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3.
Life Sci ; 233: 116696, 2019 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-31351969

RESUMO

AIMS: To explore the mechanism of how LSD1 regulates autophagy and the correlation between LSD1 and Ox-LDL-induced inflammation. MAIN METHODS: RAW264.7 cells were used during the whole study. Firstly, the effect of Ox-LDL-stimulation on LSD1 expression was detected. Through loss-of-function assay, the associations between LSD1 interference and SESN2 expression, autophagy, NLRP3 inflammasome and inflammatory cytokines were explored. Finally, the function of LSD1 exerted on activation of PI3K/Akt/mTOR signal pathway was detected using western blotting assay. KEY FINDINGS: The expression of LSD1 was significantly elevated in Ox-LDL-treated RAW264.7 cells. Inhibition of LSD1 promoted autophagy, inhibited inflammation and activated NLRP3 inflammasome. SESN2 was elevated by LSD1 inhibition, and thus activate the PI3K/Akt/mTOR signal pathway. What' more, Knockdown of SESN2 or deactivate the PI3K/Akt/mTOR signal pathway partly reversed the effect of LSD1 inhibition on autophagy. SIGNIFICANCE: Our present study drew the finding that the knockdown of LSD1 meliorated Ox-LDL-stimulated NLRP3 activation and inflammation through promoting autophagy via SESN2-mediated PI3K/Akt/mTOR pathway.


Assuntos
Autofagia , Regulação da Expressão Gênica/efeitos dos fármacos , Histona Desmetilases/metabolismo , Inflamação/patologia , Lipoproteínas LDL/efeitos adversos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteínas Nucleares/metabolismo , Animais , Células Cultivadas , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/genética , Inflamação/etiologia , Inflamação/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteínas Nucleares/genética , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Interferente Pequeno/genética , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo
4.
Molecules ; 24(9)2019 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-31060229

RESUMO

Background: KDM5 enzymes are H3K4 specific histone demethylases involved in transcriptional regulation and DNA repair. These proteins are overexpressed in different kinds of cancer, including breast, prostate and bladder carcinomas, with positive effects on cancer proliferation and chemoresistance. For these reasons, these enzymes are potential therapeutic targets. Methods: In the present study, we analyzed the effects of three different inhibitors of KDM5 enzymes in MCF-7 breast cancer cells over-expressing one of them, namely KDM5B/JARID1B. In particular we tested H3K4 demethylation (western blot); radio-sensitivity (cytoxicity and clonogenic assays) and damage accumulation (COMET assay and kinetics of H2AX phosphorylation). Results: we show that all three compounds with completely different chemical structures can selectively inhibit KDM5 enzymes and are capable of increasing sensitivity of breast cancer cells to ionizing radiation and radiation-induced damage. Conclusions: These findings confirm the involvement of H3K4 specific demethylases in the response to DNA damage, show a requirement of the catalytic function and suggest new strategies for the therapeutic use of their inhibitors.


Assuntos
Neoplasias da Mama/enzimologia , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases com o Domínio Jumonji/genética , Proteínas Nucleares/genética , Radiossensibilizantes/farmacologia , Proteínas Repressoras/genética , Bibliotecas de Moléculas Pequenas/farmacologia , Neoplasias da Mama/genética , Neoplasias da Mama/terapia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/efeitos da radiação , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Histonas/metabolismo , Humanos , Histona Desmetilases com o Domínio Jumonji/metabolismo , Células MCF-7 , Modelos Moleculares , Estrutura Molecular , Proteínas Nucleares/metabolismo , Tolerância a Radiação/efeitos dos fármacos , Radiossensibilizantes/química , Proteínas Repressoras/metabolismo , Bibliotecas de Moléculas Pequenas/química , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/efeitos da radiação
5.
Eur J Med Chem ; 175: 357-372, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31096156

RESUMO

Lysine-specific demethylase 1 (LSD1), demethylase against mono- and di - methylated histone3 lysine 4, has emerged as a promising target in oncology. More specifically, it has been demonstrated as a key promoter in acute myeloid leukemia (AML), and several LSD1 inhibitors have already entered into clinical trials for the treatment of AML. In this paper, a series of new indole derivatives were designed and synthesized based on a lead compound obtained by a high-throughput screening with our in-house compound library. Among the synthetic compounds, 9e was characterized as a potent LSD1 inhibitor with an IC50 of 1.230 µM and can inhibit the proliferation of THP-1 cells effectively. And most importantly, this is the first irreversible LSD1 inhibitor that is not derived from monoamine oxidase inhibitors. Hence, the discovery of 9e may serve as a proof of concept work for AML treatment.


