RESUMO
Sirtuins (SIRT1-SIRT7) are an evolutionary conserved family of NAD+-dependent protein deacylases regulating the acylation state of ε-N-lysine residues of proteins thereby controlling key biological processes. Numerous studies have found association of the aberrant enzymatic activity of SIRTs with various diseases like diabetes, cancer and neurodegenerative disorders. Previously, we have shown that substituted 2-alkyl-chroman-4-one/chromone derivatives can serve as selective inhibitors of SIRT2 possessing an antiproliferative effect in two human cancer cell lines. In this study, we have explored the bioisosteric replacement of the chroman-4-one/chromone core structure with different less lipophilic bicyclic scaffolds to overcome problems associated to poor physiochemical properties due to a highly lipophilic substitution pattern required for achieve a good inhibitory effect. Various new derivatives based on the quinolin-4(1H)-one scaffold, bicyclic secondary sulfonamides or saccharins were synthesized and evaluated for their SIRT inhibitory effect. Among the evaluated scaffolds, the benzothiadiazine-1,1-dioxide-based compounds showed the highest SIRT2 inhibitory activity. Molecular modeling studies gave insight into the binding mode of the new scaffold-replacement analogues.
Assuntos
Cromonas/farmacologia , Inibidores Enzimáticos/farmacologia , Sirtuína 2/antagonistas & inibidores , Cromonas/síntese química , Cromonas/química , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Modelos Moleculares , Estrutura Molecular , Sirtuína 2/metabolismo , Relação Estrutura-AtividadeRESUMO
Sirtuinâ 6 (SIRT6) is an NAD+-dependent histone deacetylase enzyme that is involved in multiple molecular pathways related to aging. Initially, it was reported that SIRT6 selectively deacetylated H3K9Ac and H3K56Ac; however, it has more recently been shown to preferentially hydrolyze long-chain fatty acyl groups over acetyl groups in vitro. Subsequently, fatty acids were demonstrated to increase the catalytic activity of SIRT6. In this study, we investigated whether a series of N-acylethanolamines (NAEs), quercetin, and luteolin could regulate SIRT6 activity. NAEs increased SIRT6 activity, with oleoylethanolamide having the strongest activity (EC50 value of 3.1 µm). Quercetin and luteolin were demonstrated to have dual functionality with respect to SIRT6 activity; namely, they inhibited SIRT6 activity with IC50 values of 24 and 2 µm, respectively, and stimulated SIRT6 activity more than sixfold (EC50 values of 990 and 270 µm, respectively).
Assuntos
Etanolaminas/química , Sirtuínas/química , Etanolaminas/metabolismo , Humanos , Estrutura Molecular , Sirtuínas/isolamento & purificação , Sirtuínas/metabolismoRESUMO
Control of histone acetylation is a part of the epigenetic mechanism that regulates gene expression and chromatin architecture. The members of the bromodomain and extra terminal domain (BET) protein family are a group of epigenetic readers that recognize histone acetylation, whereas histone deacetyl- ases such as sirtuinâ 1 (SIRT1) function as epigenetic erasers. We observed that BET inhibition by the specific inhibitor JQ1 upregulated SIRT1 expression and activated SIRT1. Moreover, we observed that BET inhibition functionally reversed the pro-inflammatory effect of SIRT1 inhibition in a cellular lung disease model. SIRT1 activation is desirable in many age-related, metabolic and inflammatory diseases; our results suggest that BET protein inhibition would be beneficial in treatment of those conditions. Most importantly, our findings demonstrate a novel mechanism of SIRT1 activation by inhibition of the BET proteins.
