addition, a study showing the promising biological activity of selected enantioenriched products was accomplished.

Development of 3,5-Dinitrophenyl-Containing 1,2,4-Triazoles and Their Trifluoromethyl Analogues as Highly Efficient Antitubercular Agents Inhibiting Decaprenylphosphoryl-ß-d-ribofuranose 2'-Oxidase.

We report herein the discovery of 3,5-dinitrophenyl 1,2,4-triazoles with excellent and selective antimycobacterial activities against Mycobacterium tuberculosis strains, including clinically isolated multidrug-resistant strains. Thorough structure-activity relationship studies of 3,5-dinitrophenyl-containing 1,2,4-triazoles and their trifluoromethyl analogues revealed the key role of the position of the 3,5-dinitrophenyl fragment in the antitubercular efficiency. Among the prepared compounds, the highest in vitro antimycobacterial activities against M. tuberculosis H37Rv and against seven clinically isolated multidrug-resistant strains of M. tuberculosis were found with S-substituted 4-alkyl-5-(3,5-dinitrophenyl)-4H-1,2,4-triazole-3-thiols and their 3-nitro-5-(trifluoromethyl)phenyl analogues. The minimum inhibitory concentrations of these compounds reached 0.03 µM, which is superior to all the current first-line anti-tuberculosis drugs. Furthermore, almost all compounds with excellent antimycobacterial activities exhibited very low in vitro cytotoxicities against two proliferating mammalian cell lines. The docking study indicated that these compounds acted as the inhibitors of decaprenylphosphoryl-ß-d-ribofuranose 2'-oxidase enzyme, which was experimentally confirmed by two independent radiolabeling experiments.

Design, Synthesis and Evaluation of N-pyrazinylbenzamides as Potential Antimycobacterial Agents.

Three series of N-(pyrazin-2-yl)benzamides were designed as retro-amide analogues of previously published N-phenylpyrazine-2-carboxamides with in vitro antimycobacterial activity. The synthesized retro-amides were evaluated for in vitro growth inhibiting activity against Mycobacterium tuberculosis H37Rv (Mtb), three non-tuberculous mycobacterial strains (M. avium, M. kansasii, M. smegmatis) and selected bacterial and fungal strains of clinical importance. Regarding activity against Mtb, most N-pyrazinylbenzamides (retro-amides) possessed lower or no activity compared to the corresponding N-phenylpyrazine-2-carboxamides with the same substitution pattern. However, the active retro-amides tended to have lower HepG2 cytotoxicity and better selectivity. Derivatives with 5-chloro substitution on the pyrazine ring were generally more active compared to their 6-cloro positional isomers or non-chlorinated analogues. The best antimycobacterial activity against Mtb was found in N-(5-chloropyrazin-2-yl)benzamides with short alkyl (2h: R² = Me; 2i: R² = Et) in position 4 of the benzene ring (MIC = 6.25 and 3.13 µg/mL, respectively, with SI > 10). N-(5-Chloropyrazin-2-ylbenzamides with hydroxy substitution (2b: R² = 2-OH; 2d: R² = 4-OH) on the benzene ring or their acetylated synthetic precursors possessed the broadest spectrum of activity, being active in all three groups of mycobacterial, bacterial and fungal strains. The substantial differences in in silico calculated properties (hydrogen-bond pattern analysis, molecular electrostatic potential, HOMO and LUMO) can justify the differences in biological activities between N-pyrazinylbenzamides and N-phenylpyrazine-2-carboxamides.

Design, synthesis and antimycobacterial activity of hybrid molecules combining pyrazinamide with a 4-phenylthiazol-2-amine scaffold.

Hybrid compounds based on a combination of the first-line antitubercular pyrazinamide (PZA) and a formerly identified antimycobacterial scaffold of 4-arylthiazol-2-amine were designed. Eighteen compounds were prepared, characterized and tested for in vitro growth inhibition activity against M. tuberculosis H37Rv, M. kansasii, M. avium and M. smegmatis by Microplate Alamar Blue Assay at neutral pH. Active compounds were tested for in vitro cytotoxicity in the human hepatocellular carcinoma cell line (HepG2). The most active 6-chloro-N-[4-(4-fluorophenyl)thiazol-2-yl]pyrazine-2-carboxamide (9b) also had the broadest spectrum of activity and inhibited M. tuberculosis, M. kansasii, and M. avium with MIC = 0.78 µg mL-1 (2.3 µM) and a selectivity index related to HepG2 cells of SI > 20. Structure-activity relationships within the series are discussed. Based on its structural similarity to known inhibitors and the results of a molecular docking study, we suggest mycobacterial beta-ketoacyl-(acyl-carrier-protein) synthase III (FabH) as a potential target.