RESUMO
Bioactive compound design based on natural product (NP) structure may be limited because of partial coverage of NP-like chemical space and biological target space. These limitations can be overcome by combining NP-centered strategies with fragment-based compound design through combination of NP-derived fragments to afford structurally unprecedented "pseudo-natural products" (pseudo-NPs). The design, synthesis, and biological evaluation of a collection of indomorphan pseudo-NPs that combine biosynthetically unrelated indole- and morphan-alkaloid fragments are described. Indomorphane derivative Glupin was identified as a potent inhibitor of glucose uptake by selectively targeting and upregulating glucose transporters GLUT-1 and GLUT-3. Glupin suppresses glycolysis, reduces the levels of glucose-derived metabolites, and attenuates the growth of various cancer cell lines. Our findings underscore the importance of dual GLUT-1 and GLUT-3 inhibition to efficiently suppress tumor cell growth and the cellular rescue mechanism, which counteracts glucose scarcity.
Assuntos
Produtos Biológicos/farmacologia , Proliferação de Células , Transportador de Glucose Tipo 1/antagonistas & inibidores , Transportador de Glucose Tipo 3/antagonistas & inibidores , Glucose/metabolismo , Morfinanos/síntese química , Neoplasias/tratamento farmacológico , Transporte Biológico , Ciclo Celular , Glicólise , Humanos , Células Tumorais CultivadasRESUMO
Due to their enhanced metabolic needs many cancers need a sufficient supply of glucose, and novel inhibitors of glucose import are in high demand. Cytochalasin B (CB) is a potent natural glucose import inhibitor which also impairs the actin cytoskeleton leading to undesired toxicity. With a view to identifying selective glucose import inhibitors we have developed an enantioselective trienamine catalyzed synthesis of a CB-inspired compound collection. Biological analysis revealed that indeed actin impairment can be distinguished from glucose import inhibition and led to the identification of the first selective glucose import inhibitor based on the basic structural architecture of cytochalasin B.
Assuntos
Alcenos/química , Aminas/química , Citocalasina B/química , Citocalasina B/farmacologia , Glucose/antagonistas & inibidores , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Catálise , Glucose/metabolismo , Células HCT116 , Células HeLa , Humanos , Estrutura Molecular , EstereoisomerismoRESUMO
In a recent study we discovered that a ring-fused 2-pyridone compound triggered fibrillization of a key protein in Parkinson's disease, α-synuclein. To reveal how variations in compound structure affect protein aggregation, we now prepared a number of strategic analogs and tested their effects on α-synuclein amyloid fiber formation in vitro. We find that, in contrast to the earlier templating effect, some analogs inhibit α-synuclein fibrillization. For both templating and inhibiting compounds, the key species formed in the reactions are α-synuclein oligomers that contain compound. Despite similar macroscopic appearance, the templating and inhibiting oligomers are distinctly different in secondary structure content. When the inhibitory oligomers are added in seed amounts, they inhibit fresh α-synuclein aggregation reactions. Our study demonstrates that small chemical changes to the same central fragment can result in opposite effects on protein aggregation.
Assuntos
Amiloide/antagonistas & inibidores , Amiloide/metabolismo , Antiparkinsonianos/química , Antiparkinsonianos/farmacologia , Piridonas/química , Piridonas/farmacologia , alfa-Sinucleína/metabolismo , Amiloide/química , Amiloide/ultraestrutura , Humanos , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/metabolismo , Estrutura Secundária de Proteína , alfa-Sinucleína/química , alfa-Sinucleína/ultraestruturaRESUMO
Small organic molecules that inhibit functional bacterial amyloid fibers, curli, are promising new antibiotics. Here we investigated the mechanism by which the ring-fused 2-pyridone FN075 inhibits fibrillation of the curli protein CsgA. Using a variety of biophysical techniques, we found that FN075 promotes CsgA to form off-pathway, non-amyloidogenic oligomeric species. In light of the generic properties of amyloids, we tested whether FN075 would also affect the fibrillation reaction of human α-synuclein, an amyloid-forming protein involved in Parkinson's disease. Surprisingly, FN075 stimulates α-synuclein amyloid fiber formation as measured by thioflavin T emission, electron microscopy (EM), and atomic force microscopy (AFM). NMR data on (15)N-labeled α-synuclein show that upon FN075 addition, α-synuclein oligomers with 7 nm radius form in which the C-terminal 40 residues remain disordered and solvent exposed. The polypeptides in these oligomers contain ß-like secondary structure, and the oligomers are detectable by AFM, EM, and size-exclusion chromatography (SEC). Taken together, FN075 triggers oligomer formation of both proteins: in the case of CsgA, the oligomers do not proceed to fibers, whereas for α-synuclein, the oligomers are poised to rapidly form fibers. We conclude that there is a fine balance between small-molecule inhibition and templation that depends on protein chemistry.