Assuntos
Descoberta de Drogas , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Histona Desmetilases/antagonistas & inibidores , Indóis/síntese química , Indóis/farmacologia , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Furanos/química , Ensaios de Triagem em Larga Escala , Histona Desmetilases/metabolismo , Humanos , Indóis/química , Indóis/metabolismo , Concentração Inibidora 50 , Leucemia Mieloide Aguda/patologia , Proteínas Recombinantes/efeitos dos fármacos , Proteínas Recombinantes/metabolismo , Relação Estrutura-Atividade
6.
Biochim Biophys Acta Gene Regul Mech ; 1862(5): 535-546, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30951900

RESUMO

Senescence is a stress-responsive cellular program that leads to cell cycle arrest. In cancer cells, senescence has profound implications for tumor aggressiveness and clinical outcome, but the molecular events that provoke cancer cells to undergo senescence remain unclear. Herein, we provide evidence that the histone demethylase LSD1/KDM1A supports the growth of Glioblastoma tumor cells and its inhibition triggers senescence response. LSD1 is a histone modifier that participates in key aspects of gene transcription as well as in the regulation of methylation dynamics of non-histone proteins. We found that down-regulation of LSD1 inhibits Glioblastoma cell growth, impairs mTOR pathway and cell migration and induces senescence. At mechanistic level, we found that LSD1 regulates HIF-1α protein stability. Pharmacological inhibition or siRNA-mediated silencing of LSD1 expression effectively reduces HIF-1α protein levels, which suffices for the induction of senescence. Our findings elucidate a mechanism whereby LSD1 controls senescence in Glioblastoma tumor cells through the regulation of HIF-1α, and we propose the novel defined LSD1/HIF-1α axis as a new target for the therapy of Glioblastoma tumors.


Assuntos
Senescência Celular , Glioblastoma/enzimologia , Histona Desmetilases/fisiologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Hipóxia Celular , Linhagem Celular Tumoral , Sobrevivência Celular , Inibidores Enzimáticos/farmacologia , Glioblastoma/metabolismo , Glioblastoma/patologia , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/metabolismo , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Mitocôndrias/metabolismo , Tranilcipromina/farmacologia
7.
Nat Chem Biol ; 15(5): 529-539, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30992567

RESUMO

Understanding the mechanism of small molecules is a critical challenge in chemical biology and drug discovery. Medicinal chemistry is essential for elucidating drug mechanism, enabling variation of small molecule structure to gain structure-activity relationships (SARs). However, the development of complementary approaches that systematically vary target protein structure could provide equally informative SARs for investigating drug mechanism and protein function. Here we explore the ability of CRISPR-Cas9 mutagenesis to profile the interactions between lysine-specific histone demethylase 1 (LSD1) and chemical inhibitors in the context of acute myeloid leukemia (AML). Through this approach, termed CRISPR-suppressor scanning, we elucidate drug mechanism of action by showing that LSD1 enzyme activity is not required for AML survival and that LSD1 inhibitors instead function by disrupting interactions between LSD1 and the transcription factor GFI1B on chromatin. Our studies clarify how LSD1 inhibitors mechanistically operate in AML and demonstrate how CRISPR-suppressor scanning can uncover novel aspects of target biology.


