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1.
Mol Cell ; 79(6): 1008-1023.e4, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32871104

RESUMO

TMPRSS2-ERG gene fusion occurs in approximately 50% of cases of prostate cancer (PCa), and the fusion product is a key driver of prostate oncogenesis. However, how to leverage cellular signaling to ablate TMPRSS2-ERG oncoprotein for PCa treatment remains elusive. Here, we demonstrate that DNA damage induces proteasomal degradation of wild-type ERG and TMPRSS2-ERG oncoprotein through ERG threonine-187 and tyrosine-190 phosphorylation mediated by GSK3ß and WEE1, respectively. The dual phosphorylation triggers ERG recognition and degradation by the E3 ubiquitin ligase FBW7 in a manner independent of a canonical degron. DNA damage-induced TMPRSS2-ERG degradation was abolished by cancer-associated PTEN deletion or GSK3ß inactivation. Blockade of DNA damage-induced TMPRSS2-ERG oncoprotein degradation causes chemotherapy-resistant growth of fusion-positive PCa cells in culture and in mice. Our findings uncover a previously unrecognized TMPRSS2-ERG protein destruction mechanism and demonstrate that intact PTEN and GSK3ß signaling are essential for effective targeting of ERG protein by genotoxic therapeutics in fusion-positive PCa.


Assuntos
Proteínas de Ciclo Celular/genética , Glicogênio Sintase Quinase 3 beta/genética , Proteínas de Fusão Oncogênica/genética , PTEN Fosfo-Hidrolase/genética , Neoplasias da Próstata/genética , Proteínas Tirosina Quinases/genética , Animais , Carcinogênese/genética , Linhagem Celular Tumoral , Dano ao DNA/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Tratamento Farmacológico , Proteína 7 com Repetições F-Box-WD/genética , Xenoenxertos , Humanos , Masculino , Camundongos , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Proteólise/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
2.
Nat Commun ; 11(1): 4311, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32855420

RESUMO

Pulmonary disease increases the risk of developing abdominal aortic aneurysms (AAA). However, the mechanism underlying the pathological dialogue between the lungs and aorta is undefined. Here, we find that inflicting acute lung injury (ALI) to mice doubles their incidence of AAA and accelerates macrophage-driven proteolytic damage of the aortic wall. ALI-induced HMGB1 leaks and is captured by arterial macrophages thereby altering their mitochondrial metabolism through RIPK3. RIPK3 promotes mitochondrial fission leading to elevated oxidative stress via DRP1. This triggers MMP12 to lyse arterial matrix, thereby stimulating AAA. Administration of recombinant HMGB1 to WT, but not Ripk3-/- mice, recapitulates ALI-induced proteolytic collapse of arterial architecture. Deletion of RIPK3 in myeloid cells, DRP1 or MMP12 suppression in ALI-inflicted mice repress arterial stress and brake MMP12 release by transmural macrophages thereby maintaining a strengthened arterial framework refractory to AAA. Our results establish an inter-organ circuitry that alerts arterial macrophages to regulate vascular remodeling.


Assuntos
Lesão Pulmonar Aguda/complicações , Aneurisma da Aorta Abdominal/patologia , Proteína HMGB1/metabolismo , Macrófagos/metabolismo , Remodelação Vascular , Lesão Pulmonar Aguda/patologia , Animais , Aorta Abdominal/citologia , Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/etiologia , Aneurisma da Aorta Abdominal/prevenção & controle , Células Cultivadas , Modelos Animais de Doenças , Dinaminas/antagonistas & inibidores , Dinaminas/metabolismo , Humanos , Macrófagos/citologia , Metaloproteinase 12 da Matriz/genética , Metaloproteinase 12 da Matriz/metabolismo , Camundongos , Camundongos Knockout , Dinâmica Mitocondrial/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Fosforilação , Cultura Primária de Células , Proteólise/efeitos dos fármacos , Doença Pulmonar Obstrutiva Crônica/complicações , Doença Pulmonar Obstrutiva Crônica/patologia , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Estudos Retrospectivos , Regulação para Cima
3.
Nat Commun ; 11(1): 3660, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32694521