Assuntos
Azepinas/farmacologia , Inflamação/tratamento farmacológico , Proteínas Nucleares/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Sirtuína 1/genética , Fatores de Transcrição/antagonistas & inibidores , Triazóis/farmacologia , Regulação para Cima/efeitos dos fármacos , Animais , Proteínas de Ciclo Celular , Linhagem Celular , Linhagem Celular Tumoral , Epigênese Genética , Células HEK293 , Humanos , Inflamação/genética , Inflamação/imunologia , Células MCF-7 , Camundongos , Proteínas Nucleares/genética , Proteínas Nucleares/imunologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Interferência de RNA , RNA Interferente Pequeno/genética , Sirtuína 1/imunologia , Fatores de Transcrição/genética , Fatores de Transcrição/imunologiaRESUMO
SIRT3 is a member of the sirtuin family of histone deacetylases. It is a mitochondrial protein, which has an important role in metabolic homeostasis but it may also act as a tumor suppressor or promoter. Increased SIRT3 transcription has been associated with node-positive breast cancer and oral squamous cell carcinoma. To identify novel SIRT3 inhibitors we have established a virtual screening workflow by using shape-based filtering and flexible docking protocol. The Chembridge database was screened and 40 molecules were selected and tested in an in vitro assay. Two novel scaffolds were identified among the tested hits. The 5-amino-2-phenyl-benzoxazole scaffold was selected for further structure-activity studies and a series of its analogs was tested. The SIRT3 inhibition for this series ranged between 13% and 71%.
Assuntos
Benzoxazóis/farmacologia , Desenho de Fármacos , Inibidores Enzimáticos/farmacologia , Sirtuína 3/antagonistas & inibidores , Animais , Benzoxazóis/química , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Humanos , Camundongos , Modelos Moleculares , Estrutura Molecular , Sirtuína 3/metabolismo , Relação Estrutura-AtividadeRESUMO
This paper reports the synthesis of a series of evodiamine derivatives. We assayed the ability to inhibit cell growth on three human tumour cell lines (H460, MCF-7 and HepG2) and we evaluated the capacity to interfere with the catalytic activity of topoisomerase I both by the relaxation assay and the occurrence of the cleavable complex. Moreover, whose effect on sirtuins 1, 2 and 3 was investigated. Finally, molecular docking analyses were performed in an attempt to rationalize the biological results.
Assuntos
DNA Topoisomerases Tipo I/química , Quinazolinas/química , Sirtuínas/antagonistas & inibidores , Inibidores da Topoisomerase I/química , Alcaloides/química , Sítios de Ligação , Carbolinas/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , DNA Topoisomerases Tipo I/metabolismo , Ativação Enzimática/efeitos dos fármacos , Células Hep G2 , Humanos , Células MCF-7 , Simulação de Acoplamento Molecular , Estrutura Terciária de Proteína , Quinazolinas/síntese química , Quinazolinas/farmacologia , Sirtuínas/metabolismo , Inibidores da Topoisomerase I/síntese química , Inibidores da Topoisomerase I/farmacologiaRESUMO
Sirtuins catalyze the NAD(+) dependent deacetylation of N(epsilon)-acetyl lysine residues to nicotinamide, O'-acetyl-ADP-ribose (OAADPR) and N(epsilon)-deacetylated lysine. Here, an easy-to-synthesize Ac-Ala-Lys-Ala sequence has been used as a probe for the screening of novel N(epsilon)-modified lysine containing inhibitors against SIRT1 and SIRT2. N(epsilon)-Selenoacetyl and N(epsilon)-isothiovaleryl were the most potent moieties found in this study, comparable to the widely studied N(epsilon)-thioacetyl group. The N(epsilon)-3,3-dimethylacryl and N(epsilon)-isovaleryl moieties gave significant inhibition in comparison to the N(epsilon)-acetyl group present in the substrates. In addition, the studied N(epsilon)-alkanoyl, N(epsilon)-alpha,beta-unsaturated carbonyl and N(epsilon)-aroyl moieties showed that the acetyl binding pocket can accept rather large groups, but is sensitive to even small changes in electronic and steric properties of the N(epsilon)-modification. These results are applicable for further screening of N(epsilon)-acetyl analogues.
