RESUMO
A growing body of evidence suggests that low nephron numbers at birth can increase the risk of chronic kidney disease or hypertension later in life. Environmental stressors, such as maternal malnutrition, medication and smoking, can influence renal size at birth. Using metanephric organ cultures to model single-variable environmental conditions, models of maternal disease were evaluated for patterns of developmental impairment. While hyperthermia had limited effects on renal development, fetal iron deficiency was associated with severe impairment of renal growth and nephrogenesis with an all-proximal phenotype. Culturing kidney explants under high glucose conditions led to cellular and transcriptomic changes resembling human diabetic nephropathy. Short-term high glucose culture conditions were sufficient for long-term alterations in DNA methylation-associated epigenetic memory. Finally, the role of epigenetic modifiers in renal development was tested using a small compound library. Among the selected epigenetic inhibitors, various compounds elicited an effect on renal growth, such as HDAC (entinostat, TH39), histone demethylase (deferasirox, deferoxamine) and histone methyltransferase (cyproheptadine) inhibitors. Thus, metanephric organ cultures provide a valuable system for studying metabolic conditions and a tool for screening for epigenetic modifiers in renal development.
Assuntos
Nefropatias Diabéticas/genética , Meio Ambiente , Epigênese Genética , Glucose/toxicidade , Rim/metabolismo , Efeitos Tardios da Exposição Pré-Natal/genética , Animais , Metilação de DNA , Feminino , Humanos , Deficiências de Ferro , Rim/efeitos dos fármacos , Camundongos , Técnicas de Cultura de Órgãos/métodos , Gravidez , TranscriptomaRESUMO
We designed and synthesized carbamates of the clinically-approved HDAC (histone deacetylase) inhibitor vorinostat (suberoylanilide hydroxamic acid, SAHA) in order to validate our previously-proposed hypothesis that these carbamates might serve as prodrugs for hydroxamic acid containing HDAC inhibitors. Biochemical assays proved our new compounds to be potent inhibitors of histone deacetylases in vitro, and they also showed antiproliferative effects in leukemic cells. These results, as well as stability analysis led to the suggestion that the intact carbamates are inhibitors of histone deacetylases themselves, representing a new zinc-binding warhead in HDAC inhibitor design. This suggestion was further supported by the synthesis and evaluation of a carbamate derivative of the HDAC6-selective inhibitor bufexamac.
Assuntos
Carbamatos/síntese química , Inibidores de Histona Desacetilases/síntese química , Histona Desacetilases/metabolismo , Ácidos Hidroxâmicos/síntese química , Pró-Fármacos/síntese química , Acetilação/efeitos dos fármacos , Motivos de Aminoácidos , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Bufexamac/química , Bufexamac/farmacologia , Carbamatos/farmacologia , Epigênese Genética , Células HL-60 , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/genética , Histonas/genética , Histonas/metabolismo , Humanos , Ácidos Hidroxâmicos/farmacologia , Simulação de Acoplamento Molecular , Pró-Fármacos/farmacologia , Relação Estrutura-Atividade , Células U937 , Vorinostat , Zinco/químicaRESUMO
The treatment of schistosomiasis, a disease caused by blood flukes parasites of the Schistosoma genus, depends on the intensive use of a single drug, praziquantel, which increases the likelihood of the development of drug-resistant parasite strains and renders the search for new drugs a strategic priority. Currently, inhibitors of human epigenetic enzymes are actively investigated as novel anti-cancer drugs and have the potential to be used as new anti-parasitic agents. Here, we report that Schistosoma mansoni histone deacetylase 8 (smHDAC8), the most expressed class I HDAC isotype in this organism, is a functional acetyl-L-lysine deacetylase that plays an important role in parasite infectivity. The crystal structure of smHDAC8 shows that this enzyme adopts a canonical α/ß HDAC fold, with specific solvent exposed loops corresponding to insertions in the schistosome HDAC8 sequence. Importantly, structures of smHDAC8 in complex with generic HDAC inhibitors revealed specific structural changes in the smHDAC8 active site that cannot be accommodated by human HDACs. Using a structure-based approach, we identified several small-molecule inhibitors that build on these specificities. These molecules exhibit an inhibitory effect on smHDAC8 but show reduced affinity for human HDACs. Crucially, we show that a newly identified smHDAC8 inhibitor has the capacity to induce apoptosis and mortality in schistosomes. Taken together, our biological and structural findings define the framework for the rational design of small-molecule inhibitors specifically interfering with schistosome epigenetic mechanisms, and further support an anti-parasitic epigenome targeting strategy to treat neglected diseases caused by eukaryotic pathogens.