RESUMO

High expression or aberrant activation of epidermal growth factor receptor (EGFR) is related to tumor progression and therapy resistance across cancer types, including non-small cell lung cancer (NSCLC). EGFR tyrosine kinase inhibitors (TKIs) are first-line therapy for NSCLC. However, patients eventually deteriorate after inevitable acquisition of EGFR TKI-resistant mutations, highlighting the need for therapeutics with alternative mechanisms of action. Here, we report that the elevated tribbles pseudokinase 3 (TRIB3) is positively associated with EGFR stability and NSCLC progression. TRIB3 interacts with EGFR and recruits PKCα to induce a Thr654 phosphorylation and WWP1-induced Lys689 ubiquitination in the EGFR juxtamembrane region, which enhances EGFR recycling, stability, downstream activity, and NSCLC stemness. Disturbing the TRIB3-EGFR interaction with a stapled peptide attenuates NSCLC progression by accelerating EGFR degradation and sensitizes NSCLC cells to chemotherapeutic agents. These findings indicate that targeting EGFR degradation is a previously unappreciated therapeutic option in EGFR-related NSCLC.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Proteínas de Ciclo Celular/metabolismo , Neoplasias Pulmonares/patologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Repressoras/metabolismo , Adulto , Animais , Antineoplásicos/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/mortalidade , Linhagem Celular Tumoral , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Receptores ErbB/metabolismo , Feminino , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidade , Masculino , Camundongos , Pessoa de Meia-Idade , Mutação , Fosforilação/efeitos dos fármacos , Proteína Quinase C-alfa/metabolismo , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Estabilidade Proteica/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/metabolismo , Proteólise/efeitos dos fármacos , Taxa de Sobrevida , Ubiquitinação/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
4.
PLoS One ; 15(7): e0236403, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32716961

RESUMO

Autophagy, a self-degradative physiological process, is critical for homeostasis maintenance and energy source balancing in response to various stresses, including nutrient deprivation. It is a highly conserved catabolic process in eukaryotes and is indispensable for cell survival as it involves degradation of unessential or excessive components and their subsequent recycling as building blocks for the synthesis of necessary molecules. Although the dysregulation of autophagy has been reported to broadly contribute to various diseases, including cancers and neurodegenerative diseases, the molecular mechanisms underlying the epigenetic regulation of autophagy are poorly elucidated. Here, we report that the level of lysine demethylase 3B (KDM3B) increases in nutrient-deprived HCT116 cells, a colorectal carcinoma cell line, resulting in transcriptional activation of the autophagy-inducing genes. KDM3B was found to enhance the transcription by demethylating H3K9me2 on the promoter of these genes. Furthermore, we observed that the depletion of KDM3B inhibited the autophagic flux in HCT116 cells. Collectively, these data suggested the critical role of KDM3B in the regulation of autophagy-related genes via H3K9me2 demethylation and induction of autophagy in nutrient-starved HCT116 cells.


Assuntos
Autofagia/genética , Histona Desmetilases com o Domínio Jumonji/metabolismo , Ativação Transcricional/genética , Aminoácidos/deficiência , Autofagia/efeitos dos fármacos , Epigênese Genética/efeitos dos fármacos , Células HCT116 , Células HEK293 , Histonas/metabolismo , Humanos , Regiões Promotoras Genéticas , Complexo de Endopeptidases do Proteassoma/metabolismo , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Proteólise/efeitos dos fármacos , Sirolimo/farmacologia , Ativação Transcricional/efeitos dos fármacos , Proteína com Valosina/metabolismo
5.
PLoS One ; 15(7): e0236175, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32697798

RESUMO

Adenoviruses cause upper respiratory infections, conjunctivitis, keratitis, and gastrointestinal illness. These can be fatal in immunocompromised individuals. Adenoviruses have also been engineered into viral vectors to deliver therapeutic genes or induce immunity as vaccine carriers. The success of ocular gene therapy is driven partly by the immunologic and biochemical influences of the intraocular environment. We have shown that versican and hyaluronan modulate adenoviral vector transgene expression through CD44 signaling. Herein we explored the role of these pathways on virus replication and viral protein expression of wild type adenovirus. We report that the addition of vitreous humor (which contains both versican and hyaluronan) increases viral hexon protein levels. Vitreous humor also increased wild type adenovirus DNA replication in vitro. Metalloproteinase and γ-secretase inhibitors, which inhibit CD44 proteolytic activation, blocked adenoviral replication in vitro. Similarly, protein kinase C and RhoA kinase inhibitors, both proteins associated with CD44 mediated pathways, also inhibited wild type adenoviral replication in vitro. Application of metalloproteinase and γ-secretase inhibitors to human conjunctival explants sharply decreased adenoviral vector gene expression. Our results demonstrate that pharmacologic delivery of these inhibitors is easily achievable. The inhibition of these enzymes should be explored as potential therapies of wild type adenoviral infections.