Assuntos
Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Leucemia Mieloide Aguda/metabolismo , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/efeitos dos fármacos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Histona Desmetilases/antagonistas & inibidores , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Modelos Moleculares , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia
8.
Eur J Med Chem ; 166: 432-444, 2019 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-30739825

RESUMO

Phenelzine was first employed to design new aryl hydrazine-based LSD1 inhibitors based on the experience-based discovery (EBD) strategy. Among these compounds, D8 potently inhibited LSD1 (IC50 = 882.30 nM) in a reversible manner. Compound D8 was selective to LSD1 over MAO-A/B and showed H3K4me2 competitive binding to LSD1. The interaction between H3K4me2 and LSD1 was also confirmed by the Co-IP assay. In LSD1 overexpressed A549 cells, compound D8 dose-dependently induced accumulation of LSD1 substrates H3K4me1/2 and H3K9me1/2, showed cellular target engagement to LSD1 and significantly inhibited cell migration of A549 cells. Docking studies suggested that compound D8 occupied the peptide binding region and therefore blocked the access of the peptide substrate to the FAD, finally leading to the demethylase activity inhibition of LSD1. The findings indicate that aryl hydrazines are new scaffolds for the design of LSD1 inhibitors, the identification of D8 provides further evidence for our previously proposed general principle that fused heterocycles with an amine group are potentially active toward LSD1 by competitive binding to LSD1 with H3 peptide substrates.


Assuntos
Desenho de Drogas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Histona Desmetilases/antagonistas & inibidores , Hidrazinas/química , Hidrazinas/farmacologia , Células A549 , Ligação Competitiva , Sobrevivência Celular/efeitos dos fármacos , Inibidores Enzimáticos/metabolismo , Histona Desmetilases/metabolismo , Histonas/metabolismo , Humanos , Hidrazinas/metabolismo , Metilação/efeitos dos fármacos , Relação Estrutura-Atividade
9.
Eur J Med Chem ; 167: 388-401, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-30780087

RESUMO

The histone lysine specific demethylase 1 (LSD1/KDM1A) is implicated in the development of cancers, targeting LSD1 has been recognized as a promising strategy for cancer therapy. To date, some small-molecule inhibitors are currently being investigated in clinical trials. Herein we report the design, synthesis and biochemical characterization of [1,2,4]triazolo[1,5-a]pyrimidine derivatives as new LSD1 inhibitors. Of these compounds, compound C26 inhibited LSD1 in a reversible manner (IC50 = 1.72 µM) and showed selectivity to LSD1 over MAO-A/B. Besides, compound C26 displayed FAD-competitive binding to LSD1. Interestingly, C26 did not inhibit horseradish peroxidase (HRP) and quench H2O2, thus excluding the possibility that LSD1 inhibition by C26 was due to the HRP inhibition and consumption of H2O2. In LSD1 overexpressed A549 cells, compound C26 concentration-dependently induced accumulation of H3K4me1/me2 and H3K9me2 and showed cellular target engagement to LSD1. Additionally, compound C26 significantly inhibited migration of A549 cells in a concentration-dependent manner, further western blot analysis showed that C26 increased expression levels of epithelial cell markers E-Cadherin and Claudin-1, down-regulated mesenchymal cell marker N-Cadherin and the upstream transcription factors Snail and Slug. Docking studies were also performed to rationalize the potency of C26 toward LSD1. To conclude, the [1,2,4]triazolo[1,5-a]pyrimidine could serve as a promising scaffold for the development of new LSD1 inhibitors.


Assuntos
Desenho de Drogas , Inibidores Enzimáticos/síntese química , Histona Desmetilases/antagonistas & inibidores , Pirimidinas/farmacologia , Triazóis/farmacologia , Ligação Competitiva , Caderinas/metabolismo , Claudina-1/metabolismo , Inibidores Enzimáticos/farmacologia , Humanos , Simulação de Acoplamento Molecular , Pirimidinas/síntese química , Relação Estrutura-Atividade , Fatores de Transcrição/metabolismo , Triazóis/síntese química
10.
Leukemia ; 33(6): 1411-1426, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30679800

