RESUMEN
Proteins of the bromodomain and extra-terminal (BET) domain family are epigenetic readers that bind acetylated histones through their bromodomains to regulate gene transcription. Dual-bromodomain BET inhibitors (DbBi) that bind with similar affinities to the first (BD1) and second (BD2) bromodomains of BRD2, BRD3, BRD4 and BRDt have displayed modest clinical activity in monotherapy cancer trials. A reduced number of thrombocytes in the blood (thrombocytopenia) as well as symptoms of gastrointestinal toxicity are dose-limiting adverse events for some types of DbBi1-5. Given that similar haematological and gastrointestinal defects were observed after genetic silencing of Brd4 in mice6, the platelet and gastrointestinal toxicities may represent on-target activities associated with BET inhibition. The two individual bromodomains in BET family proteins may have distinct functions7-9 and different cellular phenotypes after pharmacological inhibition of one or both bromodomains have been reported10,11, suggesting that selectively targeting one of the bromodomains may result in a different efficacy and tolerability profile compared with DbBi. Available compounds that are selective to individual domains lack sufficient potency and the pharmacokinetics properties that are required for in vivo efficacy and tolerability assessment10-13. Here we carried out a medicinal chemistry campaign that led to the discovery of ABBV-744, a highly potent and selective inhibitor of the BD2 domain of BET family proteins with drug-like properties. In contrast to the broad range of cell growth inhibition induced by DbBi, the antiproliferative activity of ABBV-744 was largely, but not exclusively, restricted to cell lines of acute myeloid leukaemia and prostate cancer that expressed the full-length androgen receptor (AR). ABBV-744 retained robust activity in prostate cancer xenografts, and showed fewer platelet and gastrointestinal toxicities than the DbBi ABBV-07514. Analyses of RNA expression and chromatin immunoprecipitation followed by sequencing revealed that ABBV-744 displaced BRD4 from AR-containing super-enhancers and inhibited AR-dependent transcription, with less impact on global transcription compared with ABBV-075. These results underscore the potential value of selectively targeting the BD2 domain of BET family proteins for cancer therapy.
Asunto(s)
Proteínas de Ciclo Celular/antagonistas & inhibidores , Proteínas de Ciclo Celular/química , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/metabolismo , Dominios Proteicos/efectos de los fármacos , Piridinas/farmacología , Pirroles/farmacología , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/química , Animales , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Elementos de Facilitación Genéticos/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Masculino , Ratones , Piridinas/efectos adversos , Piridinas/toxicidad , Pirroles/efectos adversos , Pirroles/toxicidad , Ratas , Receptores Androgénicos/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Transcripción Genética/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
In the originally published version of this Letter, the authors Arthur F. Kluge, Michael A. Patane and Ce Wang were inadvertently omitted from the author list. Their affiliations are: I-to-D, Inc., PO Box 6177, Lincoln, Massachusetts 01773, USA (A.F.K.); Mitobridge, Inc. 1030 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA (M.A.P.); and China Novartis Institutes for BioMedical Research, No. 4218 Jinke Road, Zhangjiang Hi-Tech Park, Pudong District, Shanghai 201203, China (C.W.). These authors contributed to the interpretation of results and design of compounds. In addition, author 'Edward A. Kesicki' was misspelled as 'Ed Kesicki'. These errors have been corrected online.
RESUMEN
The dynamic and reversible acetylation of proteins, catalysed by histone acetyltransferases (HATs) and histone deacetylases (HDACs), is a major epigenetic regulatory mechanism of gene transcription and is associated with multiple diseases. Histone deacetylase inhibitors are currently approved to treat certain cancers, but progress on the development of drug-like histone actyltransferase inhibitors has lagged behind. The histone acetyltransferase paralogues p300 and CREB-binding protein (CBP) are key transcriptional co-activators that are essential for a multitude of cellular processes, and have also been implicated in human pathological conditions (including cancer). Current inhibitors of the p300 and CBP histone acetyltransferase domains, including natural products, bi-substrate analogues and the widely used small molecule C646, lack potency or selectivity. Here, we describe A-485, a potent, selective and drug-like catalytic inhibitor of p300 and CBP. We present a high resolution (1.95 Å) co-crystal structure of a small molecule bound to the catalytic active site of p300 and demonstrate that A-485 competes with acetyl coenzyme A (acetyl-CoA). A-485 selectively inhibited proliferation in lineage-specific tumour types, including several haematological malignancies and androgen receptor-positive prostate cancer. A-485 inhibited the androgen receptor transcriptional program in both androgen-sensitive and castration-resistant prostate cancer and inhibited tumour growth in a castration-resistant xenograft model. These results demonstrate the feasibility of using small molecule inhibitors to selectively target the catalytic activity of histone acetyltransferases, which may provide effective treatments for transcriptional activator-driven malignancies and diseases.