Assuntos
Infecções por Adenoviridae/tratamento farmacológico , Adenoviridae/efeitos dos fármacos , Antivirais/farmacologia , Vetores Genéticos/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Adenoviridae/fisiologia , Infecções por Adenoviridae/virologia , Administração Oftálmica , Amidas/farmacologia , Amidas/uso terapêutico , Antivirais/uso terapêutico , Túnica Conjuntiva/metabolismo , DNA Viral/genética , DNA Viral/isolamento & purificação , Diaminas/farmacologia , Diaminas/uso terapêutico , Dipeptídeos/farmacologia , Dipeptídeos/uso terapêutico , Terapia Genética/métodos , Vetores Genéticos/administração & dosagem , Vetores Genéticos/fisiologia , Células HeLa , Humanos , Receptores de Hialuronatos/metabolismo , Ácido Hialurônico/metabolismo , Ácidos Hidroxâmicos/farmacologia , Ácidos Hidroxâmicos/uso terapêutico , Indóis/farmacologia , Indóis/uso terapêutico , Maleimidas/farmacologia , Maleimidas/uso terapêutico , Metaloproteases/antagonistas & inibidores , Metaloproteases/metabolismo , Permeabilidade , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/metabolismo , Proteólise/efeitos dos fármacos , Piridinas/farmacologia , Piridinas/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Tiazóis/farmacologia , Tiazóis/uso terapêutico , Versicanas/metabolismo , Proteínas Virais/genética , Proteínas Virais/metabolismo , Corpo Vítreo/metabolismo , Quinases Associadas a rho/metabolismo
6.
Environ Health Prev Med ; 25(1): 30, 2020 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-32680455

RESUMO

Methylmercury is an environmental pollutant that causes neurotoxicity. Recent studies have reported that the ubiquitin-proteasome system is involved in defense against methylmercury toxicity through the degradation of proteins synthesizing the pyruvate. Mitochondrial accumulation of pyruvate can enhance methylmercury toxicity. In addition, methylmercury exposure induces several immune-related chemokines, specifically in the brain, and may cause neurotoxicity. This summary highlights several molecular mechanisms of methylmercury-induced neurotoxicity.


Assuntos
Quimiocinas/efeitos dos fármacos , Compostos de Metilmercúrio/toxicidade , Neurotoxinas/toxicidade , Proteólise/efeitos dos fármacos , Animais , Quimiocinas/metabolismo , Humanos , Camundongos , Ratos , Saccharomyces cerevisiae/efeitos dos fármacos
7.
PLoS One ; 15(6): e0232068, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32559187

RESUMO

Cyclin Dependent Kinase 9 (CDK9) associates with Bromodomain and Extra-Terminal Domain (BET) proteins to promote transcriptional elongation by phosphorylation of serine 2 of RNAP II C-terminal domain. We examined the therapeutic potential of selective CDK9 inhibitors (AZD 4573 and MC180295) against human multiple myeloma cells in vitro. Short-hairpin RNA silencing of CDK9 in Multiple Myeloma (MM) cell lines reduced cell viability compared to control cells showing the dependency of MM cells on CDK9. In order to explore synergy with the CDK9 inhibitor, proteolysis targeting chimeric molecule (PROTAC) ARV 825 was added. This latter drug causes ubiquitination of BET proteins resulting in their rapid and efficient degradation. Combination treatment of MM cells with ARV 825 and AZD 4573 markedly reduced their protein expression of BRD 2, BRD 4, MYC and phosphorylated RNA pol II as compared to each single agent alone. Combination treatment synergistically inhibited multiple myeloma cells both in vitro and in vivo with insignificant weight loss. The combination also resulted in marked increase of apoptotic cells at low dose compared to single agent alone. Taken together, our studies show for the first time that the combination of a BET PROTAC (ARV 825) plus AZD 4573 (CDK9 inhibitor) is effective against MM cells.


Assuntos
Quinase 9 Dependente de Ciclina/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Proteínas/metabolismo , Proteólise/efeitos dos fármacos , Animais , Azepinas/farmacologia , Azepinas/uso terapêutico , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Quinase 9 Dependente de Ciclina/genética , Quinase 9 Dependente de Ciclina/metabolismo , Regulação para Baixo/efeitos dos fármacos , Sinergismo Farmacológico , Humanos , Camundongos , Camundongos SCID , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/patologia , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas/genética , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Interferência de RNA , RNA Polimerase II/metabolismo , RNA Interferente Pequeno/metabolismo , Talidomida/análogos & derivados , Talidomida/farmacologia , Talidomida/uso terapêutico , Transplante Heterólogo
8.
Am J Physiol Renal Physiol ; 319(1): F84-F92, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32475130