RESUMO

LSD1 has emerged as a promising epigenetic target in the treatment of acute myeloid leukemia (AML). We used two murine AML models based on retroviral overexpression of Hoxa9/Meis1 (H9M) or MN1 to study LSD1 loss of function in AML. The conditional knockout of Lsd1 resulted in differentiation with both granulocytic and monocytic features and increased ATRA sensitivity and extended the survival of mice with H9M-driven AML. The conditional knockout led to an increased expression of multiple genes regulated by the important myeloid transcription factors GFI1 and PU.1. These include the transcription factors GFI1B and IRF8. We also compared the effect of different irreversible and reversible inhibitors of LSD1 in AML and could show that only tranylcypromine derivatives were capable of inducing a differentiation response. We employed a conditional knock-in model of inactive, mutant LSD1 to study the effect of only interfering with LSD1 enzymatic activity. While this was sufficient to initiate differentiation, it did not result in a survival benefit in mice. Hence, we believe that targeting both enzymatic and scaffolding functions of LSD1 is required to efficiently treat AML. This finding as well as the identified biomarkers may be relevant for the treatment of AML patients with LSD1 inhibitors.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Proteínas de Ligação a DNA/metabolismo , Histona Desmetilases/antagonistas & inibidores , Leucemia Mieloide Aguda/patologia , Proteínas Proto-Oncogênicas/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Tranilcipromina/farmacologia , Animais , Antidepressivos/farmacologia , Proteínas de Ligação a DNA/genética , Regulação Leucêmica da Expressão Gênica , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Histona Desmetilases/fisiologia , Humanos , Fatores Reguladores de Interferon/genética , Fatores Reguladores de Interferon/metabolismo , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Camundongos , Camundongos Knockout , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Proteínas Proto-Oncogênicas/genética , Transativadores/genética , Fatores de Transcrição/genética , Células Tumorais Cultivadas
11.
Biomed Pharmacother ; 109: 1994-2004, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30551455

RESUMO

The prognosis of oral squamous cell carcinoma (OSCC) patients remains unclear, and a better understanding of the underlying molecular mechanisms is urgently required. Jumonji-C (JmjC) domain-containing protein 5 (JMJD5), renamed KDM8, has been implicated in tumorigenesis, circadian rhythm modulation, embryological development, and osteoclastogenesis. In the present study, we found that JMJD5 was over-expressed in patients with OSCC by real-time quantitative PCR (qPCR), western blot and immunohistochemical assays. When knockdown using small interfering RNA (siRNA) in OSCCs, JMJD5 was exhibited to be important for sustaining cell migration and invasion. JMJD5-knockdown increased E-cadherin expressions, and decreased N-cadherin and Vimentin expression levels in OSCC cells. Further, apoptosis was induced by JMJD5-silence through both the intrinsic and extrinsic pathways, as evidenced by the increased cleavage of Caspase-8/-9/-3 and PARP. Meanwhile, p53 expression levels were also up-regulated by JMJD5-knockdown. Suppressing p53 expressions with its inhibitor, PFTα, blocked apoptotic response in JMJD5-silenced cells. JMJD5 inhibition-induced decrease of nuclear factor-kappaB (NF-κB) was rescued by pifithrin-α (PFTα) pre-treatment. Consistently, over-expressing JMJD5 decreased p53, cleaved Caspase-3 and poly (ADP-ribose) polymerase-1 (PARP-1), whereas increased nuclear NF-κB expressions in OSCC cell lines. More importantly, targeting JMJD5 reduced xenograft tumor growth in vivo through the same molecular mechanisms evidenced in vitro. Thus, the data supplied mechanistic insights into the effects of JMJD5 on the modulation of OSCC development, illustrating that JMJD5 might be an essential prognostic indicator and therapeutic target against OSCC progression.


Assuntos
Carcinoma de Células Escamosas/metabolismo , Regulação para Baixo/fisiologia , Histona Desmetilases/metabolismo , Neoplasias Bucais/metabolismo , NF-kappa B/fisiologia , Proteína Supressora de Tumor p53/metabolismo , Animais , Biomarcadores Tumorais/antagonistas & inibidores , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Técnicas de Silenciamento de Genes/métodos , Histona Desmetilases/antagonistas & inibidores , Humanos , Masculino , Camundongos , Camundongos Nus , NF-kappa B/antagonistas & inibidores , Invasividade Neoplásica/prevenção & controle , Transdução de Sinais/fisiologia , Carga Tumoral/fisiologia , Proteína Supressora de Tumor p53/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
12.
Nat Commun ; 9(1): 5230, 2018 12 07.
Artigo em Inglês | MEDLINE | ID: mdl-30531796