Assuntos
Inibidores de Histona Desacetilases/química , Lisina/análogos & derivados , Peptídeos/química , Sirtuína 1/antagonistas & inibidores , Sirtuína 2/antagonistas & inibidores , Sequência de Aminoácidos , Inibidores de Histona Desacetilases/síntese química , Inibidores de Histona Desacetilases/farmacologia , Humanos , Lisina/síntese química , Lisina/farmacologia , Peptídeos/síntese química , Peptídeos/farmacologia , Sirtuína 1/metabolismo , Sirtuína 2/metabolismo , Relação Estrutura-AtividadeRESUMO
Flavonoids are polyphenolic secondary metabolites synthesized by plants and fungus with various pharmacological effects. Due to their plethora of biological activities, they have been studied extensively in drug development. They have been shown to modulate the activity of a NAD+-dependent histone deacetylase, SIRT6. Because SIRT6 has been implicated in longevity, metabolism, DNA-repair, and inflammatory response reduction, it is an interesting target in inflammatory and metabolic diseases as well as in cancer. Here we show, that flavonoids can alter SIRT6 activity in a structure dependent manner. Catechin derivatives with galloyl moiety displayed significant inhibition potency against SIRT6 at 10 µM concentration. The most potent SIRT6 activator, cyanidin, belonged to anthocyanidins, and produced a 55-fold increase in SIRT6 activity compared to the 3-10 fold increase for the others. Cyanidin also significantly increased SIRT6 expression in Caco-2 cells. Results from the docking studies indicated possible binding sites for the inhibitors and activators. Inhibitors likely bind in a manner that could disturb NAD+ binding. The putative activator binding site was found next to a loop near the acetylated peptide substrate binding site. In some cases, the activators changed the conformation of this loop suggesting that it may play a role in SIRT6 activation.
Assuntos
Antocianinas , Simulação de Acoplamento Molecular , Sirtuínas , Antocianinas/química , Antocianinas/farmacologia , Sítios de Ligação , Células CACO-2 , Inibidores de Histona Desacetilases/química , Inibidores de Histona Desacetilases/farmacologia , Humanos , Polifenóis , Estrutura Secundária de Proteína , Sirtuínas/antagonistas & inibidores , Sirtuínas/química , Sirtuínas/metabolismoRESUMO
The effects of a novel prolyl oligopeptidase (POP) inhibitor KYP-2047 on spatial memory of young (3-month-old) and old (8- to 9-month-old) scopolamine-treated rats (0.4 mg/kg intraperitoneally) was investigated in the Morris water maze. In addition, the concentrations of promnesic neuropeptide substrates of POP, substance P and neurotensin in various brain areas after acute and chronic POP inhibition were measured in young rats. In addition, inositol-1,4,5-trisphosphate (IP(3)) levels were assayed in rat cortex and hippocampus after effective 2.5-day POP inhibition. KYP-2047 (1 or 5 mg/kg 30 min. before daily testing) dose-dependently improved the escape performance (i.e. latency to find the hidden platform and swimming path length) of the young but not the old rats in the water maze. POP inhibition had no consistent effect on substance P levels in cortex, hippocampus or hypothalamus, and only a modest increase in neurotensin concentration was observed in the hypothalamus after a single dose of KYP-2047. Moreover, IP(3) concentrations remained unaffected in cortex and hippocampus after POP inhibition. In conclusion, the behavioural data support the earlier findings of the promnesic action of POP inhibitors, but the mechanism of the memory-enhancing action remains unclear.