RESUMO

Loss of muscle proteins increases the morbidity and mortality of patients with chronic kidney disease (CKD), and there are no reliable preventive treatments. We uncovered a STAT3/CCAAT-enhancer-binding protein-δ to myostatin signaling pathway that activates muscle protein degradation in mice with CKD or cancer; we also identified a small-molecule inhibitor of STAT3 (TTI-101) that blocks this pathway. To evaluate TTI-101 as a treatment for CKD-induced cachexia, we measured TTI-101 pharmacokinetics and pharmacodynamics in control and CKD rats that were orally administered TTI-101or its diluent. The following two groups of gavage-fed rats were studied: sham-operated control rats and CKD rats. Plasma was collected serially (0, 0.25, 0.5, 1, 2, 4, 8, and 24 h) following TTI-101 administration (at oral doses of 0, 10, 30, or 100 mg/kg). Plasma levels of TTI-101 were measured by LC-MS/MS, and pharmacokinetic results were analyzed with the PKSolver program. Plasma TTI-101 levels increased linearly with doses; the maximum plasma concentrations and time to maximal plasma levels (~1 h) were similar in sham-operated control rats and CKD rats. Notably, gavage treatment of TTI-101 for 3 days produced TTI-101 muscle levels in sham control rats and CKD rats that were not significantly different. CKD rats that received TTI-101 for 7 days had suppression of activated STAT3 and improved muscle grip strength; there also was a trend for increasing body and muscle weights. TTI-101 was tolerated at doses of 100 mg·kg-1·day-1 for 7 days. These results with TTI-101 in rats warrant its development as a treatment for cachexia in humans.


Assuntos
Inibidores Enzimáticos/farmacologia , Músculo Esquelético/efeitos dos fármacos , Proteólise/efeitos dos fármacos , Insuficiência Renal Crônica/metabolismo , Fator de Transcrição STAT3/antagonistas & inibidores , Animais , Peso Corporal/efeitos dos fármacos , Cromatografia Líquida , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacocinética , Força da Mão , Músculo Esquelético/metabolismo , Ratos , Espectrometria de Massas em Tandem
9.
Int J Food Microbiol ; 331: 108732, 2020 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-32521374

RESUMO

The present study aimed to evaluate the anti-quorum sensing (anti-QS) and anti-proteolytic potentials of tarragon essential oil (TEO) and its major compounds against food-associated Pseudomonas spp. The activities were verified by in vitro, in silico and in situ approaches. In this work, methyl eugenol (ME)- and ß-phellandrene (ß-PH)-rich TEO was investigated. TEO at subMIC increased the percentage of saturated fatty acids in the bacterial membranes (from 7 to 22%) and exhibited anti-quorum sensing via decreasing the efficiency of QS autoinducer synthesis [3-oxo-C12-HSL (from 2.028 µg/mL to

Assuntos
Artemisia/química , Óleos Voláteis/farmacologia , Proteólise/efeitos dos fármacos , Pseudomonas/efeitos dos fármacos , Percepção de Quorum/efeitos dos fármacos , 4-Butirolactona/análogos & derivados , 4-Butirolactona/isolamento & purificação , 4-Butirolactona/farmacologia , Animais , Antibacterianos/isolamento & purificação , Antibacterianos/farmacologia , Simulação por Computador , Peixes/microbiologia , Homosserina/análogos & derivados , Homosserina/isolamento & purificação , Homosserina/farmacologia , Óleos Voláteis/química , Óleos Voláteis/isolamento & purificação , Pseudomonas/isolamento & purificação
10.
Nat Struct Mol Biol ; 27(7): 605-614, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32541897

RESUMO

Controlled perturbation of protein activity is essential to study protein function in cells and living organisms. Small molecules that hijack the cellular protein ubiquitination machinery to selectively degrade proteins of interest, so-called degraders, have recently emerged as alternatives to selective chemical inhibitors, both as therapeutic modalities and as powerful research tools. These systems offer unprecedented temporal and spatial control over protein function. Here, we review recent developments in this field, with a particular focus on the use of degraders as research tools to interrogate complex biological problems.