Assuntos
Epigênese Genética , Proteínas de Helminto/química , Histona Desacetilases/química , Dobramento de Proteína , Schistosoma mansoni/enzimologia , Animais , Proteínas de Helminto/genética , Proteínas de Helminto/metabolismo , Inibidores de Histona Desacetilases/química , Histona Desacetilases/genética , Histona Desacetilases/metabolismo , Humanos , Estrutura Secundária de Proteína , Schistosoma mansoni/genéticaRESUMO
Schistosomiasis, caused by S. mansoni, is a tropical disease that affects over 200 million people worldwide. A novel approach for targeting eukaryotic parasites is to tackle their dynamic epigenetic machinery that is necessary for the extensive phenotypic changes during their life cycle. We recently identified S. mansoni histone deacetylase 8 (smHDAC8) as a potential target for antiparasitic therapy. Here we present results from a virtual screening campaign on smHDAC8. Besides hydroxamates, several sulfonamide-thiazole derivatives were identified by a target-based virtual screening using a homology model of smHDAC8. In vitro testing of 75 compounds identified 8 hydroxamates as potent and lead-like inhibitors of the parasitic HDAC8. Solving of the crystal structure of smHDAC8 with two of the virtual screening hits confirmed the predicted binding mode.
Assuntos
Proteínas de Helminto/antagonistas & inibidores , Inibidores de Histona Desacetilases/química , Histona Desacetilases/química , Ácidos Hidroxâmicos/química , Schistosoma mansoni/química , Sulfonamidas/química , Tiazóis/química , Animais , Sítios de Ligação , Cristalografia por Raios X , Descoberta de Drogas , Proteínas de Helminto/química , Ensaios de Triagem em Larga Escala , Ligantes , Simulação de Acoplamento Molecular , Ligação Proteica , Schistosoma mansoni/enzimologia , Homologia Estrutural de Proteína , Relação Estrutura-Atividade , Interface Usuário-ComputadorRESUMO
A series of hydroxamates (4a-4l) were prepared from p-aminobenzoic acid to inhibit HDAC8. The idea is to substitute rigid aromatic ring in place of less rigid piperazine ring of hydroxamates reported earlier by our group. It is expected to increase potency retaining the selectivity. Result obtained suggested that the modifications carried out retained the selectivity towards HDAC8 isoform and increasing the potency in very few cases. Increase in potency is also associated with variation in cap aryl region. Two compounds (4f &4l) were found to inhibit HDAC8 at concentrations (IC50) less than 20µM.
