RESUMO
Leishmaniasis is a highly prevalent, yet neglected disease caused by protozoan parasites of the genus Leishmania. In the search for newer, safer, and more effective antileishmanial compounds, we herein present a study of the mode of action in addition to a detailed structural and biological characterization of LQOF-G6 [N-benzoyl-N'-benzyl-Nâ³-(4-tertbutylphenyl)guanidine]. X-ray crystallography and extensive NMR experiments revealed that LQOF-G6 nearly exclusively adopts the Z conformation stabilized by an intramolecular hydrogen bond. The investigated guanidine showed selective inhibitory activity on Leishmania major cysteine protease LmCPB2.8ΔCTE (CPB) with ~73% inhibition and an IC50-CPB of 6.0 µM. This compound did not show any activity against the mammalian homologues cathepsin L and B. LQOF-G6 has been found to be nontoxic toward both organs and several cell lines, and no signs of hepatotoxicity or nephrotoxicity were observed from the analysis of biochemical clinical plasma markers in the treated mice. Docking simulations and experimental NMR measurements showed a clear contribution of the conformational parameters to the strength of the binding in the active site of the enzyme, and thus fit the differences in the inhibition values of LQOF-G6 compared to the other guanidines. Furthermore, the resulting data render LQOF-G6 suitable for further development as an antileishmanial drug.
Assuntos
Cisteína Proteases , Leishmania major , Leishmaniose , Animais , Camundongos , Cisteína Proteases/metabolismo , Guanidina , Virulência , Leishmaniose/tratamento farmacológico , Mamíferos/metabolismoRESUMO
The species Streptomyces venezuelae is represented by several distinct strains with variable abilities to biosynthesize structurally diverse secondary metabolites. In this work, we examined the effect of ethanol shock on the transcriptome and metabolome of Streptomyces venezuelae NRRL B-65442 using high-throughput RNA sequencing (RNA-seq) and high-resolution liquid chromatography-tandem mass spectrometry (LC-MS/MS). Ethanol shock caused massive changes in the gene expression profile, differentially affecting genes for secondary metabolite biosynthesis and central metabolic pathways. Most of the data from the transcriptome analysis correlated well with the metabolome changes, including the overproduction of jadomycin congeners and a downshift in the production of desferrioxamines, legonoxamine, foroxymithin, and a small cryptic ribosomally synthesized peptide. Some of the metabolome changes, such as the overproduction of chloramphenicol, could not be explained by overexpression of the cognate biosynthetic genes but correlated with the expression profiles of genes for precursor biosynthesis. Changes in the transcriptome were also observed for several genes known to play a role in stress response in other bacteria and included at least 10 extracytoplasmic function σ factors. This study provides important new insights into the stress response in antibiotic-producing bacteria and will help to understand the complex mechanisms behind the environmental factor-induced regulation of secondary metabolite biosynthesis. IMPORTANCE Streptomyces spp. are filamentous Gram-positive bacteria known as versatile producers of secondary metabolites, of which some have been developed into human medicines against infections and cancer. The genomes of these bacteria harbor dozens of gene clusters governing the biosynthesis of secondary metabolites (BGCs), of which most are not expressed under laboratory conditions. Detailed knowledge of the complex regulation of BGC expression is still lacking, although certain growth conditions are known to trigger the production of previously undetected secondary metabolites. In this work, we investigated the effect of ethanol shock on the production of secondary metabolites by Streptomyces venezuelae and correlated these findings with the expression of cognate BGCs and primary metabolic pathways involved in the generation of cofactors and precursors. The findings of this study set the stage for the rational manipulation of bacterial genomes aimed at enhanced production of industrially important bioactive natural products.
Assuntos
Streptomyces , Transcriptoma , Humanos , Etanol/metabolismo , Cromatografia Líquida , Espectrometria de Massas em Tandem , Streptomyces/metabolismo , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga EscalaRESUMO
Dopamine is an important neurotransmitter that regulates numerous essential functions, including cognition and voluntary movement. As such, it serves as an important scaffold for synthesis of novel analogues as part of drug development effort to obtain drugs for treatment of neurodegenerative diseases, such as Parkinson's disease. To that end, similarity search of the ZINC database based on two known dopamine-1 receptor (D1R) agonists, dihydrexidine (DHX) and SKF 38393, respectively, was used to predict novel chemical entities with potential binding to D1R. Three compounds that showed the highest similarity index were selected for synthesis and bioactivity profiling. All main synthesis products as well as the isolated intermediates, were properly characterized. The physico-chemical analyses were performed using HRESIMS, GC/MS, LC/MS with UV-Vis detection, and FTIR, 1H NMR and 13C NMR spectroscopy. Binding to D1 and D2 receptors and inhibition of dopamine reuptake via dopamine transporter were measured for the synthesized analogues of DHX and SKF 38393.
Assuntos
Catecolaminas , Receptores de Dopamina D1 , 2,3,4,5-Tetra-Hidro-7,8-Di-Hidroxi-1-Fenil-1H-3-Benzazepina/farmacologia , Fenantridinas/farmacologia , Receptores de Dopamina D1/metabolismoRESUMO
In a survey of plants from Ecuador with antiprotozoal activity, Cupania cinerea was found to show significant in vitro activity against the Plasmodium falciparum K1 strain and Trypanosoma brucei rhodesiense. Subsequently, activity-guided isolation of the n-hexane and dichloromethane extracts from the bark of C. cinerea afforded two diterpene glycosides (1 and 2), named cupacinoside and 6'-de-O-acetylcupacinoside, and a lactonized triterpene bearing an oxepin moiety named cupacinoxepin (3), together with the known compounds scopoletin (4), caryophyllene oxide (5), two bisabolane sesquiterpenes (6 and 7), lichexanthone (8), gustastatin (9), lupenone (10), betulone (11), 17ß,21ß-epoxyhopan-3-one (12), taraxerol (13), and taraxerone (14). For compound 3, X-ray crystallography was employed to elucidate the relative configuration. For cupacinosides (1) and (2) and cupacinoxepin (3), in vitro activities against the P. falciparum K1 strain (IC(50)1, 1.3; 2, 1.8; and 3, 8.7 µM) and T. b. rhodesiense (IC(50)1, 4.5; 2, 15.8; and 3, 71.6 µM) were found. Cytotoxicity toward L-6 cells is discussed for all the compounds isolated.