Assuntos
Descoberta de Drogas/métodos , Fatores Imunológicos/farmacologia , Proteínas/metabolismo , Humanos , Fatores Imunológicos/química , Ácidos Indolacéticos/metabolismo , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Proteólise/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Sulfonamidas/química , Sulfonamidas/farmacologia , Ubiquitinação/efeitos dos fármacos
11.
Mol Pharmacol ; 98(3): 222-233, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32587095

RESUMO

DNA topoisomerase II (TOP2) is required for the unwinding and decatenation of DNA through the induction of an enzyme-linked double-strand break (DSB) in one DNA molecule and passage of another intact DNA duplex through the break. Anticancer drugs targeting TOP2 (TOP2 poisons) prevent religation of the DSB and stabilize a normally transient intermediate of the TOP2 reaction mechanism called the TOP2-DNA covalent complex. Subsequently, TOP2 remains covalently bound to each end of the enzyme-bridged DSB, which cannot be repaired until TOP2 is removed from the DNA. One removal mechanism involves the proteasomal degradation of the TOP2 protein, leading to the liberation of a protein-free DSB. Proteasomal degradation is often regulated by protein ubiquitination, and here we show that inhibition of ubiquitin-activating enzymes reduces the processing of TOP2A- and TOP2B-DNA complexes. Depletion or inhibition of ubiquitin-activating enzymes indicated that ubiquitination was required for the liberation of etoposide-induced protein-free DSBs and is therefore an important layer of regulation in the repair of TOP2 poison-induced DNA damage. TOP2-DNA complexes stabilized by etoposide were shown to be conjugated to ubiquitin, and this was reduced by inhibition or depletion of ubiquitin-activating enzymes. SIGNIFICANCE STATEMENT: There is currently great clinical interest in the ubiquitin-proteasome system and ongoing development of specific inhibitors. The results in this paper show that the therapeutic cytotoxicity of DNA topoisomerase II (TOP2) poisons can be enhanced through combination therapy with ubiquitin-activating enzyme inhibitors or by specific inhibition of the BMI/RING1A ubiquitin ligase, which would lead to increased cellular accumulation or persistence of TOP2-DNA complexes.


Assuntos
DNA Topoisomerases Tipo II/metabolismo , Nucleosídeos/farmacologia , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , Sulfonamidas/farmacologia , Enzimas Ativadoras de Ubiquitina/antagonistas & inibidores , Linhagem Celular , DNA/metabolismo , DNA Topoisomerases Tipo II/química , Humanos , Células K562 , Proteínas de Ligação a Poli-ADP-Ribose/química , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteólise/efeitos dos fármacos , Ubiquitina/metabolismo , Ubiquitinação/efeitos dos fármacos
12.
Toxicol Appl Pharmacol ; 401: 115092, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32512068

RESUMO

Inflammatory breast cancer (IBC) is a highly metastatic and lethal breast cancer. As many as 25-30% of IBCs are triple negative (TN) and associated with low survival rates and poor prognosis. We found that the microenvironment of IBC is characterized by high infiltration of tumor associated macrophages (TAMs) and by over-expression of the cysteine protease cathepsin B (CTSB). TAMs in IBC secrete high levels of the cytokines interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1/CCL2) compared to non-IBC patients. Herein, we tested the roles of IL-8 and MCP-1/CCL2 in modulating proteolytic activity and invasiveness of TN-non-IBC as compared to TN-IBC and addressed the underlying molecular mechanism(s) for both cytokines. Quantitative real time PCR results showed that IL-8 and MCP-1/CCL2 were significantly overexpressed in tissues of TN-IBCs. IL-8 and MCP-1/CCL2 induced CTSB expression and activity of the p-Src and p-Erk1/2 signaling pathways relevant for invasion and metastasis in TN-non-IBC, HCC70 cells and TN-IBC, SUM149 cells. Dasatinib, an inhibitor of p-Src, and U0126, an inhibitor of p-Erk1/2, down-regulated invasion and expression of CTSB by HCC70 and SUM149 cells, a mechanism that is reversed by IL-8 and MCP-1/CCL2. Our study shows that targeting the cytokines IL-8 and MCP-1/CCL2 and associated signaling molecules may represent a promising therapeutic strategy in TN-IBC patients.


Assuntos
Quimiocina CCL2/biossíntese , Genes src/fisiologia , Neoplasias Inflamatórias Mamárias/metabolismo , Interleucina-8/biossíntese , Sistema de Sinalização das MAP Quinases/fisiologia , Neoplasias de Mama Triplo Negativas/metabolismo , Adulto , Idoso , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Dasatinibe/farmacologia , Feminino , Genes src/efeitos dos fármacos , Humanos , Neoplasias Inflamatórias Mamárias/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Pessoa de Meia-Idade , Proteólise/efeitos dos fármacos , Neoplasias de Mama Triplo Negativas/patologia , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/fisiologia
13.
Life Sci ; 256: 117983, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32565252