RESUMO

Left ventricular hypertrophy (LVH) is a major risk factor for cardiovascular morbidity and mortality. Pathological LVH engages transcriptional programs including reactivation of canonical fetal genes and those inducing fibrosis. Histone lysine demethylases (KDMs) are emerging regulators of transcriptional reprogramming in cancer, though their potential role in abnormal heart growth and fibrosis remains little understood. Here, we investigate gain and loss of function of an H3K9me2 specific demethylase, Kdm3a, and show it promotes LVH and fibrosis in response to pressure-overload. Cardiomyocyte KDM3A activates Timp1 transcription with pro-fibrotic activity. By contrast, a pan-KDM inhibitor, JIB-04, suppresses pressure overload-induced LVH and fibrosis. JIB-04 inhibits KDM3A and suppresses the transcription of fibrotic genes that overlap with genes downregulated in Kdm3a-KO mice versus WT controls. Our study provides genetic and biochemical evidence for a pro-hypertrophic function of KDM3A and proof-of principle for pharmacological targeting of KDMs as an effective strategy to counter LVH and pathological fibrosis.


Assuntos
Cardiomegalia/genética , Regulação da Expressão Gênica/genética , Histona Desmetilases/genética , Miocárdio/metabolismo , Aminopiridinas/farmacologia , Animais , Animais Recém-Nascidos , Cardiomegalia/enzimologia , Células Cultivadas , Fibrose/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/metabolismo , Humanos , Hidrazonas/farmacologia , Camundongos Knockout , Camundongos Transgênicos , Miocárdio/enzimologia , Miocárdio/patologia , Miócitos Cardíacos/metabolismo , Ratos Sprague-Dawley
13.
Molecules ; 23(5)2018 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-29734782

RESUMO

Lysine-specific demethylase 1 (LSD1) mainly removes methyl groups of mono- or di-methylated lysine residues at the fourth position of histone H3 to epigenetically regulate the expression of genes associated with several diseases, such as cancer. Therefore, LSD1 inactivators are expected to be used as therapeutic agents. In this study, to identify novel peptide-based LSD1 inactivators, we focused on the X-ray structure of LSD1 complexed with a H3 peptide-based suicide substrate. It has been proposed that a methylated histone substrate forms three consecutive γ-turn structures in the active pocket of LSD1. Based on this, we designed and synthesized novel histone H3 peptide-based LSD1 inactivators 2a⁻c by incorporating various α,α-disubstituted amino acids with γ-turn-inducing structures. Among synthetic peptides 2a⁻c, peptide 2b incorporating two 1-aminocyclohexanecarboxylic acids at both sides of a lysine residue bearing a trans-2-phenylcyclopropylamine (PCPA) moiety, which is a pharmacophore for LSD1 inactivation, was the most potent and selective LSD1 inactivator. These findings are useful for the further development of histone H3 peptide-based LSD1 inactivators.


Assuntos
Aminoácidos/síntese química , Inibidores Enzimáticos/síntese química , Histona Desmetilases/antagonistas & inibidores , Histonas/química , Lisina/química , Peptídeos/síntese química , Aminoácidos Cíclicos/química , Domínio Catalítico , Ácidos Cicloexanocarboxílicos/química , Desenho de Drogas , Ensaios Enzimáticos , Inibidores Enzimáticos/farmacologia , Histona Desmetilases/química , Histona Desmetilases/metabolismo , Histonas/metabolismo , Humanos , Hidrólise , Isoenzimas/química , Isoenzimas/metabolismo , Lisina/metabolismo , Metilação , Simulação de Acoplamento Molecular , Monoaminoxidase/química , Monoaminoxidase/metabolismo , Peptídeos/farmacologia , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Relação Estrutura-Atividade , Tranilcipromina/química
14.
Diabetes ; 67(6): 1113-1127, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29618580

