RESUMO
PURPOSE: Hypereosinophilia represents a heterogenous group of severe medical conditions characterized by elevated numbers of eosinophil granulocytes in peripheral blood, bone marrow or tissue. Treatment options for hypereosinophilia remain limited despite recent approaches including IL-5-targeted monoclonal antibodies and tyrosine kinase inhibitors. METHODS: To understand aberrant survival patterns and options for pharmacologic intervention, we characterized BCL-2-regulated apoptosis signaling by testing for BCL-2 family expression levels as well as pharmacologic inhibition using primary patient samples from diverse subtypes of hypereosinophilia (hypereosinophilic syndrome n = 18, chronic eosinophilic leukemia not otherwise specified n = 9, lymphocyte-variant hypereosinophilia n = 2, myeloproliferative neoplasm with eosinophilia n = 2, eosinophilic granulomatosis with polyangiitis n = 11, reactive eosinophilia n = 3). RESULTS: Contrary to published literature, we found no difference in the levels of the lncRNA Morrbid and its target BIM. Yet, we identified a near complete loss of expression of pro-apoptotic PUMA as well as a reduction in anti-apoptotic BCL-2. Accordingly, BCL-2 inhibition using venetoclax failed to achieve cell death induction in eosinophil granulocytes and bone marrow mononuclear cells from patients with hypereosinophilia. In contrast, MCL1 inhibition using S63845 specifically decreased the viability of bone marrow progenitor cells in patients with hypereosinophilia. In patients diagnosed with Chronic Eosinophilic Leukemia (CEL-NOS) or Myeloid and Lymphatic Neoplasia with hypereosinophilia (MLN-Eo) repression of survival was specifically powerful. CONCLUSION: Our study shows that MCL1 inhibition might be a promising therapeutic option for hypereosinophilia patients specifically for CEL-NOS and MLN-Eo.
Assuntos
Eosinófilos/metabolismo , Síndrome Hipereosinofílica/genética , Síndrome Hipereosinofílica/terapia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos/uso terapêutico , Proteína 11 Semelhante a Bcl-2/fisiologia , Compostos Bicíclicos Heterocíclicos com Pontes/uso terapêutico , Estudos de Casos e Controles , Células Cultivadas , Eosinofilia/genética , Eosinofilia/mortalidade , Eosinofilia/patologia , Eosinofilia/terapia , Eosinófilos/patologia , Granulomatose com Poliangiite/genética , Granulomatose com Poliangiite/patologia , Granulomatose com Poliangiite/terapia , Células HL-60 , Humanos , Síndrome Hipereosinofílica/mortalidade , Síndrome Hipereosinofílica/patologia , Transtornos Mieloproliferativos/genética , Transtornos Mieloproliferativos/patologia , Transtornos Mieloproliferativos/terapia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Pirimidinas/uso terapêutico , Sulfonamidas/uso terapêutico , Tiofenos/uso terapêuticoRESUMO
While apoptosis plays a role in B-cell self-tolerance, its significance in preventing autoimmunity remains unclear. Here, we report that dysregulated B cell apoptosis leads to delayed onset autoimmune phenotype in mice. Our longitudinal studies revealed that mice with B cell-specific deletion of pro-apoptotic Bim (BBimfl/fl ) have an expanded B cell compartment with a notable increase in transitional, antibody secreting and recently described double negative (DN) B cells. They develop greater hypergammaglobulinemia than mice lacking Bim in all cells and accumulate several autoantibodies characteristic of Systemic Lupus Erythematosus (SLE) and related Sjögren's Syndrome (SS) including anti-nuclear, anti-Ro/SSA and anti-La/SSB at a level comparable to NODH2h4 autoimmune mouse model. Furthermore, lymphocytes infiltrated the tissues including submandibular glands and formed follicle-like structures populated with B cells, plasma cells and T follicular helper cells indicative of ongoing immune reaction. This autoimmunity was ameliorated upon deletion of Bruton's tyrosine kinase (Btk) gene, which encodes a key B cell signaling protein. These studies suggest that Bim-mediated apoptosis suppresses and B cell tyrosine kinase signaling promotes B cell-mediated autoimmunity.