Assuntos
Memória/efeitos dos fármacos , Serina Endopeptidases/metabolismo , Inibidores de Serina Proteinase/farmacologia , Envelhecimento/fisiologia , Amnésia/induzido quimicamente , Amnésia/tratamento farmacológico , Amnésia/fisiopatologia , Animais , Lobo Frontal/efeitos dos fármacos , Lobo Frontal/metabolismo , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Hipotálamo/efeitos dos fármacos , Hipotálamo/metabolismo , Inositol 1,4,5-Trifosfato/metabolismo , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Memória/fisiologia , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Antagonistas Muscarínicos , Neurotensina/metabolismo , Prolil Oligopeptidases , Pirrolidinas/farmacologia , Ratos , Ratos Wistar , Escopolamina , Substância P/metabolismoRESUMO
Prolyl oligopeptidase (POP) is a serine protease that specifically hydrolyses small peptides at the carboxyl end of the proline residue. POP has gained pharmaceutical interest, since its inhibitors have been shown to have antiamnesic properties in rat. We examined the effect of the 2(S)-substituents CN and COCH(2)OH at the P1 site of the parent inhibitors isophthalic acid 2(S)-(cyclopentanecarbonyl)pyrrolidine-l-prolyl-pyrrolidine amide and 4-phenylbutanoyl-l-prolyl-pyrrolidine and bulky 5-t-butyl group at the P2 site l-prolyl residue of the parent inhibitor 4-phenylbutanoyl-l-prolyl-pyrrolidine on the binding kinetics to the enzyme. In addition, we studied the duration of POP inhibition in the rat tissues in vivo after i.p. administration. CN and COCH(2)OH substituents at the P1 site pyrrolidine group were found to greatly increase the affinity of the inhibitor and the enzyme-inhibitor complex half-life. In addition, 5-t-butyl group at the P2 site l-prolyl residue increased the dissociation half-life of the enzyme-inhibitor complex, without much affecting the inhibitory potency. The duration of the inhibition in the rat tissues followed the inhibition kinetic properties in that the compounds with fast dissociation produced shorter inhibition in the rat tissues than the compounds with slow dissociation. The duration of POP inhibition of compounds was evidently not governed by their serum clearance. The fact that the in vivo pharmacodynamic behaviour of POP inhibitors can be predicted by their in vitro-properties may be of importance when designing therapeutically useful POP inhibitors.
Assuntos
Inibidores de Proteases/farmacocinética , Serina Endopeptidases/efeitos dos fármacos , Animais , Sequência de Bases , Western Blotting , Primers do DNA , Meia-Vida , Humanos , Cinética , Masculino , Prolil Oligopeptidases , Inibidores de Proteases/sangue , Ratos , Ratos Wistar , SuínosRESUMO
Photoaffinity labeling (PAL) was used to identify the binding site of chroman-4-one-based SIRT2-selective inhibitors. The photoactive diazirine 4, a potent SIRT2 inhibitor, was subjected to detailed photochemical characterization. In PAL experiments with SIRT2, a tryptic peptide originating from the covalent attachment of photoactivated 4 was identified. The peptide covers both the active site of SIRT2 and the proposed binding site of chroman-4-one-based inhibitors. A high-power LED was used as source for the monochromatic UV light enabling rapid photoactivation.
Assuntos
Cromonas/farmacologia , Inibidores Enzimáticos/farmacologia , Marcadores de Fotoafinidade/química , Sirtuína 2/antagonistas & inibidores , Sítios de Ligação/efeitos dos fármacos , Cromonas/síntese química , Cromonas/química , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Modelos Moleculares , Estrutura Molecular , Sirtuína 2/metabolismo , Relação Estrutura-Atividade , Espectrometria de Massas em TandemRESUMO
Sirtuins are NAD(+)-dependent histone deacetylases regulating important metabolic pathways in prokaryotes and eukaryotes and are involved in many biological processes such as cell survival, senescence, proliferation, apoptosis, DNA repair, cell metabolism, and caloric restriction. The seven members of this family of enzymes are considered potential targets for the treatment of human pathologies including neurodegenerative diseases, cardiovascular diseases, and cancer. Furthermore, recent interest focusing on sirtuin modulators as epigenetic players in the regulation of fundamental biological pathways has prompted increased efforts to discover new small molecules able to modify sirtuin activity. Here, we review the role, mechanism of action, and biological function of the seven sirtuins, as well as their inhibitors and activators.
Assuntos
Inibidores Enzimáticos/farmacologia , Sirtuínas/metabolismo , Proliferação de Células , Sobrevivência Celular , Senescência Celular , Reparo do DNA , Epigênese Genética , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Sirtuínas/antagonistas & inibidoresRESUMO
A series of dicarboxylic acid azacycle l-prolyl-pyrrolidine amides was synthesized, and their inhibitory activity against prolyl oligopeptidase (POP) from porcine brain was tested. Three different azacycles were tested at the position beyond P3 and six different dicarboxylic acids at the P3 position. l-Prolyl-pyrrolidine and l-prolyl-2(S)-cyanopyrrolidine were used at the P2-P1 positions. The IC(50) values ranged from 0.39 to 19000 nM. The most potent inhibitor was the 3,3-dimethylglutaric acid azepane l-prolyl-2(S)-cyanopyrrolidine amide. Molecular docking (GOLD) was used to analyze binding interactions between different POP inhibitors of this type and the POP enzyme. The data set consisted of the novel inhibitors, inhibitors published previously by our group, and well-known reference compounds. The alignments were further analyzed using comparative molecular similarity indices analysis. The binding of the inhibitors was consistent at the P1-P3 positions. Beyond the P3 position, two different binding modes were found, one that favors lipophilic structures and one that favors nonhydrophobic structures.