Asunto(s)
Linaje de la Célula , Compuestos Heterocíclicos de 4 o más Anillos/farmacología , Compuestos Heterocíclicos de 4 o más Anillos/uso terapéutico , Histona Acetiltransferasas/antagonistas & inhibidores , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Factores de Transcripción p300-CBP/antagonistas & inhibidores , Acetilcoenzima A/metabolismo , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Unión Competitiva , Biocatálisis/efectos de los fármacos , Dominio Catalítico/efectos de los fármacos , Línea Celular Tumoral , Linaje de la Célula/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Neoplasias Hematológicas/tratamiento farmacológico , Neoplasias Hematológicas/enzimología , Neoplasias Hematológicas/patología , Compuestos Heterocíclicos de 4 o más Anillos/química , Histona Acetiltransferasas/química , Histona Acetiltransferasas/metabolismo , Humanos , Masculino , Ratones , Ratones SCID , Modelos Moleculares , Neoplasias/enzimología , Neoplasias de la Próstata Resistentes a la Castración/tratamiento farmacológico , Neoplasias de la Próstata Resistentes a la Castración/enzimología , Neoplasias de la Próstata Resistentes a la Castración/patología , Conformación Proteica , Receptores Androgénicos/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto , Factores de Transcripción p300-CBP/química , Factores de Transcripción p300-CBP/metabolismoRESUMEN
BACKGROUND: Venetoclax (ABT-199), a first-in-class orally bioavailable BCL-2-selective inhibitor, was recently approved by the FDA for use in patients with 17p-deleted chronic lymphocytic leukemia who have received prior therapy. It is also being evaluated in numerous clinical trials for treating patients with various hematologic malignancies. As with any targeted cancer therapy, it is critically important to identify potential mechanisms of resistance, both for patient stratification and developing strategies to overcome resistance, either before it develops or as it emerges. METHODS: In order to gain a more comprehensive insight into the nature of venetoclax resistance mechanisms, we evaluated the changes in the BCL-2 family members at the genetic and expression levels in seven different venetoclax-resistant derived leukemia and lymphoma cell lines. RESULTS: Gene and protein expression analyses identified a number of different alterations in the expression of pro- and anti-apoptotic BCL-2 family members. In the resistant derived cells, an increase in either or both the anti-apoptotic proteins BCL-XL or MCL-1, which are not targeted by venetoclax was observed, and either concomitant or exclusive with a decrease in one or more pro-apoptotic proteins. In addition, mutational analysis also revealed a mutation in the BH3 binding groove (F104L) that could potentially interfere with venetoclax-binding. Not all changes may be causally related to venetoclax resistance and may only be an epiphenomenon. For resistant cell lines showing elevations in BCL-XL or MCL-1, strong synergistic cell killing was observed when venetoclax was combined with either BCL-XL- or MCL-1-selective inhibitors, respectively. This highlights the importance of BCL-XL- and MCL-1 as causally contributing to venetoclax resistance. CONCLUSIONS: Overall our study identified numerous changes in multiple resistant lines; the changes were neither mutually exclusive nor universal across the cell lines tested, thus exemplifying the complexity and heterogeneity of potential resistance mechanisms. Identifying and evaluating their contribution has important implications for both patient selection and the rational development of strategies to overcome resistance.
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Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Leucemia/tratamiento farmacológico , Linfoma/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Sulfonamidas/farmacología , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Linaje de la Célula/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Leucemia/genética , Leucemia/patología , Linfoma/genética , Linfoma/patología , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteína bcl-X/genéticaRESUMEN
BACKGROUND: Defects in programmed cell death, or apoptosis, are a hallmark of cancer. The anti-apoptotic B-cell lymphoma 2 (BCL-2) family proteins, including BCL-2, BCL-X(L), and MCL-1 have been characterized as key survival factors in multiple cancer types. Because cancer types with BCL2 and MCL1 amplification are more prone to inhibition of their respectively encoded proteins, we hypothesized that cancers with a significant frequency of BCL2L1 amplification would have greater dependency on BCL-X(L) for survival. METHODS: To identify tumor subtypes that have significant frequency of BCL2L1 amplification, we performed data mining using The Cancer Genome Atlas (TCGA) database. We then assessed the dependency on BCL-X(L) in a panel of cell lines using a selective and potent BCL-X(L) inhibitor, A-1155463, and BCL2L1 siRNA. Mechanistic studies on the role of BCL-X(L) were further undertaken via a variety of genetic manipulations. RESULTS: We identified colorectal cancer as having the highest frequency of BCL2L1 amplification across all tumor types examined. Colorectal cancer cell lines with BCL2L1 copy number >3 were more sensitive to A-1155463. Consistently, cell lines with high expression of BCL-XL and NOXA, a pro-apoptotic protein that antagonizes MCL-1 activity were sensitive to A-1155463. Silencing the expression of BCL-X(L) via siRNA killed the cell lines that were sensitive to A-1155463 while having little effect on lines that were resistant. Furthermore, silencing the expression of MCL-1 in resistant cell lines conferred sensitivity to A-1155463, whereas silencing NOXA abrogated sensitivity. CONCLUSIONS: This work demonstrates the utility of characterizing frequent genomic alterations to identify cancer survival genes. In addition, these studies demonstrate the utility of the highly potent and selective compound A-1155463 for investigating the role of BCL-X(L) in mediating the survival of specific tumor types, and indicate that BCL-X(L) inhibition could be an effective treatment for colorectal tumors with high BCL-X(L) and NOXA expression.