Assuntos
Tirosina Quinase da Agamaglobulinemia/antagonistas & inibidores , Apoptose/fisiologia , Doenças Autoimunes/imunologia , Autoimunidade/imunologia , Linfócitos B/imunologia , Proteína 11 Semelhante a Bcl-2/fisiologia , Tirosina Quinase da Agamaglobulinemia/deficiência , Tirosina Quinase da Agamaglobulinemia/fisiologia , Animais , Especificidade de Anticorpos , Autoanticorpos/sangue , Linfócitos B/enzimologia , Linfócitos B/patologia , Proteína 11 Semelhante a Bcl-2/deficiência , Divisão Celular , Células Cultivadas , Hipergamaglobulinemia/imunologia , Tolerância Imunológica/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Antígenos/imunologia , Síndrome de Sjogren/imunologia , Linfócitos T/imunologiaRESUMO
MicroRNAs have become pivotal modulators in the pathogenesis of Alzheimer's disease. MiR-338-5p is associated with neuronal differentiation and neurogenesis, and expressed aberrantly in patients with cognitive dysfunction. However, its role and potential mechanism involved in Alzheimer's disease remain to be elucidated. Herein, we showed that the expression of miR-338-5p decreased in APP/PS1 mice, accompanied by the elevation in the expression level of amyloid ß, which indicated a reverse relationship between Alzheimer's disease progression and miR-338-5p. In addition, lentiviral overexpression of miR-338-5p through intrahippocampal injection mitigated the amyloid plaque deposition and cognitive dysfunction in APP/PS1 mice, suggesting a protecting role of miR-338-5p against the development of Alzheimer's disease. Moreover, miR-338-5p decelerated apoptotic loss of neurons in APP/PS1 mice. MiR-338-5p decreased neuronal apoptosis in vitro induced by amyloid ß accumulation, which was attributed to the negative regulation of BCL2L11 by miR-338-5p, since the restoration of BCL2L11 eliminated the protective role of miR-338-5p against neuronal apoptosis. Taken together, all of these results may indicate miR-338-5p as an innovative modulator in the pathogenesis of Alzheimer's disease, and also suggest that the protective effect of miR-338-5p on neuronal apoptosis may underlie its beneficial effect on APP/PS1 mice.
Assuntos
Doença de Alzheimer/etiologia , Apoptose/fisiologia , Proteína 11 Semelhante a Bcl-2/fisiologia , MicroRNAs/fisiologia , Neurônios/fisiologia , Animais , Células Cultivadas , Disfunção Cognitiva , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
JQ1 is a BET-bromodomain inhibitor that has immunomodulatory effects. However, the precise molecular mechanism that JQ1 targets to elicit changes in antibody production is not understood. Our results show that JQ1 induces apoptosis, reduces cell proliferation, and as a consequence, inhibits antibody-secreting cell differentiation. ChIP-sequencing reveals a selective displacement of Brd4 in response to acute JQ1 treatment (<2 h), resulting in specific transcriptional repression. After 8 h, subsequent alterations in gene expression arise as a result of the global loss of Brd4 occupancy. We demonstrate that apoptosis induced by JQ1 is solely attributed to the pro-apoptotic protein Bim (Bcl2l11). Conversely, cell-cycle regulation by JQ1 is associated with multiple Myc-associated gene targets. Our results demonstrate that JQ1 drives temporal changes in Brd4 displacement that results in a specific transcriptional profile that directly affects B cell survival and proliferation to modulate the humoral immune response.
Assuntos
Proteína 11 Semelhante a Bcl-2/metabolismo , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Animais , Apoptose/efeitos dos fármacos , Azepinas/farmacologia , Linfócitos B/metabolismo , Proteína 11 Semelhante a Bcl-2/fisiologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Nucleares/genética , Proteínas Nucleares/fisiologia , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/fisiologia , Triazóis/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
It has been widely accepted that mitochondria-dependent apoptosis initiates when select BH3-only proteins (BID, BIM, etc.) directly engage and allosterically activate effector proteins BAX/BAK. Here, through reconstitution of cells lacking all eight pro-apoptotic BH3-only proteins, we demonstrate that all BH3-only proteins primarily target the anti-apoptotic BCL-2 proteins BCL-xL/MCL-1, whose simultaneous suppression enables membrane-mediated spontaneous activation of BAX/BAK. BH3-only proteins' apoptotic activities correlate with affinities for BCL-xL/MCL-1 instead of abilities to directly activate BAX/BAK. Further, BID and BIM do not distinguish BAX from BAK or accelerate BAX/BAK activation following inactivation of BCL-xL/MCL-1. Remarkably, death ligand-induced apoptosis in cells lacking BH3-only proteins and MCL-1 is fully restored by BID mutants capable of neutralizing BCL-xL, but not direct activation of BAX/BAK. Taken together, our findings provide a "Membrane-mediated Permissive" model, in which the BH3-only proteins only indirectly activate BAX/BAK by neutralizing the anti-apoptotic BCL-2 proteins, and thus allowing BAX/BAK to undergo unimpeded, spontaneous activation in the mitochondrial outer membrane milieu, leading to apoptosis initiation.