RESUMO

Recent studies have emphasized the association of adipose oxidative stress (Fat reactive oxygen species [ROS]) with the pathogenesis of metabolic disorders in obesity. However, the causal roles of Fat ROS in metabolic disturbances in vivo remain unclear because no mouse model has been available in which oxidative stress is manipulated by targeting adipocytes. In this research, we generated two models of Fat ROS-manipulated mice and evaluated the metabolic features in diet-induced obesity. Fat ROS-eliminated mice, in which Cat and Sod1 were overexpressed in adipocytes, exhibited adipose expansion with decreased ectopic lipid accumulation and improved insulin sensitivity. Conversely, Fat ROS-augmented mice, in which glutathione was depleted specifically in adipocytes, exhibited restricted adipose expansion associated with increased ectopic lipid accumulation and deteriorated insulin sensitivity. In the white adipose tissues of these mice, macrophage polarization, tissue fibrosis, and de novo lipogenesis were significantly changed. In vitro approaches identified KDM1A-mediated attenuation of sterol-regulatory element-binding transcription factor 1 (SREBF1) transcriptional activities as the underlying mechanism for the suppression of de novo lipogenesis by oxidative stress. Thus, our study uncovered the novel roles of Fat ROS in healthy adipose expansion, ectopic lipid accumulation, and insulin resistance, providing the possibility for the adipocyte-targeting antioxidant therapy.


Assuntos
Tecido Adiposo Branco/metabolismo , Regulação Enzimológica da Expressão Gênica , Histona Desmetilases/antagonistas & inibidores , Lipogênese , Obesidade/metabolismo , Estresse Oxidativo , Proteína de Ligação a Elemento Regulador de Esterol 1/antagonistas & inibidores , Células 3T3-L1 , Adipogenia , Tecido Adiposo Branco/imunologia , Tecido Adiposo Branco/patologia , Animais , Biomarcadores/sangue , Biomarcadores/metabolismo , Cruzamentos Genéticos , Dieta Ocidental/efeitos adversos , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Humanos , Resistência à Insulina , Ativação de Macrófagos , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Obesidade/etiologia , Obesidade/imunologia , Obesidade/patologia , Interferência de RNA , Ratos , Organismos Livres de Patógenos Específicos , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo
15.
Cell Prolif ; 51(4): e12459, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29656462

RESUMO

OBJECTIVES: Dental tissue-derived mesenchymal stem cells (MSCs)-mediated pulp-dentin regeneration is considered a potential approach for the regeneration of damaged teeth. Enhancing MSC-mediated pulp-dentin regeneration is based on an understanding of the molecular mechanisms underlying directed cell differentiation process. Histone demethylation enzyme, lysine demethylase 1A (KDM1A) can regulate the differentiation of some MSCs, but its role in dental tissue-derived MSCs is unclear. MATERIAL AND METHODS: We obtained SCAPs from immature teeth. Alkaline phosphatase (ALP) activity assay, Alizarin red staining, quantitative calcium analysis, osteogenesis-related genes expression and in vivo transplantation experiment were used to explore the osteo/dentinogenic differentiation. Co-immunoprecipitation (Co-IP) assay was used to investigate the binding protein. RESULTS: Knock-down of KDM1A reduced ALP activity and mineralization, promoted the expressions of osteo/dentinogenic differentiation markers DSPP, DMP1, BSP and key transcript factors, RUNX2, OSX, DLX2 in SCAPs, and also enhanced the osteo/dentinogenesis in vivo. In addition, KDM1A could associate with PLOD2 to form protein complex. And knock-down of PLOD2 inhibited ALP activity and mineralization, and promoted the expressions of DSPP, DMP1, BSP, RUNX2, OSX and DLX2 in SCAPs. CONCLUSIONS: KDM1A might have different role in different stages of osteo/dentinogenic differentiation process by binding partner with PLOD2, and finally resulted in the inhibited function for the osteo/dentinogenesis in SCAPs. Our studies provided a further understanding of the regulatory mechanisms of dynamic osteo/dentinogenic differentiation process in dental tissue MSCs.