Assuntos
Ácido 4-Aminobenzoico/química , Ácido 4-Aminobenzoico/farmacologia , Inibidores de Histona Desacetilases/química , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/química , Ácidos Hidroxâmicos/farmacologia , Proteínas Repressoras/antagonistas & inibidores , Cristalografia por Raios X , Desenho de Fármacos , Histona Desacetilases/química , Histona Desacetilases/metabolismo , Humanos , Simulação de Acoplamento Molecular , Ligação Proteica , Proteínas Repressoras/química , Proteínas Repressoras/metabolismo , Relação Estrutura-AtividadeRESUMO
Anti-parasitic treatment of neglected tropical diseases caused by cestodes such as echinococcosis and cysticercosis relies on a small number of approved anthelmintic drugs. Furthermore, the treatment is usually prolonged and often partially effective and not well tolerated by some patients. Therefore, the identification of novel drug targets and their associated compounds is critical. In this study, we identified and characterised sirtuin enzymes in cestodes and evaluated the cestocidal potential of sirtuin inhibitors as new cestocidal molecules. Sirtuins are a highly conserved family of nicotinamide-adenine dinucleotide-lysine deacylases involved in multiple cellular functions. Here, we described the full repertoire of sirtuin-encoding genes in several cestode species. We identified six sirtuin-encoding genes that were classified into sirtuins Class I (SIRT1, SIRT2, and SIRT3), Class III (SIRT5), and Class IV (SIRT6 and SIRT7). In Echinococcus spp., sirtuin genes showed transcriptional expression throughout several developmental stages, sirtuin 2 (SIRT2) being the most expressed. To evaluate the potential of sirtuin inhibitors as new cestocidal molecules, we determined the in vitro effect of several Class I sirtuin inhibitors by motility assay. Of those, the selective SIRT2 inhibitor Mz25 showed a strong cestocidal activity in Mesocestoides vogae (syn. Mesocestoides corti) tetrathyridia at various concentrations. The Mz25 cestocidal activity was time- and dose-dependent with a half-maximal inhibitory concentration value significantly lower than that of albendazole. Additionally, Mz25 induced extensive damage in the general morphology with marked alterations in the tegument and ultrastructural features. By homology modelling, we found that cestode SIRT2s showed a high conservation of the canonical sirtuin structure as well as in the residues related to Mz25 binding. Interestingly, some non-conservative mutations were found on the selectivity pocket (an Mz25-induced structural rearrangement on the active site), which represent a promising lead for developing selective cestode SIRT2 inhibitors derived from Mz25. Nevertheless, the Mz25 molecular target in M. vogae is unknown and remains to be determined. This report provides the basis for further studies of sirtuins to understand their roles in cestode biology and to develop selective sirtuin inhibitors to treat these neglected tropical diseases.
Assuntos
Cestoides , Mesocestoides , Sirtuínas , Albendazol/farmacologia , Animais , Cestoides/genética , Mesocestoides/metabolismo , Sirtuínas/genética , Sirtuínas/metabolismoRESUMO
In utero renal development is subject to maternal metabolic and environmental influences affecting long-term renal function and the risk of developing chronic kidney failure and cardiovascular disease. Epigenetic processes have been implicated in the orchestration of renal development and prenatal programming of nephron number. However, the role of many epigenetic modifiers for kidney development is still unclear. Bromodomain and extra-terminal domain (BET) proteins act as histone acetylation reader molecules and promote gene transcription. BET family members Brd2, Brd3 and Brd4 are expressed in the nephrogenic zone during kidney development. Here, the effect of the BET inhibitor JQ1 on renal development is evaluated. Inhibition of BET proteins via JQ1 leads to reduced growth of metanephric kidney cultures, loss of the nephron progenitor cell population, and premature and disturbed nephron differentiation. Gene expression of key nephron progenitor transcription factor Osr1 is downregulated after 24 h BET inhibition, while Lhx1 and Pax8 expression is increased. Mining of BRD4 ChIP-seq and gene expression data identify Osr1 as a key factor regulated by BRD4-controlled gene activation. Inhibition of BRD4 by BET inhibitor JQ1 leads to downregulation of Osr1, thereby causing a disturbance in the balance of nephron progenitor cell self-renewal and premature differentiation of the nephron, which ultimately leads to kidney hypoplasia and disturbed nephron development. This raises questions about the potential teratogenic effects of BET inhibitors for embryonic development. In summary, our work highlights the role of BET proteins for prenatal programming of nephrogenesis and identifies Osr1 as a potential target of BET proteins.