Assuntos
Amiloide/antagonistas & inibidores , Antibacterianos/química , Antibacterianos/farmacologia , Proteínas de Escherichia coli/metabolismo , Piridonas/química , Piridonas/farmacologia , alfa-Sinucleína/metabolismo , Amiloide/metabolismo , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Humanos , Doença de Parkinson/metabolismoRESUMO
Fluorescent multi-ring-fused 2-pyridones, with chemical resemblance to other biologically active 2-pyridone systems, were solubilized in spherical micelles formed by the ganglioside G(M1) and studied with respect to their spatial localization and rotational mobility. For this, electronic energy transfer between the multi-ring-fused 2-pyridone (donor) and BODIPY-FL-labeled G(M1) was determined, as well as their fluorescence depolarization. From the obtained efficiency of energy transfer to the acceptor group (BODIPY-FL), either localized in the polar or in the nonpolar part of the ganglioside, it has been possible to estimate the most likely localization of the multi-ring-fused 2-pyridones. The center of mass of the studied multi-ring-fused 2-pyridones are located at approximately 33 Å from the micellar center of mass, which corresponds to the internal hydrophobic-hydrophilic interfacial region. At this location, the reorienting rates of the multi-ring-fused 2-pyridones are surprisingly slow with typical correlation times of 35-55 ns. No evidence was found for the formation of ground and excited state dimers, even when two monomers were forced to be near each other via a short covalent linker.
Assuntos
Gangliosídeo G(M1)/química , Micelas , Piridonas/química , Portadores de Fármacos/química , Corantes Fluorescentes/química , Modelos Moleculares , Conformação Molecular , Solubilidade , Espectrometria de FluorescênciaRESUMO
UV-VIS absorption and fluorescence spectroscopic properties of six related polyaromatic 2-pyridones have been studied. Excitation of the lowest and rather weak and structure-less transition [ε (max) (430 nm) ≈ 3,000 mol(-1)dm(3)cm(-1)] gives rise to a broad fluorescence band in the visible region, for these compounds. These S (0) â S (1) transitions are compatible with symmetrically forbidden transitions, promoted by intensity borrowing, as is revealed by fluorescence depolarisation data. With one exception, all compounds exhibit strong fluorescence, with quantum yields in glycerol varying between 40% and 70%. The corresponding fluorescence lifetimes range from 11 ns to 17 ns, while the radiative lifetimes are very similar (≈26 ns), for all compounds. Interestingly and rarely observed, the calculated radiative lifetimes for the weak absorption band are significantly longer, i.e. between 37 and 40 ns.
Assuntos
Fluorescência , Piridonas/química , Estrutura Molecular , Piridonas/síntese química , Teoria Quântica , Espectrometria de Fluorescência , Espectrofotometria UltravioletaRESUMO
Finding a new drug candidate for a selected target is an expensive and time-consuming process. Target guided-synthesis, or in situ click chemistry, is a concept where the drug target is used to template the formation of its own inhibitors from reactive building blocks. This could simplify the identification of drug candidates. However, with the exception of one example of an RNA-target, target-guided synthesis has always employed purified targets. This limits the number of targets that can be screened by the method. By applying methods from the field of metabolomics, we demonstrate that target-guided synthesis with protein targets also can be performed directly in cell-based systems. These methods offer new possibilities to conduct screening for drug candidates of difficult protein targets in cellular environments.
RESUMO
An emerging method to help elucidate the mode of action of experimental drugs is to use untargeted metabolomics of cell-systems. The interpretations of such screens are however complex and more examples with inhibitors of known targets are needed. Here two T-cell lines were treated with an inhibitor of aspartate aminotransferase and analyzed with untargeted GC-MS. The interpretation of the data was enhanced by the use of two different cell-lines and supports aspartate aminotransferase as a target. In addition, the data suggest an unexpected off-target effect on glutamate decarboxylase. The results exemplify the potency of metabolomics to provide insight into both mode of action and off-target effects of drug candidates.