Assuntos
Dentinogênese/fisiologia , Histona Desmetilases/metabolismo , Osteogênese/fisiologia , Pró-Colágeno-Lisina 2-Oxoglutarato 5-Dioxigenase/metabolismo , Animais , Diferenciação Celular , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Papila Dentária/citologia , Feminino , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Camundongos Nus , Pró-Colágeno-Lisina 2-Oxoglutarato 5-Dioxigenase/antagonistas & inibidores , Pró-Colágeno-Lisina 2-Oxoglutarato 5-Dioxigenase/genética , Ligação Proteica , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Fator de Transcrição Sp7/metabolismo , Transplante de Células-Tronco , Células-Tronco/citologia , Células-Tronco/metabolismo , Fatores de Transcrição/metabolismo
16.
Anal Chim Acta ; 1016: 59-68, 2018 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-29534805

RESUMO

Countercurrent chromatography (CCC) has gradually become a widely used method for preparative separation of bioactive natural molecules. These molecules generally contain distinct scaffolds and characteristics that cannot be readily isolated from plants. While one-dimensional CCC is typically used for the initial purification with insufficiently resolved peaks after locating bioactive components, two-dimensional (2D) or multi-dimensional CCC strategies are employed to improve the resolution of peaks. However, these methods usually present certain disadvantages, such as complicated procedures and increased time consumption, experimental costs, and equipment requirements. Here, a bioactivity-guided cut CCC strategy was established to isolate lysine-specific demethylase 1 (LSD1) inhibitors from Scutellaria baicalensis Georgi. Gradient-elution CCC coupled with real-time detection of LSD1 inhibition by the collected fractions was developed. Next, an online-storage recycling CCC mode was designed to enable the active fractions to be stored in coils, and these active fractions were further separated to obtain pure compounds by using sequential recycling elution. In this strategy, active fractions are first identified, and then pure LSD1 inhibitors are isolated in the 2D CCC mode through continuous separation on a single instrument. By using our bioactivity-guided cut CCC strategy, we successfully isolated six natural LSD1 inhibitors from S. baicalensis Georgi, five of which were identified for the first time as natural LSD1 inhibitors.


Assuntos
Inibidores Enzimáticos/isolamento & purificação , Extratos Vegetais/isolamento & purificação , Scutellaria baicalensis/química , Linhagem Celular Tumoral , Distribuição Contracorrente , Inibidores Enzimáticos/química , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/metabolismo , Humanos , Extração Líquido-Líquido , Simulação de Acoplamento Molecular , Extratos Vegetais/química
17.
Proc Natl Acad Sci U S A ; 115(18): E4179-E4188, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29581250

RESUMO

Medical castration that interferes with androgen receptor (AR) function is the principal treatment for advanced prostate cancer. However, clinical progression is universal, and tumors with AR-independent resistance mechanisms appear to be increasing in frequency. Consequently, there is an urgent need to develop new treatments targeting molecular pathways enriched in lethal prostate cancer. Lysine-specific demethylase 1 (LSD1) is a histone demethylase and an important regulator of gene expression. Here, we show that LSD1 promotes the survival of prostate cancer cells, including those that are castration-resistant, independently of its demethylase function and of the AR. Importantly, this effect is explained in part by activation of a lethal prostate cancer gene network in collaboration with LSD1's binding protein, ZNF217. Finally, that a small-molecule LSD1 inhibitor-SP-2509-blocks important demethylase-independent functions and suppresses castration-resistant prostate cancer cell viability demonstrates the potential of LSD1 inhibition in this disease.


Assuntos
Redes Reguladoras de Genes , Histona Desmetilases/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias de Próstata Resistentes à Castração/enzimologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Histona Desmetilases/antagonistas & inibidores , Histona Desmetilases/genética , Humanos , Hidrazinas/farmacologia , Masculino , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Sulfonamidas/farmacologia , Transativadores/genética , Transativadores/metabolismo
18.
Eur J Med Chem ; 148: 210-220, 2018 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-29459279