Assuntos
Apoptose/fisiologia , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/fisiologia , Proteína 11 Semelhante a Bcl-2/fisiologia , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , Células HCT116 , Células HEK293 , Humanos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo , Proteína bcl-X/metabolismoRESUMO
High circulating insulin-like growth factor-1 (IGF-1) levels increase the risk of prostate cancer. However, whether circulating IGF-1 levels directly aggravate prostate cancer remains elusive. In this study, we crossed a transgenic prostate adenocarcinoma mouse model, Hi-Myc mice, with a liver-specific IGF-1 transgenic mouse model (HIT) to increase their circulating IGF-1 levels to investigate the impact of the elevated circulating IGF-1 on prostate cancer development in vivo. The Hi-Myc/HIT mice had increased incidence and invasiveness of prostate cancer. IGF-1 elevation led to the accumulation of FOXO3A in the cytosol of prostate tumor cells and downregulation of its target gene Bim, which resulted in the apoptosis inhibition and prostate cancer overgrowth. The differential expressions of IGF-1R, FOXO3A, and BIM in the benign versus malignant prostate tissues supported a negative association between the FOXO3A/BIM axis and IGF-1R expression in human prostate adenocarcinoma. Our findings suggest that targeting the IGF-1/FOXO3A/BIM signaling axis could be an attractive strategy for prostate cancer prevention or treatment.
Assuntos
Adenocarcinoma , Proteína 11 Semelhante a Bcl-2/fisiologia , Proteína Forkhead Box O3/fisiologia , Fator de Crescimento Insulin-Like I/fisiologia , Neoplasias da Próstata , Adenocarcinoma/sangue , Adenocarcinoma/genética , Adenocarcinoma/patologia , Animais , Proteína 11 Semelhante a Bcl-2/genética , Linhagem Celular Tumoral , Estudos de Coortes , Modelos Animais de Doenças , Progressão da Doença , Proteína Forkhead Box O3/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Fator de Crescimento Insulin-Like I/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Camundongos Transgênicos , Células PC-3 , Neoplasias da Próstata/sangue , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Transdução de Sinais/genéticaRESUMO
Estrogen dependence is major driver of ER + breast cancer, which is associated with PI3K mutation. PI3K inhibition (PI3Ki) can restore dependence on ER signaling for some hormone therapy-resistant ER + breast cancers, but is ineffective in others. Here we show that short-term supplementation with estrogen strongly enhanced Pik3caH1047R-induced mammary tumorigenesis in mice that resulted exclusively in ER + tumors, demonstrating the cooperation of the hormone and the oncogene in tumor development. Similar to human ER + breast cancers that are endocrine-dependent or endocrine-independent at diagnosis, tumor lines from this model retained ER expression but were sensitive or resistant to hormonal therapies. PI3Ki did not induce cell death but did cause upregulation of the pro-apoptotic gene BIM. BH3 mimetics or PI3Ki were unable to restore hormone sensitivity in several resistant mouse and human tumor lines. Importantly however, combination of PI3Ki and BH3 mimetics had a profound, BIM-dependent cytotoxic effect in PIK3CA-mutant cancer cells while sparing normal cells. We propose that addition of BH3 mimetics offers a therapeutic strategy to markedly improve the cytotoxic activity of PI3Ki in hormonal therapy-resistant and ER-independent PIK3CA-mutant breast cancer.