RESUMO

Estrogen receptor (ER) positive accounts for a large proportion of breast cancer. Although there are many targeted therapeutic drugs, the emergence of drug resistance urgently requires the development of new drugs. Arctigenin (Arc), a lignan found in certain plants of the Asteraceae, has the effect on inhibiting breast cancer, but its molecular mechanism has not been clear. AIMS: To this end, the current study focuses on understanding the mechanism of Arc on ER-positive breast cancer cells. MAIN METHODS: Colony formation experiments and sulforhodamine B methods were used to determine the growth-inhibitory effect of Arc. The cell cycle and apoptosis were analyzed by flow cytometry. Alterations of signaling proteins were measured by Western blotting. Protein degradation was determined by comparing protein half-lives and inhibiting proteasome. KEY FINDINGS: The experimental results show that Arc did not induce apoptosis in ER-positive breast cancer cell, rather caused G1 cycle arrest by decreasing cyclin D1 levels without effect on altering CDK4/6 levels. Moreover, we have demonstrated that Arc decreases cyclin D1 levels through prompting Akt/GSK3ß-mediated degradation. SIGNIFICANCE: These findings warrant the potential of Arc as a candidate treatment for ER-positive breast cancer.


Assuntos
Neoplasias da Mama/metabolismo , Pontos de Checagem do Ciclo Celular/fisiologia , Ciclina D1/metabolismo , Furanos/farmacologia , Quinase 3 da Glicogênio Sintase/metabolismo , Lignanas/farmacologia , Receptores Estrogênicos/metabolismo , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Relação Dose-Resposta a Droga , Feminino , Furanos/uso terapêutico , Humanos , Lignanas/uso terapêutico , Células MCF-7 , Proteólise/efeitos dos fármacos
14.
Nature ; 585(7824): 293-297, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32494016

RESUMO

Molecular glue compounds induce protein-protein interactions that, in the context of a ubiquitin ligase, lead to protein degradation1. Unlike traditional enzyme inhibitors, these molecular glue degraders act substoichiometrically to catalyse the rapid depletion of previously inaccessible targets2. They are clinically effective and highly sought-after, but have thus far only been discovered serendipitously. Here, through systematically mining databases for correlations between the cytotoxicity of 4,518 clinical and preclinical small molecules and the expression levels of E3 ligase components across hundreds of human cancer cell lines3-5, we identify CR8-a cyclin-dependent kinase (CDK) inhibitor6-as a compound that acts as a molecular glue degrader. The CDK-bound form of CR8 has a solvent-exposed pyridyl moiety that induces the formation of a complex between CDK12-cyclin K and the CUL4 adaptor protein DDB1, bypassing the requirement for a substrate receptor and presenting cyclin K for ubiquitination and degradation. Our studies demonstrate that chemical alteration of surface-exposed moieties can confer gain-of-function glue properties to an inhibitor, and we propose this as a broader strategy through which target-binding molecules could be converted into molecular glues.


Assuntos
Ciclinas/deficiência , Ciclinas/metabolismo , Proteólise/efeitos dos fármacos , Purinas/química , Purinas/farmacologia , Piridinas/química , Piridinas/farmacologia , Linhagem Celular Tumoral , Quinases Ciclina-Dependentes/antagonistas & inibidores , Quinases Ciclina-Dependentes/química , Quinases Ciclina-Dependentes/metabolismo , Ciclinas/química , Proteínas de Ligação a DNA/metabolismo , Humanos , Modelos Moleculares , Complexo de Endopeptidases do Proteassoma/metabolismo , Ligação Proteica/efeitos dos fármacos , Purinas/toxicidade , Piridinas/toxicidade , Bibliotecas de Moléculas Pequenas/análise , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Ubiquitinação/efeitos dos fármacos
15.
Chem Biol Interact ; 325: 109088, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32360554

RESUMO

Osteoarthritis (OA) is one of the most common degenerative joint diseases in aging people. The activation of chondrocytes and their dysregulation are closely related to the pathogenesis of OA. GPR55 is an unique orphan G-receptor which binds to cannabinoids. In this study, we explored the role of GPR55 in advanced glycation end productions (AGEs)- induced chondrocytes activation in cultured cells. We showed that AGEs dose dependently induced GPR55 expression in ATDC5 chondrocytes. The blockage of GPR55 by its newly discovered antagonist-CID16020046 mitigated AGEs- induced increase in cellular ROS and decrease in antioxidant NRF2. Moreover, CID16020046 showed a dose-response suppressive effect on AGEs- induced expression of the major inflammatory mediators, including COX-2 and iNOS, and the production of NO and PGE2. CID16020046 also dose responsively inhibited AGEs- induced key effectors of cartilage degradation such as MMP-3 and MMP-13. In consequence, CID16020046 showed robust inhibition on AGEs- induced type II collagen degradation. Mechanistically, our data demonstrated that CID16020046 mediated GPR55 blockage ameliorated AGEs- induced NF-κB activation as revealed by its inhibition on IκBα, nuclear p65 translocation and NF-κB promoter activity. Collectively, our study demonstrates that GPR55 signaling mediates AGEs- induced chondrocyte activation, and the targeted blockage of GPR55 pathway could be therapeutic choice in the treatment of osteoarthritis.