Assuntos
Compostos Aza/síntese química , Prolina/análogos & derivados , Prolina/síntese química , Pirrolidinas/síntese química , Serina Endopeptidases/química , Inibidores de Serina Proteinase/síntese química , Amidas/síntese química , Amidas/farmacologia , Animais , Compostos Aza/farmacologia , Encéfalo/efeitos dos fármacos , Encéfalo/enzimologia , Ácidos Dicarboxílicos/síntese química , Ácidos Dicarboxílicos/farmacologia , Técnicas In Vitro , Modelos Moleculares , Prolina/farmacologia , Prolil Oligopeptidases , Pirrolidinas/farmacologia , Relação Quantitativa Estrutura-Atividade , Serina Endopeptidases/metabolismo , Inibidores de Serina Proteinase/farmacologia , SuínosRESUMO
POP (prolyl oligopeptidase) specifically hydrolyses a number of small proline-containing peptides at the carboxy end of the proline residue and POP inhibitors have been shown to have cognition-enhancing properties. It has been noted that certain functional groups at the P1 site of the inhibitor, which correspond to the substrate residue on the N-terminal side of the bond to be cleaved, increase the inhibitory potency. However, detailed mechanistic and kinetic analysis of the inhibition has not been studied. In the present study, we examined the effect of different functional groups at the P1 site of the parent inhibitor isophthalic acid bis-(L-prolylpyrrolidine) amide on the binding kinetics to POP. Addition of CHO, CN or COCH(2)OH groups to the P1 site increased the inhibitory potency by two orders of magnitude (K(i)=11.8-0.1 nM) and caused a clear slow-binding inhibition. The inhibitor containing a CHO group had the lowest association rate constant, k(on)=(2.43+/-0.12) x 10(5) M(-1) x s(-1), whereas the inhibitor with a CN group exhibited the fastest binding, k(on)=(12.0+/-0.08)x10(5) M(-1) x s(-1). In addition, the dissociation rate was found to be crucially dependent on the type of the functional group. Compounds with COCH(2)OH and CHO groups had much longer half-lives of dissociation (over 5 h) compared with the compound with the CN group (25 min), although the K(i) values of the compounds were relatively similar. A possibility to optimize the duration of inhibition by changing the functional group at the P1 site is important when planning therapeutically useful POP inhibitors.
Assuntos
Ácidos Ftálicos/farmacologia , Pirrolidinas/farmacologia , Serina Endopeptidases/metabolismo , Inibidores de Serina Proteinase/química , Animais , Sítios de Ligação , Cinética , Modelos Químicos , Estrutura Molecular , Ácidos Ftálicos/química , Ácidos Ftálicos/metabolismo , Prolil Oligopeptidases , Ligação Proteica , Pirrolidinas/química , Pirrolidinas/metabolismo , Inibidores de Serina Proteinase/metabolismo , Relação Estrutura-Atividade , SuínosRESUMO
Sirtuins (SIRT1-SIRT7) are NAD dependent deacetylases and intriguing drug targets in for example neurodegenerative diseases and cancer. Virtual screening has been shown to be a fast and efficient method of discovering new sirtuin inhibitors. In this study, a new putative binding site on the zinc binding domain of sirtuins was utilized to screen the ZINC database virtually in order to discover new sirtuin inhibiting scaffolds. Altogether 26 compounds were tested in vitro and initially 15 inhibitors displayed >30% SIRT3 inhibition. However, the evaluation of raw data from in vitro assay revealed that many of the compounds had intrinsic property to interfere with the fluorescence signal at the assay wavelengths resulting in false positives. All compounds with over 30% SIRT3 inhibition were studied more closely for their behavior in the assay and eventually, three compounds were identified as novel sirtuin inhibitors. They displayed 32-40% SIRT3 and 21-60% SIRT2 inhibition. The inhibitors display two new scaffolds, the smaller of which can be considered as a promising fragment, which offers a base for structural optimization.