RESUMO
BACKGROUND: Echinococcosis and cysticercosis are neglected tropical diseases caused by cestode parasites (family Taeniidae). Not only there is a small number of approved anthelmintics for the treatment of these cestodiases, but also some of them are not highly effective against larval stages, such that identifying novel drug targets and their associated compounds is critical. Histone deacetylase (HDAC) enzymes are validated drug targets in cancers and other diseases, and have been gaining relevance for developing new potential anti-parasitic treatments in the last years. Here, we present the anthelmintic profile for a panel of recently developed HDAC inhibitors against the model cestode Mesocestoides vogae (syn. M. corti). METHODOLOGY/PRINCIPAL FINDINGS: Phenotypic screening was performed on M. vogae by motility measurements and optical microscopic observations. Some HDAC inhibitors showed potent anthelmintic activities; three of them -entinostat, TH65, and TH92- had pronounced anthelmintic effects, reducing parasite viability by ~100% at concentrations of ≤ 20 µM. These compounds were selected for further characterization and showed anthelmintic effects in the micromolar range and in a time- and dose-dependent manner. Moreover, these compounds induced major alterations on the morphology and ultrastructural features of M. vogae. The potencies of these compounds were higher than albendazole and the anthelmintic effects were irreversible. Additionally, we evaluated pairwise drug combinations of these HDAC inhibitors and albendazole. The results suggested a positive interaction in the anthelmintic effect for individual pairs of compounds. Due to the maximum dose approved for entinostat, adjustments in the dose regime and/or combinations with currently-used anthelmintic drugs are needed, and the selectivity of TH65 and TH92 towards parasite targets should be assessed. CONCLUSION, SIGNIFICANCE: The results presented here suggest that HDAC inhibitors represent novel and potent drug candidates against cestodes and pave the way to understanding the roles of HDACs in these parasites.
Assuntos
Anti-Helmínticos/farmacologia , Benzamidas/farmacologia , Inibidores de Histona Desacetilases/farmacologia , Mesocestoides/efeitos dos fármacos , Piridinas/farmacologia , Albendazol/farmacologia , Animais , Infecções por Cestoides , Larva/anatomia & histologia , Larva/efeitos dos fármacos , Mesocestoides/anatomia & histologiaRESUMO
The cell type diversity and complexity of the nervous system is generated by a network of signaling events, transcription factors, and epigenetic regulators. Signaling and transcriptional control have been easily amenable to forward genetic screens in model organisms like zebrafish. In contrast, epigenetic mechanisms have been somewhat elusive in genetic screens, likely caused by broad action in multiple developmental pathways that masks specific phenotypes, but also by genetic redundancies of epigenetic factors. Here, we performed a screen using small molecule inhibitors of epigenetic mechanisms to reveal contributions to specific aspects of neurogenesis in zebrafish. We chose development of dopaminergic and noradrenergic neurons from neural progenitors as target of epigenetic regulation. We performed the screen in two phases: First, we tested a small molecule inhibitor library that targets a broad range of epigenetic protein classes and mechanisms, using expression of the dopaminergic and noradrenergic marker tyrosine hydroxylase as readout. We identified 10 compounds, including HDAC, Bromodomain and HAT inhibitors, which interfered with dopaminergic and noradrenergic development in larval zebrafish. In the second screening phase, we aimed to identify neurogenesis stages affected by these 10 inhibitors. We analyzed treated embryos for effects on neural stem cells, growth progression of the retina, and apoptosis in neural tissues. In addition, we analyzed effects on islet1 expressing neuronal populations to determine potential selectivity of compounds for transmitter phenotypes. In summary, our targeted screen of epigenetic inhibitors identified specific compounds, which reveal chromatin regulator classes that contribute to dopaminergic and noradrenergic neurogenesis in vivo.