Assuntos
Compostos de Anilina/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/antagonistas & inibidores , Proteína 11 Semelhante a Bcl-2/agonistas , Estradiol , Receptor alfa de Estrogênio/fisiologia , Neoplasias Mamárias Experimentais/tratamento farmacológico , Proteínas de Neoplasias/fisiologia , Neoplasias Hormônio-Dependentes/tratamento farmacológico , Neuropeptídeos/antagonistas & inibidores , Inibidores de Fosfoinositídeo-3 Quinase , Sulfonamidas/farmacologia , Tiazóis/farmacologia , Compostos de Anilina/administração & dosagem , Animais , Antineoplásicos Hormonais/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Proteína 11 Semelhante a Bcl-2/biossíntese , Proteína 11 Semelhante a Bcl-2/genética , Proteína 11 Semelhante a Bcl-2/fisiologia , Linhagem Celular Tumoral , Classe I de Fosfatidilinositol 3-Quinases , Cocarcinogênese , Resistencia a Medicamentos Antineoplásicos , Ensaios de Seleção de Medicamentos Antitumorais , Sinergismo Farmacológico , Estradiol/toxicidade , Receptor alfa de Estrogênio/efeitos dos fármacos , Feminino , Fulvestranto/administração & dosagem , Fulvestranto/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Técnicas de Introdução de Genes , Neoplasias Mamárias Experimentais/induzido quimicamente , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Nus , Mutação de Sentido Incorreto , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Neoplasias Hormônio-Dependentes/induzido quimicamente , Neoplasias Hormônio-Dependentes/genética , Neoplasias Hormônio-Dependentes/patologia , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/fisiologia , Sulfonamidas/administração & dosagem , Tiazóis/administração & dosagemRESUMO
Metastatic breast cancer is responsible for most breast cancer-related deaths. Disseminated cancer cells have developed an intrinsic ability to resist anchorage-dependent apoptosis (anoikis). Anoikis is caused by the absence of cellular adhesion, a process that underpins lumen formation and maintenance during mammary gland development and homeostasis. In healthy cells, anoikis is mostly governed by B-cell lymphoma-2 (BCL2) protein family members. Metastatic cancer cells, however, have often developed autocrine BCL2-dependent resistance mechanisms to counteract anoikis. In this Review, we discuss how a pro-apoptotic subgroup of the BCL2 protein family, known as the BH3-only proteins, controls apoptosis and anoikis during mammary gland homeostasis and to what extent their inhibition confers tumor suppressive functions in metastatic breast cancer. Specifically, the role of the two pro-apoptotic BH3-only proteins BCL2-modifying factor (BMF) and BCL2-interacting mediator of cell death (BIM) will be discussed here. We assess current developments in treatment that focus on mimicking the function of the BH3-only proteins to induce apoptosis, and consider their applicability to restore normal apoptotic responses in anchorage-independent disseminating tumor cells.
Assuntos
Proteínas Reguladoras de Apoptose/fisiologia , Neoplasias da Mama/terapia , Morte Celular/fisiologia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Animais , Proteína 11 Semelhante a Bcl-2/fisiologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Feminino , Humanos , Terapia de Alvo Molecular/métodos , Terapia de Alvo Molecular/tendências , Metástase NeoplásicaRESUMO
Phosphorylation of Ser/Thr residues is a well-established modulating mechanism of the pro-apoptotic function of the BH3-only protein Bim. However, nothing is known about the putative tyrosine phosphorylation of this Bcl-2 family member and its potential impact on Bim function and subsequent Bax/Bak-mediated cytochrome c release and apoptosis. As we have previously shown that the tyrosine kinase Lyn could behave as an anti-apoptotic molecule, we investigated whether this Src family member could directly regulate the pro-apoptotic function of Bim. In the present study, we show that Bim is phosphorylated onto tyrosine residues 92 and 161 by Lyn, which results in an inhibition of its pro-apoptotic function. Mechanistically, we show that Lyn-dependent tyrosine phosphorylation of Bim increases its interaction with anti-apoptotic members such as Bcl-xL, therefore limiting mitochondrial outer membrane permeabilization and subsequent apoptosis. Collectively, our data uncover one molecular mechanism through which the oncogenic tyrosine kinase Lyn negatively regulates the mitochondrial apoptotic pathway, which may contribute to the transformation and/or the chemotherapeutic resistance of cancer cells.