Assuntos
Desenho Assistido por Computador , Desenho de Fármacos , Inibidores de Histona Desacetilases/farmacologia , Simulação de Acoplamento Molecular , Sirtuína 2/antagonistas & inibidores , Sirtuína 3/antagonistas & inibidores , Regulação Alostérica , Artefatos , Sítios de Ligação , Domínio Catalítico , Inibidores de Histona Desacetilases/química , Inibidores de Histona Desacetilases/metabolismo , Ligação Proteica , Conformação Proteica , Sirtuína 2/química , Sirtuína 2/metabolismo , Sirtuína 3/química , Sirtuína 3/metabolismo , Espectrometria de Fluorescência , Relação Estrutura-AtividadeRESUMO
Isophthalic acid bis(l-prolyl-pyrrolidine) amide is a very potent prolyl oligopeptidase inhibitor, but it has a log P value of -0.2, which is very low for a compound targeted to the brain. Therefore, these types of compounds were further modified to improve the structure-activity relationships, with the focus on increasing the log P value. The inhibitory activity against prolyl oligopeptidase from pig brain was tested in vitro. The most promising compounds resulted from replacing the pyrrolidinyl group at the P5 site by cycloalkyl groups, such as cyclopentyl and cyclohexyl groups, and by a phenyl group. These compounds are slightly more potent, and they have a significantly higher log P value. The potency of these compounds was further increased by replacing the pyrrolidinyl group at the P1 site by 2(S)-cyanopyrrolidinyl and 2(S)-(hydroxyacetyl)pyrrolidinyl groups.
Assuntos
Prolina/análogos & derivados , Inibidores de Proteases/síntese química , Inibidores de Proteases/farmacologia , Pirrolidinas/síntese química , Pirrolidinas/farmacologia , Serina Endopeptidases/efeitos dos fármacos , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/enzimologia , Indicadores e Reagentes , Espectroscopia de Ressonância Magnética , Prolina/síntese química , Prolina/farmacologia , Prolil Oligopeptidases , Pirróis/farmacologia , Relação Estrutura-Atividade , SuínosRESUMO
With the aim to replace the natural amino acid proline by a proline mimetic structure, a cyclopent-2-enecarbonyl moiety was studied at the P2 position of prolyl oligopeptidase (POP) inhibitors. The cyclopent-2-enecarbonyl moiety proved to be an excellent proline mimetic at the P2 position of POP inhibitors. The replacement is particularly useful when increased lipophilicity is needed.
Assuntos
Ciclopentanos/química , Prolina/química , Serina Endopeptidases/metabolismo , Inibidores de Serina Proteinase/química , 1-Octanol , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/enzimologia , Soluções Tampão , Ciclopentanos/síntese química , Ciclopentanos/farmacologia , Técnicas In Vitro , Mimetismo Molecular , Prolil Oligopeptidases , Inibidores de Serina Proteinase/síntese química , Inibidores de Serina Proteinase/farmacologia , Solubilidade , Relação Estrutura-Atividade , SuínosRESUMO
The modulation of protein deacetylase SIRT1 has a vast therapeutic potential in treatment of several aging-associated diseases. Active regulator of SIRT1 (AROS) is a small endogenous protein which was originally reported to activate SIRT1 through a direct interaction in cancer cells. We show that the interaction between the two proteins is weak and does not alter the activity of SIRT1 in non-cancerous human cells. The results of different in vitro SIRT1 activity assays disclosed AROS as an inhibitor of SIRT1. The functional relationship between AROS and SIRT1 proved to be dependent on the biological context and experimental setting.