Assuntos
Inibidores Enzimáticos/farmacologia , Epigênese Genética/fisiologia , Quinazolinas/farmacologia , Autoantígenos , Azepinas/farmacologia , Sistemas de Liberação de Medicamentos , Inibidores Enzimáticos/química , Proteínas da Matriz do Complexo de Golgi , Antígenos de Histocompatibilidade , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Humanos , Estrutura Molecular , Proteínas/antagonistas & inibidores , Quinazolinas/química , Transdução de Sinais/efeitos dos fármacos , Relação Estrutura-AtividadeRESUMO
DNA and histone methylation belong to the key regulatory components in the epigenetic machinery, and dysregulations of these processes have been associated with various human diseases. Small molecule modulators of these epigenetic targets are highly valuable both as chemical probes to study the biological roles of the target proteins, and as potential therapeutics. Indeed, recent years have seen the discovery of chemical modulators of several epigenetic targets, some of which are already marketed drugs or undergoing clinical trials. In this review, we will focus on small molecule modulators of DNA and histone methylation.
Assuntos
Metilação de DNA/efeitos dos fármacos , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Epigênese Genética/efeitos dos fármacos , Histonas/genética , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Antineoplásicos/farmacologia , Metilases de Modificação do DNA/antagonistas & inibidores , Metilases de Modificação do DNA/metabolismo , Descoberta de Drogas/métodos , Inibidores Enzimáticos/química , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/metabolismo , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Proteína-Arginina N-Metiltransferases/antagonistas & inibidores , Proteína-Arginina N-Metiltransferases/metabolismo , Bibliotecas de Moléculas Pequenas/químicaRESUMO
Schistosomiasis is a major neglected parasitic disease that affects more than 265 million people worldwide and for which the control strategy consists of mass treatment with the only available drug, praziquantel. In this study, a series of new benzohydroxamates were prepared as potent inhibitors of Schistosoma mansoni histone deacetylase 8 (smHDAC8). Crystallographic analysis provided insights into the inhibition mode of smHDAC8 activity by these 3-amidobenzohydroxamates. The newly designed inhibitors were evaluated in screens for enzyme inhibitory activity against schistosome and human HDACs. Twenty-seven compounds were found to be active in the nanomolar range, and some of them showed selectivity toward smHDAC8 over the major human HDACs (1 and 6). The active benzohydroxamates were additionally screened for lethality against the schistosome larval stage using a fluorescence-based assay. Four of these showed significant dose-dependent killing of the schistosome larvae and markedly impaired egg laying of adult worm pairs maintained in culture.
Assuntos
Proteínas de Helminto/efeitos dos fármacos , Histona Desacetilases/efeitos dos fármacos , Schistosoma mansoni/efeitos dos fármacos , Esquistossomicidas/síntese química , Esquistossomicidas/farmacologia , Animais , Relação Dose-Resposta a Droga , Sistemas de Liberação de Medicamentos , Humanos , Larva , Modelos Moleculares , Schistosoma mansoni/genética , Esquistossomose mansoni/tratamento farmacológico , Relação Estrutura-Atividade , Especificidade por Substrato , Difração de Raios XRESUMO
Schistosomiasis, caused by the parasitic flatworm Schistosoma mansoni and related species, is a tropical disease that affects over 200 million people worldwide. A new approach for targeting eukaryotic parasites is to tackle their dynamic epigenetic machinery that is necessary for the extensive phenotypic changes during the life cycle of the parasite. Recently, we identified S. mansoni histone deacetylase 8 (smHDAC8) as a potential target for antiparasitic therapy. Here, we present results on the investigations of a focused set of HDAC (histone deacetylase) inhibitors on smHDAC8. Besides several active hydroxamates, we identified a thiol-based inhibitor that inhibited smHDAC8 activity in the micromolar range with unexpected selectivity over the human isotype, which has not been observed so far. The crystal structure of smHDAC8 complexed with the thiol derivative revealed that the inhibitor is accommodated in the catalytic pocket, where it interacts with both the catalytic zinc ion and the essential catalytic tyrosine (Y341) residue via its mercaptoacetamide warhead. To our knowledge, this is the first complex crystal structure of any HDAC inhibited by a mercaptoacetamide inhibitor, and therefore, this finding offers a rationale for further improvement. Finally, an ester prodrug of the thiol HDAC inhibitor exhibited antiparasitic activity on cultured schistosomes in a dose-dependent manner.