Assuntos
Apoptose/genética , Proteína 11 Semelhante a Bcl-2/fisiologia , Quinases da Família src/fisiologia , Animais , Proteína 11 Semelhante a Bcl-2/antagonistas & inibidores , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Células Cultivadas , Resistencia a Medicamentos Antineoplásicos/genética , Células HEK293 , Células HeLa , Humanos , Células K562 , Camundongos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Oncogenes/fisiologia , Transdução de Sinais/genética , Quinases da Família src/genéticaRESUMO
BCL-2 proteins are known to engage each other to determine the fate of a cell after a death stimulus. However, their evolutionary conservation and the many other reported binding partners suggest an additional function not directly linked to apoptosis regulation. To identify such a function, we studied mice lacking the BH3-only protein BIM. BIM-/- cells had a higher mitochondrial oxygen consumption rate that was associated with higher mitochondrial complex IV activity. The consequences of increased oxygen consumption in BIM-/- mice were significantly lower body weights, reduced adiposity and lower hepatic lipid content. Consistent with reduced adiposity, BIM-/- mice had lower fasting blood glucose, improved insulin sensitivity and hepatic insulin signalling. Lipid oxidation was increased in BIM-/- mice, suggesting a mechanism for their metabolic phenotype. Our data suggest a role for BIM in regulating mitochondrial bioenergetics and metabolism and support the idea that regulation of metabolism and cell death are connected.
Assuntos
Adiposidade , Proteína 11 Semelhante a Bcl-2/fisiologia , Metabolismo dos Lipídeos , Mitocôndrias/metabolismo , Animais , Proteína 11 Semelhante a Bcl-2/genética , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Metabolismo Energético , Glucose/metabolismo , Hepatócitos/metabolismo , Resistência à Insulina , Fígado/metabolismo , Potencial da Membrana Mitocondrial , Camundongos , Oxirredução , Consumo de Oxigênio , Redução de PesoRESUMO
The members of the BCL-2 family are crucial regulators of the mitochondrial pathway of apoptosis in normal physiology and disease. Besides their role in cell death, BCL-2 proteins have been implicated in the regulation of mitochondrial oxidative phosphorylation and cellular metabolism. It remains unclear, however, whether these proteins have a physiological role in glucose homeostasis and metabolism in vivo. In this study, we report that fat accumulation in the liver increases c-Jun N-terminal kinase-dependent BCL-2 interacting mediator of cell death (BIM) expression in hepatocytes. To determine the consequences of hepatic BIM deficiency in diet-induced obesity, we generated liver-specific BIM-knockout (BLKO) mice. BLKO mice had lower hepatic lipid content, increased insulin signaling, and improved global glucose metabolism. Consistent with these findings, lipogenic and lipid uptake genes were downregulated and lipid oxidation enhanced in obese BLKO mice. Mechanistically, BIM deficiency improved mitochondrial function and decreased oxidative stress and oxidation of protein tyrosine phosphatases, and ameliorated activation of peroxisome proliferator-activated receptor γ/sterol regulatory element-binding protein 1/CD36 in hepatocytes from high fat-fed mice. Importantly, short-term knockdown of BIM rescued obese mice from insulin resistance, evidenced by reduced fat accumulation and improved insulin sensitivity. Our data indicate that BIM is an important regulator of liver dysfunction in obesity and a novel therapeutic target for restoring hepatocyte function.
Assuntos
Proteína 11 Semelhante a Bcl-2/fisiologia , Fígado Gorduroso/etiologia , Resistência à Insulina , Proteínas Quinases JNK Ativadas por Mitógeno/fisiologia , Fígado/metabolismo , Obesidade/metabolismo , Estresse Oxidativo , Animais , Células Cultivadas , Ativação Enzimática , Ácidos Graxos/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio/metabolismoRESUMO
OBJECTIVE: Parkinson is one of the most common neurodegenerative diseases. At present, many studies have pointed out that miRNAs play a very important role in Parkinson's development and process. MiR-181c has been shown to have a significant low expression in blood samples and brain tissues of Parkinson's patients. MATERIALS AND METHODS: We used 1-Methyl-4-Phenylpyridinium Iodide (MPP(+)) as a tool for constructing the Parkinson's cell model, using mir181c mimics to construct an experimental model of acquisition. The cell viability of PC12 was detected by MTT and CCK8. Reactive oxygen species (ROS) and caspase-3 activity were analyzed. The apoptosis of PC12 was detected by flow cytometry (FCM), and luciferase was used to study the binding of target genes. The protein levels of BCL2L11were measured by Western-blot. RESULTS: There was a significant low expression of mir181c in MPP(+)-morbid cells. PC12 cell viability was rescued by miR-181c overexpression. Flow cytometry showed that apoptosis in PC12 cells overexpressing miR-181c was significantly decreased. Also, ROS and caspase-3 activity were significantly decreased. Luciferase experiments showed that miR-181c may bind to the 3-'UTR side of BCL2L11 and inhibited its expression. By Western-blot, the BCL2L11 level was markedly decreased by miR-181c. CONCLUSIONS: miR-181c could promote the cell viability and inhibit the apoptosis of PC12 cells induced by MPP (+) by downregulating BCL2L11, which may play a protective role and provide a new target for PD drug resistance research.