RESUMO

Lysine-specific demethylase 1 (LSD1) is a chromatin-remodeling enzyme that plays an important role in cancer. Over-expression of LSD1 decreases methylation at histone 3 lysine 4, and aberrantly silences tumor suppressor genes. Inhibitors of LSD1 have been designed as chemical probes and potential antitumor agents. We recently reported the cyclic peptide 9, which potently and reversibly inhibits LSD1 (IC50 2.1 µM; Ki 385 nM). Systematic alanine mutagenesis of 9 revealed residues that are critical for LSD1 inhibition, and these mutated peptides were evaluated as LSD1 inhibitors. Alanine substitution at positions 2, 3, 4, 6 and 11-17 preserved inhibition, while substitution of alanine at positions 8 and 9 resulted in complete loss of activity. Cyclic mutant peptides 11 and 16 produced the greatest LSD1 inhibition, and 11, 16, 27 and 28 increased global H3K4me2 in K562 cells. In addition, 16, 27 and 28 promoted significant increases in H3K4me2 levels at the promoter sites of the genes IGFBP2 and FEZ1. Data from these LSD1 inhibitors will aid in the design of peptidomimetics with improved stability and pharmacokinetics.


Assuntos
Alanina/genética , Antineoplásicos/química , Histona Desmetilases/antagonistas & inibidores , Mutagênese Sítio-Dirigida , Peptídeos Cíclicos/farmacologia , Antineoplásicos/farmacologia , Humanos , Células K562
19.
Bioorg Med Chem ; 26(8): 1523-1537, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29439916

RESUMO

LSD1/KDM1 is a histone demethylase that preferentially removes methyl groups from the mono- and di-methylated lysine 4 in histone H3 (H3K4), key marks for active chromatin for transcriptional activation. LSD1 is essential for pluripotent embryonic stem cells and embryonic teratocarcinoma/carcinoma cells and its expression is often elevated in various cancers. We developed a new LSD1 inhibitor, CBB3001, which potently inhibited LSD1 activity both in vitro and in vivo. CBB3001 also selectively inhibited the growth of human ovarian teratocarcinoma PA-1 and mouse embryonic carcinoma F9 cells, caused the downregulation of pluripotent stem cell proteins SOX2 and OCT4. However, CBB3001 does not have significant inhibition on the growth of human colorectal carcinoma HCT116 cells or mouse fibroblast NIH3T3 cells that do not express these stem cell proteins. Our studies strongly indicate that CBB3001 is a specific LSD1 inhibitor that selectively inhibits teratocarcinoma and embryonic carcinoma cells that express SOX2 and OCT4.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Embrionário/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Histona Desmetilases/antagonistas & inibidores , Teratocarcinoma/tratamento farmacológico , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Carcinoma Embrionário/metabolismo , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Células HCT116 , Histona Desmetilases/metabolismo , Humanos , Camundongos , Estrutura Molecular , Células NIH 3T3 , Relação Estrutura-Atividade , Teratocarcinoma/metabolismo
20.
Bioorg Med Chem ; 26(3): 775-785, 2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29331452

RESUMO

Lysine-specific demethylase 1 (LSD1) is an attractive molecular target for cancer therapy. We have previously reported potent LSD1-selective inhibitors (i.e., NCD18, NCD38, and their analogs) consisting of trans-2-phenylcyclopropylamine (PCPA) or trans-2-arylcyclopropylamine (ACPA) and a lysine moiety that could form a γ-turn structure in the active site of LSD1. Herein we report the design, synthesis and evaluation of γ-turn mimetic compounds for further improvement of LSD1 inhibitory activity and anticancer activity. Among a series of γ-turn mimetic compounds synthesized by a Mitsunobu-reaction-based amination strategy, we identified 1n as a potent and selective LSD1 inhibitor. Compound 1n induced cell cycle arrest and apoptosis through histone methylation in human lung cancer cells. The γ-turn mimetics approach should offer new insights into drug design for LSD1-selective inhibitors.


Assuntos
Ciclopropanos/farmacologia , Desenho de Drogas , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacologia , Histona Desmetilases/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Domínio Catalítico , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ciclopropanos/síntese química , Ciclopropanos/química , Ensaios Enzimáticos , Inibidores Enzimáticos/química , Histona Desmetilases/metabolismo , Humanos , Monoaminoxidase/química , Monoaminoxidase/metabolismo
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