Assuntos
Proteínas Nucleares/metabolismo , Sirtuína 1/metabolismo , Fatores de Transcrição/metabolismo , Acetilação , Células Epiteliais/enzimologia , Células HEK293 , Histonas/química , Humanos , NAD/química , Proteínas Nucleares/química , Ligação Proteica , Processamento de Proteína Pós-Traducional , Epitélio Pigmentado da Retina/citologia , Sirtuína 1/química , Fatores de Transcrição/química , Proteína Supressora de Tumor p53/metabolismoRESUMO
SIRT6 is a modulator of chromatin structure having an important role in healthy ageing, and there is a crucial need to find specific modulators for it. Therefore, the activity of SIRT6 should be studied using a variety of methods. We examined the capability of SIRT6 to deacetylate a set of five fluorogenic substrates based on p53 and histone H3 sequences. The substrate designed around H3K56 deacetylation site exhibited the best signal-to-background ratio and was chosen for further studies. Nicotinamide is a known inhibitor for sirtuins, and it was found to be less potent inhibitor for SIRT6 than it is for SIRT1. In addition, we studied 15 other small molecule sirtuin modulators using the H3K56 based substrate. EX-527, quercetin and three pseudopeptidic compounds were found to be the most potent SIRT6 inhibitors, exhibiting over 50% deacetylation inhibition. These findings describe the first modulators of SIRT6 activity at the physiologically important H3K56 deacetylation site.
Assuntos
Histonas/metabolismo , Sirtuínas/metabolismo , Acetilação/efeitos dos fármacos , Carbazóis/química , Carbazóis/farmacologia , Histonas/química , Humanos , Estrutura Molecular , Niacinamida/química , Niacinamida/farmacologia , Peptídeos/química , Peptídeos/farmacologia , Quercetina/química , Quercetina/farmacologia , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/metabolismo , Sirtuínas/antagonistas & inibidores , Sirtuínas/biossíntese , Relação Estrutura-AtividadeRESUMO
Sirtuins (SIRTs) catalyze the NAD(+)-dependent deacetylation of N(ε)-acetyl lysines on various protein substrates. SIRTs are interesting drug targets as they are considered to be related to important pathologies such as inflammation and aging-associated diseases. We have previously shown that chroman-4-ones act as potent and selective inhibitors of SIRT2. Herein we report novel chroman-4-one and chromone-based SIRT2 inhibitors containing various heterofunctionalities to improve pharmacokinetic properties. The compounds retained both high SIRT2 selectivity and potent inhibitory activity. Two compounds were tested for their antiproliferative effects in breast cancer (MCF-7) and lung carcinoma (A549) cell lines. Both compounds showed antiproliferative effects correlating with their SIRT2 inhibition potency. They also increased the acetylation level of α-tubulin, indicating that SIRT2 is likely to be the target in cancer cells. A binding mode of the inhibitors that is consistent with the SAR data was proposed based on a homology model of SIRT2.
Assuntos
Antineoplásicos/farmacocinética , Cromanos/síntese química , Cromonas/síntese química , Inibidores Enzimáticos/síntese química , Sirtuína 2/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cromanos/farmacocinética , Cromanos/farmacologia , Cromonas/farmacocinética , Cromonas/farmacologia , Inibidores Enzimáticos/farmacologia , Humanos , Tubulina (Proteína)/metabolismoRESUMO
Sirtuin 1 (SIRT1) is the most studied human sirtuin and it catalyzes the deacetylation reaction of acetylated lysine residues of its target proteins, for example histones. It is a promising drug target in the treatment of age-related diseases, such as neurodegenerative diseases and cancer. In this study, a series of known substrate-based sirtuin inhibitors was analyzed with comparative molecular field analysis (CoMFA), which is a three-dimensional quantitative structure-activity relationships (3D-QSAR) technique. The CoMFA model was validated both internally and externally, producing the statistical values concordance correlation coefficient (CCC) of 0.88, the mean value r(2)m of 0.66 and Q(2)F3 of 0.89. Based on the CoMFA interaction contours, 13 new potential inhibitors with high predicted activity were designed, and the activities were verified by in vitro measurements. This work proposes an effective approach for the design and activity prediction of new potential substrate-based SIRT1 inhibitors.