Assuntos
1-Metil-4-fenilpiridínio/toxicidade , Proteína 11 Semelhante a Bcl-2/fisiologia , MicroRNAs/fisiologia , Doença de Parkinson/prevenção & controle , Animais , Apoptose , Sobrevivência Celular , Células PC12 , Fatores de Proteção , Ratos , Espécies Reativas de Oxigênio/metabolismoRESUMO
A deletion polymorphism in the BIM gene was identified as an intrinsic mechanism for resistance to tyrosine kinase inhibitor in chronic myeloid leukemia patients in East Asia. BIM is also involved in the responses to glucocorticoid and chemotherapy in acute lymphoblastic leukemia (ALL), suggesting a possible association between deletion polymorphism of BIM and the chemosensitivity of ALL. Thus, we analyzed 72 B-cell precursor (BCP)-ALL cell lines established from Japanese patients. Indeed, higher BIM gene expression was associated with good in vitro sensitivities to glucocorticoid and chemotherapeutic agents used in induction therapy. We also analyzed the methylation status of the BIM gene promoter by next generation sequencing of genome bisulfite PCR products, since genetic polymorphism could be insignificant when epigenetically inactivated. Hypermethylation of the BIM gene promoter was associated with lower BIM gene expression and poorer sensitivity to vincristine. Of note, however, the prevalence of a deletion polymorphism was not associated with the BIM gene expression level or drug sensitivities in BCP-ALL cell lines, in which the BIM gene was unmethylated. These observations suggest that an association of a deletion polymorphism of BIM and the response to induction therapy in BCP-ALL may be clinically minimal.
Assuntos
Proteína 11 Semelhante a Bcl-2/genética , Resistencia a Medicamentos Antineoplásicos/genética , Deleção de Genes , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Antineoplásicos/farmacologia , Povo Asiático , Proteína 11 Semelhante a Bcl-2/fisiologia , Linhagem Celular Tumoral , Metilação de DNA , Glucocorticoides/farmacologia , Humanos , Polimorfismo Genético , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologia , Regiões Promotoras Genéticas , Vincristina/farmacologiaRESUMO
Mycolactone, the macrolide exotoxin produced by Mycobacterium ulcerans, is central to the pathogenesis of the chronic necrotizing skin disease Buruli ulcer (BU). Here we show that mycolactone acts as an inhibitor of the mechanistic Target of Rapamycin (mTOR) signaling pathway by interfering with the assembly of the two distinct mTOR protein complexes mTORC1 and mTORC2, which regulate different cellular processes. Inhibition of the assembly of the rictor containing mTORC2 complex by mycolactone prevents phosphorylation of the serine/threonine protein kinase Akt. The associated inactivation of Akt leads to the dephosphorylation and activation of the Akt-targeted transcription factor FoxO3. Subsequent up-regulation of the FoxO3 target gene BCL2L11 (Bim) increases expression of the pro-apoptotic regulator Bim, driving mycolactone treated mammalian cells into apoptosis. The central role of Bim-dependent apoptosis in BU pathogenesis deduced from our experiments with cultured mammalian cells was further verified in an experimental M. ulcerans infection model. As predicted by the model, M. ulcerans infected Bim knockout mice did not develop necrotic BU lesions with large clusters of extracellular bacteria, but were able to contain the mycobacterial multiplication. Our findings provide a new coherent and comprehensive concept of BU pathogenesis.