Assuntos
Compostos Azabicíclicos/farmacologia , Benzoatos/farmacologia , Condrócitos/citologia , Condrócitos/efeitos dos fármacos , Produtos Finais de Glicação Avançada/farmacologia , Receptores de Canabinoides/metabolismo , Linhagem Celular , Condrócitos/metabolismo , Colágeno Tipo II/metabolismo , Ciclo-Oxigenase 2/genética , Dinoprostona/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Humanos , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 3 da Matriz/genética , NF-kappa B/metabolismo , Óxido Nítrico/biossíntese , Óxido Nítrico Sintase Tipo II/biossíntese , Estresse Oxidativo/efeitos dos fármacos , Proteólise/efeitos dos fármacos
16.
Proc Natl Acad Sci U S A ; 117(23): 13044-13055, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32434920

RESUMO

Epstein-Barr virus (EBV) is associated with epithelial and lymphoid malignancies, establishes latent infection in memory B cells, and intermittently produces infectious virions through lytic replication. Released virions play a key role in latent reservoir maintenance and transmission. Lytic EBV transcription differs from cellular transcription in requiring a virus-encoded preinitiation complex that binds to TATT motifs unique to EBV late lytic promoters. Expression of 15 late lytic genes that are important for virion production and infectivity is particularly dependent on the EBV SM protein, a nuclear protein expressed early during lytic reactivation that binds to viral RNAs and enhances RNA stability. We recently discovered that spironolactone blocks EBV virion production by inhibiting EBV SM function. Since spironolactone causes degradation of xeroderma pigmentosum group B-complementing protein (XPB), a component of human transcription factor TFIIH, in both B lymphocytes and epithelial cells, we hypothesized that SM utilizes XPB to specifically activate transcription of SM target promoters. While EBV SM has been thought to act posttranscriptionally, we provide evidence that SM also facilitates EBV gene transcription. We demonstrate that SM binds and recruits XPB to EBV promoters during lytic replication. Depletion of XPB protein, by spironolactone treatment or by siRNA transfection, inhibits SM-dependent late lytic gene transcription but not transcription of other EBV genes or cellular genes. These data indicate that SM acts as a transcriptional activator that has co-opted XPB to specifically target 15 EBV promoters that have uniquely evolved to require XPB for activity, providing an additional mechanism to differentially regulate EBV gene expression.


Assuntos
DNA Helicases/metabolismo , Proteínas de Ligação a DNA/metabolismo , Herpesvirus Humano 4/genética , Interações Hospedeiro-Patógeno/genética , Fosfoproteínas/metabolismo , Transativadores/metabolismo , Linhagem Celular Tumoral , DNA Helicases/genética , Proteínas de Ligação a DNA/genética , Infecções por Vírus Epstein-Barr/tratamento farmacológico , Infecções por Vírus Epstein-Barr/virologia , Regulação Viral da Expressão Gênica/efeitos dos fármacos , Humanos , Regiões Promotoras Genéticas/genética , Proteólise/efeitos dos fármacos , RNA Interferente Pequeno/metabolismo , Espironolactona/farmacologia , Espironolactona/uso terapêutico , Ativação Transcricional/efeitos dos fármacos , Vírion/efeitos dos fármacos , Vírion/metabolismo
17.
Food Chem ; 326: 127024, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32428856

RESUMO

This study aimed to analyze the effect of elderly gastrointestinal (GI) conditions on proteolysis, lipolysis and calcium and vitamins A and D3 bioaccessibility in salmon, sardine, sea bass and hake. For this purpose, cooked fishes were in vitro subjected to three elderly in vitro digestion models: E1 (oral elderly conditions), E2 (oral and gastric elderly conditions) and E3 (oral, gastric and intestinal elderly conditions)). In parallel, samples were digested under standardized GI conditions of a healthy adult as a control. Proteolysis was highly affected by elderly GI alterations (p < 0.05) (50% of reduction compared to control), being salmon and sea bass proteolysis extent (40 and 33%, respectively) the most affected with an important descend in leucine release. Calcium and vitamins bioaccessibility seemed to be also compromised for elders; however, the extent of the reduction highly depends on the fish type. Finally, these GI disorders did not negatively influence the bioabsorbable lipids of the fishes.