Assuntos
Antiparasitários/química , Proteínas de Helminto/química , Histona Desacetilases/química , Schistosoma mansoni/enzimologia , Tioacetamida/química , Animais , Antiparasitários/metabolismo , Antiparasitários/farmacologia , Apoptose/efeitos dos fármacos , Biocatálise/efeitos dos fármacos , Cristalografia por Raios X , Proteínas de Helminto/antagonistas & inibidores , Proteínas de Helminto/metabolismo , Inibidores de Histona Desacetilases/química , Inibidores de Histona Desacetilases/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/metabolismo , Humanos , Ácidos Hidroxâmicos/química , Ácidos Hidroxâmicos/metabolismo , Ácidos Hidroxâmicos/farmacologia , Concentração Inibidora 50 , Modelos Moleculares , Estrutura Molecular , Ligação Proteica , Estrutura Terciária de Proteína , Schistosoma mansoni/efeitos dos fármacos , Schistosoma mansoni/fisiologia , Esquistossomose mansoni/parasitologia , Tioacetamida/metabolismo , Tioacetamida/farmacologia , VorinostatRESUMO
The effect of histone deacetylases (HDACs) on normal and aberrant gene expression has been studied widely, making these enzymes interesting targets for the treatment of cancer and other diseases. In this chapter, we present in vitro assays that are commonly used to detect HDAC activity that do not rely on radioactive substrates and are amenable for high-throughput testing in microtiter plates. The major focus is on in vitro screening, but we also provide protocols to monitor HDAC activity from cancer cells and peripheral white blood cells. We will discuss the advantages and drawbacks of the respective protocols and give general hints and suggestions that are valuable to obtain reliable and reproducible results.
Assuntos
Bioensaio/métodos , Linhagem Celular Tumoral/enzimologia , Histona Desacetilases/metabolismo , Leucócitos/enzimologia , Acetilação , Polarização de Fluorescência , Histona Desacetilases/genética , Histonas/metabolismo , HumanosRESUMO
Lysine demethylases play an important role in epigenetic regulation and thus in the development of diseases like cancer or neurodegenerative disorders. As the lysine specific demethylase 1 (LSD1/KDM1) has been strongly connected to androgen and estrogen dependent gene expression, it serves as a promising target for the therapy of hormone dependent cancer. Here, we report on the discovery of new small molecule inhibitors of LSD1 containing a propargylamine warhead, starting out from lysine containing substrate analogues. On the basis of these substrate mimicking inhibitors, we were able to increase potency by a combination of similarity-based virtual screening and subsequent synthetic optimization resulting in more druglike LSD1 inhibitors that led to histone hypermethylation in breast cancer cells.
Assuntos
Aminas/química , Aminas/farmacologia , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Histona Desmetilases/antagonistas & inibidores , Aminas/síntese química , Aminas/metabolismo , Animais , Linhagem Celular Tumoral , Desenho de Fármacos , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/metabolismo , Histona Desmetilases/química , Histona Desmetilases/metabolismo , Histonas/metabolismo , Metilação/efeitos dos fármacos , Simulação de Acoplamento Molecular , Conformação ProteicaRESUMO
In the past years, a lot of attention has been given to the identification and characterization of selective and potent inhibitors of chromatin-modifying enzymes to better understand their specific role in transcriptional regulation. As aberrant histone methylation is involved in different pathological processes, the search for methyltransferase and demethylase inhibitors has emerged as a crucial issue in current medicinal chemistry research. High-throughput in vitro assays are important tools for the identification of new methyltransferase or demethylase inhibitors. These usually use oligopeptide substrates derived from histone sequences, although in many cases, they are not good substrates for these enzymes. Here, the authors report about the setup and establishment of in vitro assays that use native core histones as substrates, enabling an assay environment that better resembles native conditions. They have applied these substrates for the known formaldehyde dehydrogenase assay for the histone demethylase LSD1 and have established two new antibody-based assays. For LSD1, a heterogeneous assay format was set up, and a homogeneous assay was used for the characterization of the arginine methyltransferase PRMT1. Validation of the system was achieved with reference inhibitors in each case.
Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Epigênese Genética , Ensaios de Triagem em Larga Escala/métodos , Histonas/metabolismo , Aldeído Oxirredutases/metabolismo , Anticorpos/metabolismo , Relação Dose-Resposta a Droga , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/imunologia , Histona Desmetilases/metabolismo , Proteína-Arginina N-Metiltransferases/antagonistas & inibidores , Proteína-Arginina N-Metiltransferases/metabolismo , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/metabolismo , Reprodutibilidade dos TestesRESUMO
Epigenetics is defined as heritable changes in gene expression, i.e. the presentation of a certain phenotype, without alterations to the genetic code. This is a central phenomenon in developmental biology but also for the pathogenesis of a variety of diseases, e.g. for cancer in the case of silencing of tumor suppressor proteins. Reversible histone acetylation is an important player within the epigenetic machinery and histone deacetylases are the best studied epigenetic target for drug discovery. A first drug (vorinostat) is already approved for human cancer treatment. The strategies and protocols for monitoring histone deacetylase (HDAC) activity are an integral part of drug discovery approaches towards new HDAC inhibitors and will be reviewed in this article. The main focus is on biochemical in-vitro assays but cellular hit validation models are also presented.
Assuntos
Inibidores Enzimáticos/análise , Inibidores de Histona Desacetilases , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Humanos , Relação Estrutura-AtividadeRESUMO
Epigenetics is defined as heritable changes to the transcriptome that are independent of changes in the genome. The biochemical modifications that govern epigenetics are DNA methylation and posttranslational histone modifications. Among the histone modifications, acetylation and deacetylation are well characterized, whereas the fields of histone methylation and especially demethylation are still in their infancy. This is particularly true with regard to drug discovery. There is strong evidence that these modifications play an important role in the maintenance of transcription as well as in the development of certain diseases. This article gives an overview of the mechanisms of action of histone methyltransferases and demethylases, their role in the formation of certain diseases, and available inhibitors. Special emphasis is placed on the strategies that led to the first inhibitors which are currently available and the screening approaches that were used in that process.
Assuntos
Inibidores Enzimáticos/uso terapêutico , Histona Desmetilases/antagonistas & inibidores , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Neoplasias/enzimologia , Arginina/metabolismo , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Histona Desmetilases/classificação , Histona Desmetilases/metabolismo , Histona Metiltransferases , Histona-Lisina N-Metiltransferase/classificação , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Humanos , Lisina/metabolismo , Metilação , Neoplasias/tratamento farmacológico , Processamento de Proteína Pós-TraducionalRESUMO
The term epigenetics is defined as heritable changes in gene expression patterns that occur without changes in DNA sequence. Epigenetic changes according to this definition are achieved by methylation of cytosine bases in the DNA and by histone modifications, such as acetylation, methylation or phosphorylation. These modifications play an important role in regulating gene expression and the existence of an epigenetic code which maintains these modifications even upon cell division has been underlined by many investigations. Targeting the enzymes which catalyze DNA methylation or histone modifications may be a possibility not only for cancer therapy but also for chemoprevention since disruption of epigenetic balance is known to cause diseases such as cancer. In this review, we want to present the key epigenetic targets. We highlight natural products that modulate these epigenetic mechanisms and show their potential for cancer chemoprevention.