Assuntos
Apoptose , Proteína 11 Semelhante a Bcl-2/fisiologia , Úlcera de Buruli/patologia , Macrolídeos/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Animais , Proteína 11 Semelhante a Bcl-2/genética , Proteína 11 Semelhante a Bcl-2/metabolismo , Úlcera de Buruli/microbiologia , Células Cultivadas , Técnicas de Inativação de Genes , Macrolídeos/toxicidade , Alvo Mecanístico do Complexo 1 de Rapamicina , Alvo Mecanístico do Complexo 2 de Rapamicina , Camundongos , Complexos Multiproteicos/efeitos dos fármacos , Mycobacterium ulcerans/química , Mycobacterium ulcerans/patogenicidade , Serina-Treonina Quinases TOR/efeitos dos fármacosRESUMO
Natural killer (NK) cells are innate lymphoid cells with antitumor functions. Using an N-ethyl-N-nitrosourea (ENU)-induced mutagenesis screen in mice, we identified a strain with an NK cell deficiency caused by a hypomorphic mutation in the Bcl2 (B cell lymphoma 2) gene. Analysis of these mice and the conditional deletion of Bcl2 in NK cells revealed a nonredundant intrinsic requirement for BCL2 in NK cell survival. In these mice, NK cells in cycle were protected against apoptosis, and NK cell counts were restored in inflammatory conditions, suggesting a redundant role for BCL2 in proliferating NK cells. Consistent with this, cycling NK cells expressed higher MCL1 (myeloid cell leukemia 1) levels in both control and BCL2-null mice. Finally, we showed that deletion of BIM restored survival in BCL2-deficient but not MCL1-deficient NK cells. Overall, these data demonstrate an essential role for the binding of BCL2 to BIM in the survival of noncycling NK cells. They also favor a model in which MCL1 is the dominant survival protein in proliferating NK cells.
Assuntos
Células Matadoras Naturais/fisiologia , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Animais , Antígenos Ly/fisiologia , Proteína 11 Semelhante a Bcl-2/fisiologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Ciclo Celular , Sobrevivência Celular , Feminino , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptor 1 Desencadeador da Citotoxicidade Natural/fisiologia , Sulfonamidas/farmacologiaRESUMO
3,5-Dimethyl-7H-furo[3,2-g]chromen-7-one (DMFC) is a coumarin derivative with anti-cancer activity against human hepatoma cells, but the mechanisms underlying DMFC function in cancer suppression is unknown. In this study, we aimed at elucidating the molecular mechanisms underlying DMFC anti-cancer activity and determining whether DMFC is effective in suppression of drug-resistant human hepatocellular carcinoma. We show here that DMFC effectively suppresses both the parent and the multidrug-resistant hepatoma cell growth in vitro and DMFC suppresses hepatoma cell growth at least in part through inducing tumor cell apoptosis. In the molecular level, we observed that DMFC treatment decreases Bcl-2 level by a post-transcriptional mechanism and activates Bim transcription to increase Bim mRNA and protein level in hepatoma cells. Furthermore, co-immunoprecipitation studies revealed that DMFC-induced Bim interrupts interactions between Bcl-2 and Bax and between Mcl-1 and Bak, resulting in dissociation of Bax from Bcl-2 and Bak from Mcl-1 and subsequent activation of both Bax and Bak. Activation of Bax and Bak leads to mitochondrial outer membrane permeabilization and cytochrome c release. Consistent with its potent apoptosis-inducing activity, DMFC exhibited potent activity against the multidrug-resistant hepatoma xenograft growth in vivo. Therefore, we determine that DMFC suppresses hepatoma growth through decreasing Bcl-2 and increasing Bim to induce tumor cell apoptosis and hold great promise for further development as a therapeutic agent to treat chemoresistant hepatoma.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Proteína 11 Semelhante a Bcl-2/fisiologia , Benzofuranos/farmacologia , Carcinoma Hepatocelular/metabolismo , Cumarínicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias Hepáticas/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteína Killer-Antagonista Homóloga a bcl-2/metabolismo , Proteína X Associada a bcl-2/metabolismo , Animais , Antineoplásicos/química , Proteína 11 Semelhante a Bcl-2/antagonistas & inibidores , Proteína 11 Semelhante a Bcl-2/biossíntese , Proteína 11 Semelhante a Bcl-2/genética , Benzofuranos/química , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Cumarínicos/química , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistência a Múltiplos Medicamentos/fisiologia , Resistencia a Medicamentos Antineoplásicos/fisiologia , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Nus , Estrutura Molecular , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Conformação Proteica/efeitos dos fármacos , Interferência de RNA , Transcrição Gênica/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The BET (bromodomain and extraterminal domain) bromodomain-containing proteins, such as BRD4, are highly promising targets for treating lymphoid and myeloid malignancies. They act to modulate the expression of multiple genes that control diverse cellular processes including proliferation, survival and differentiation that are consequentially disrupted by small-molecule BET bromodomain inhibitors such as JQ1. By assessing the impact of these inhibitors on normal mouse hematopoietic cells or their transformed counterparts, we establish definitively that their cytotoxic action in vitro and in vivo relies predominantly on the activation of BAX/BAK-dependent mitochondrial (intrinsic) apoptosis. In large part, this is triggered by marked upregulation of the BH3-only protein BIM when the BET inhibitors suppress miR-17-92, a key post-transcriptional repressor of BIM expression. Thus, our study strongly suggests that mutations that permit the evasion of apoptosis (for example, BCL2 overexpression, BIM inactivation) are likely to blunt the activity of the BET bromodomain inhibitors and should be anticipated when therapy resistance develops. Strikingly, we also found that certain normal hematopoietic cells, especially those of lymphoid origin, are as prone to apoptosis induced by the BET inhibitors as their transformed counterparts, indicating that their susceptibility to BET inhibitors did not arise from oncogenic transformation.