Assuntos
Aspartato Aminotransferases/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Hidrazinas/farmacologia , Metaboloma/efeitos dos fármacos , Succinatos/farmacologia , Linfócitos T/efeitos dos fármacos , Aspartato Aminotransferases/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Cromatografia Gasosa-Espectrometria de Massas , Glutamato Descarboxilase/metabolismo , Humanos , Células Jurkat , Metabolômica , Linfócitos T/metabolismoRESUMO
Enteric bacteria assemble functional amyloid fibers, curli, on their surfaces that share structural and biochemical properties with disease-associated amyloids. Here, we test rationally designed 2-pyridone compounds for their ability to alter amyloid formation of the major curli subunit CsgA. We identified several compounds that discourage CsgA amyloid formation and several compounds that accelerate CsgA amyloid formation. The ability of inhibitor compounds to stop growing CsgA fibers was compared to the same property of the CsgA chaperone, CsgE. CsgE blocked CsgA amyloid assembly and arrested polymerization when added to actively polymerizing fibers. Additionally, CsgE and the 2-pyridone inhibitors prevented biofilm formation by Escherichia coli at the air-liquid interface of a static culture. We demonstrate that curli amyloid assembly and curli-dependent biofilm formation can be modulated not only by protein chaperones, but also by "chemical chaperones."
Assuntos
Amiloide/química , Biofilmes/efeitos dos fármacos , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/farmacologia , Proteínas de Membrana Transportadoras/farmacologia , Chaperonas Moleculares/farmacologia , Piridonas/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Amiloide/metabolismo , Biofilmes/crescimento & desenvolvimento , Desenho de Fármacos , Escherichia coli/efeitos dos fármacos , Escherichia coli/metabolismo , Escherichia coli/fisiologia , Proteínas de Escherichia coli/metabolismo , Cinética , Multimerização Proteica/efeitos dos fármacos , Estrutura Secundária de Proteína/efeitos dos fármacos , Piridonas/química , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-AtividadeRESUMO
A three-component reaction forming dihydro 2,7-naphthyridine-1-ones has been developed. These unstable dihydro intermediates can be either oxidized or reduced to form naphthyridones or tetrahydro naphthyridones, respectively. The reaction tolerates a large variety of aldehydes and amines, and the produced compounds are analogs of the natural product lophocladine A.
Assuntos
Naftiridinas/síntese química , Aldeídos/química , Aminas/química , Estrutura Molecular , Naftiridinas/química , OxirreduçãoRESUMO
A novel heterocyclic scaffold with a peptidomimetic backbone structure has been synthesized. The scaffold is formed by insertion of primary amines into a cyclic sulfone to give the corresponding ring-expanded sulfonamides. By varying the amine component, a series of potentially biologically interesting compounds has been synthesized.
Assuntos
Sulfonamidas/síntese química , Sulfonas/química , Sulfonas/síntese química , Aminas/química , Catálise , Estrutura Molecular , Sulfonamidas/químicaRESUMO
An efficient method to synthesize a novel rigid tricyclic peptidomimetic scaffold through ring-closure of amino-functionalized bicyclic 2-pyridones has been discovered. The scaffold can function as a peptide backbone mimetic (highlighted) with two substituents independently variable to fine-tune biological response. Halogenation of the pyrazolo ring followed by Suzuki couplings made it possible to introduce substituents with variable electronic properties late in the synthetic route, which is preferable in library synthesis.
Assuntos
Peptídeos Cíclicos/síntese química , Conformação Molecular , Peptídeos Cíclicos/química , Pirazóis/química , Piridonas/química , Tiazóis/químicaRESUMO
Substituted bicyclic 2-pyridones, termed pilicides, are dipeptide mimetics that prevent pilus assembly in uropathogenic Escherichia coli. Here, we apply rational design to produce four classes of extended peptidomimetics based on two bioactive 2-pyridones. The key intermediate in the synthesis was an amino-functionalised 2-pyridone scaffold, which could be obtained via a mild and selective nitration and subsequent reduction. Procedures were then developed to further derivatize this amino-substituted core and a total of 24 extended peptidomimetics were synthesised and evaluated for chaperone affinity and in vivo antivirulence activity in P pili producing E. coli. Enhanced affinities for the target protein were observed within the generated set of compounds, while the ability to prevent pilus assembly in vivo was significantly decreased compared to the parent lead compounds. The results suggest that the limited in vivo potencies of the analogues are either uptake/distribution related or due to loss in binding specificity.