Assuntos
Cálcio/farmacologia , Trato Gastrointestinal/efeitos dos fármacos , Lipólise/efeitos dos fármacos , Proteólise/efeitos dos fármacos , Vitaminas/farmacologia , Idoso , Animais , Bass , Digestão , Gadiformes , Humanos , Salmão , Alimentos Marinhos/análise
18.
Mol Cell ; 78(6): 1086-1095, 2020 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-32407673

RESUMO

Transcription is epigenetically regulated by the orchestrated function of chromatin-binding proteins that tightly control the expression of master transcription factors, effectors, and supportive housekeeping genes required for establishing and propagating the normal and malignant cell state. Rapid advances in chemical biology and functional genomics have facilitated exploration of targeting epigenetic proteins, yielding effective strategies to target transcription while reducing toxicities to untransformed cells. Here, we review recent developments in conventional active site and allosteric inhibitors, peptidomimetics, and novel proteolysis-targeted chimera (PROTAC) technology that have deepened our understanding of transcriptional processes and led to promising preclinical compounds for therapeutic translation, particularly in cancer.


Assuntos
Epigênese Genética/efeitos dos fármacos , Epigênese Genética/genética , Neoplasias/genética , Animais , Antineoplásicos/farmacologia , Cromatina/genética , Cromatina/metabolismo , Epigênese Genética/fisiologia , Epigenômica/métodos , Humanos , Neoplasias/terapia , Proteólise/efeitos dos fármacos , Fatores de Transcrição/metabolismo
19.
Anticancer Res ; 40(4): 1989-1996, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32234888

RESUMO

BACKGROUND/AIM: The antitumor effect of sustained calcium supply on Src degradation was investigated in the context of hormone-dependent breast cancer, followed by elucidation of the underlying mechanisms. MATERIALS AND METHODS: Hormone-dependent T-47D breast cancer cells were used. Lactate calcium salt (LCS) was used as the source of sustained calcium supply, and the applicable concentration of LCS was determined by the colorimetric MTT assay. LCS-mediated deactivation of downstream signaling via Src degradation was identified by western blot and immunocytochemistry. RESULTS: Calcium-mediated degradation of Src decreased survival signaling via phosphoinositide 3-kinase and protein kinase B and resulted in significant inhibition of the clonogenic ability of hormone-dependent breast cancer cells. Tumor volume was significantly decreased in response to LCS injection in a heterotopic xenograft model, and immuno histochemistry revealed tumor necrosis. CONCLUSION: Sustained supply of calcium inhibited survival signaling via degradation of Src in hormone-dependent breast cancer cells.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Neoplasias Hormônio-Dependentes/tratamento farmacológico , Proteólise/efeitos dos fármacos , Quinases da Família src/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Cálcio/farmacologia , Linhagem Celular Tumoral , Feminino , Humanos , Ácido Láctico/farmacologia , Neoplasias Hormônio-Dependentes/genética , Neoplasias Hormônio-Dependentes/patologia , Proteína Oncogênica v-akt/genética , Fosfatidilinositol 3-Quinases/genética , Transdução de Sinais/efeitos dos fármacos , Quinases da Família src/antagonistas & inibidores
20.
Nat Commun ; 11(1): 1996, 2020 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-32332723

RESUMO

Small molecules that selectively kill senescent cells (SCs), termed senolytics, have the potential to prevent and treat various age-related diseases and extend healthspan. The use of Bcl-xl inhibitors as senolytics is largely limited by their on-target and dose-limiting platelet toxicity. Here, we report the use of proteolysis-targeting chimera (PROTAC) technology to reduce the platelet toxicity of navitoclax (also known as ABT263), a Bcl-2 and Bcl-xl dual inhibitor, by converting it into PZ15227 (PZ), a Bcl-xl PROTAC, which targets Bcl-xl to the cereblon (CRBN) E3 ligase for degradation. Compared to ABT263, PZ is less toxic to platelets, but equally or slightly more potent against SCs because CRBN is poorly expressed in platelets. PZ effectively clears SCs and rejuvenates tissue stem and progenitor cells in naturally aged mice without causing severe thrombocytopenia. With further improvement, Bcl-xl PROTACs have the potential to become safer and more potent senolytic agents than Bcl-xl inhibitors.


Assuntos
Envelhecimento/efeitos dos fármacos , Compostos de Anilina/farmacologia , Plaquetas/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Sulfonamidas/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Compostos de Anilina/química , Animais , Linhagem Celular , Feminino , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Modelos Animais , Cultura Primária de Células , Proteólise/efeitos dos fármacos , Sulfonamidas/química , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/metabolismo
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