Assuntos
Apoptose , Azepinas/farmacologia , Proteína 11 Semelhante a Bcl-2/fisiologia , Linfoma/patologia , MicroRNAs/antagonistas & inibidores , Fatores de Transcrição/antagonistas & inibidores , Triazóis/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Proteínas de Ciclo Celular , Linhagem Celular , Linhagem Celular Transformada , Modelos Animais de Doenças , Sistema Hematopoético/citologia , História Antiga , Humanos , Camundongos , Camundongos Transgênicos , Proteínas Nucleares/antagonistas & inibidores , RNA Longo não CodificanteRESUMO
BACKGROUND & AIMS: Acute hepatitis is often mediated by cytotoxic T lymphocytes (CTLs); however, the intrinsic parameters that limit CTL-mediated liver injury are not well understood. METHODS: To investigate whether acute liver damage is limited by molecules that decrease the lifespan or effector function of CTLs, we used a well-characterized transgenic (Tg) mouse model in which acute liver damage develops upon transfer of T cell receptor (TCR) Tg CD8 T cells. Recipient Tg mice received donor TCR Tg T cells deficient for either the pro-apoptotic molecule Bim, which regulates CTL survival, or suppressor of cytokine signaling-1 (SOCS-1), which controls expression of common gamma chain cytokines; the effects of anti-PD-L1 neutralizing antibodies were also assessed. RESULTS: Use of Bim-deficient donor T cells and/or PD-L1 blockade increased the number of intrahepatic T cells without affecting the degree and kinetic of acute hepatitis. In contrast, SOCS-1-deficient T cells induced a heightened, prolonged acute hepatitis caused by their enhanced cytotoxic function and increased expansion. Although they inflicted more severe acute liver damage, SOCS-1-deficient T cells never precipitated chronic hepatitis and became exhausted. CONCLUSIONS: The degree of acute hepatitis is regulated by the function of CD8 T cells, but is not affected by changes in CTL lifespan. Although manipulation of the examined parameters affected acute hepatitis, persistent hepatitis did not ensue, indicating that, in the presence of high intrahepatic antigen load, changes in these factors in isolation were not sufficient to prevent T cell exhaustion and mediate progression to chronic hepatitis.
Assuntos
Hepatite/etiologia , Linfócitos T Citotóxicos/imunologia , Doença Aguda , Animais , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/fisiologia , Proteína 11 Semelhante a Bcl-2/fisiologia , Sobrevivência Celular , Hepatite/imunologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/fisiologia , Receptores de Antígenos de Linfócitos T/fisiologia , Proteína 1 Supressora da Sinalização de Citocina/fisiologia , Linfócitos T Citotóxicos/fisiologiaRESUMO
Caffeine is one of the most commonly ingested neuroactive compounds and exhibits anticancer effects through induction of apoptosis and suppression of cell proliferation. However, the mechanisms underlying these effects are currently unknown. In this study, we investigated the mechanisms of caffeine-induced apoptosis in U251 cells (human glioma cell line). We analyzed the inhibitory effects of caffeine on cell proliferation by performing WST-8 and colony formation assays; in addition, cell survival was assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and flow cytometric analysis. Western blotting was used to investigate the role played by FoxO1 in the proapoptotic effects of caffeine on glioma cells. Results showed that caffeine inhibited proliferation and survival of human glioma cells, induced apoptosis, and increased the expression of FoxO1 and its proapoptotic target Bim. In addition, we found that FoxO1 enhanced the transcription of its proapoptotic target Bim. In summary, our data indicates that FoxO1-Bim mediates caffeine-induced regression of glioma growth by activating cell apoptosis, thereby providing new mechanistic insight into the possible use of caffeine in treating human cancer.