Asunto(s)
Neoplasias Colorrectales/genética , Genómica , Proteína bcl-X/genética , Benzotiazoles/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Análisis por Conglomerados , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Expresión Génica , Perfilación de la Expresión Génica , Humanos , Isoquinolinas/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteína bcl-X/antagonistas & inhibidores , Proteína bcl-X/metabolismoRESUMEN
Dual bromodomain BET inhibitors that bind with similar affinities to the first and second bromodomains across BRD2, BRD3, BRD4, and BRDT have displayed modest activity as monotherapy in clinical trials. Thrombocytopenia, closely followed by symptoms characteristic of gastrointestinal toxicity, have presented as dose-limiting adverse events that may have prevented escalation to higher dose levels required for more robust efficacy. ABBV-744 is a highly selective inhibitor for the second bromodomain of the four BET family proteins. In contrast to the broad antiproliferative activities observed with dual bromodomain BET inhibitors, ABBV-744 displayed significant antiproliferative activities largely although not exclusively in cancer cell lines derived from acute myeloid leukemia and androgen receptor positive prostate cancer. Studies in acute myeloid leukemia xenograft models demonstrated antitumor efficacy for ABBV-744 that was comparable with the pan-BET inhibitor ABBV-075 but with an improved therapeutic index. Enhanced antitumor efficacy was also observed with the combination of ABBV-744 and the BCL-2 inhibitor, venetoclax compared with monotherapies of either agent alone. These results collectively support the clinical evaluation of ABBV-744 in AML (Clinical Trials.gov identifier: NCT03360006).
Asunto(s)
Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Leucemia Mieloide Aguda/tratamiento farmacológico , Proteínas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Piridinas/farmacología , Pirroles/farmacología , Sulfonamidas/farmacología , Animales , Antineoplásicos/farmacología , Apoptosis , Proliferación Celular , Quimioterapia Combinada , Femenino , Humanos , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patología , Ratones , Ratones Endogámicos NOD , Ratones SCID , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Metastatic melanoma is responsible for approximately 80% of deaths from skin cancer. Microphthalmia-associated transcription factor (MITF) is a melanocyte-specific transcription factor that plays an important role in the differentiation, proliferation, and survival of melanocytes as well as in melanoma oncogenesis. MITF is amplified in approximately 15% of patients with metastatic melanoma. However, no small-molecule inhibitors of MITF currently exist. MITF was shown to associate with p300/CBP, members of the KAT3 family of histone acetyltransferase. p300 and CREB-binding protein (p300/CBP) regulate a wide range of cellular events such as senescence, apoptosis, cell cycle, DNA damage response, and cellular differentiation. p300/CBP act as transcriptional coactivators for multiple proteins in cancers, including oncogenic transcription factors such as MITF. In this study, we showed that our novel p300/CBP catalytic inhibitor, A-485, induces senescence in multiple melanoma cell lines, similar to silencing expression of EP300 (encodes p300) or MITF We did not observe apoptosis and increase invasiveness upon A-485 treatment. A-485 regulates the expression of MITF and its downstream signature genes in melanoma cell lines undergoing senescence. In addition, expression and copy number of MITF is significantly higher in melanoma cell lines that undergo A-485-induced senescence than resistant cell lines. Finally, we showed that A-485 inhibits histone-H3 acetylation but did not displace p300 at promoters of MITF and its putative downstream genes. Taken together, we provide evidence that p300/CBP inhibition suppressed the melanoma-driven transcription factor, MITF, and could be further exploited as a potential therapy for treating melanoma.
Asunto(s)
Proteína de Unión a CREB/antagonistas & inhibidores , Linaje de la Célula , Proteína p300 Asociada a E1A/antagonistas & inhibidores , Compuestos Heterocíclicos de 4 o más Anillos/farmacología , Melanoma/patología , Factor de Transcripción Asociado a Microftalmía/metabolismo , Transducción de Señal/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/farmacología , Acetilación , Proteína de Unión a CREB/metabolismo , Línea Celular Tumoral , Linaje de la Célula/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Senescencia Celular/efectos de los fármacos , Variaciones en el Número de Copia de ADN/genética , Proteína p300 Asociada a E1A/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Compuestos Heterocíclicos de 4 o más Anillos/química , Histonas/metabolismo , Humanos , Melanoma/genética , Regiones Promotoras Genéticas/genéticaRESUMEN
Competitive inhibitors of acetyl-lysine binding to the bromodomains of the BET (bromodomain and extra terminal) family are being developed for the treatment of solid and hematologic malignancies. The function of BET family member BRD4 at enhancers/superenhancers has been shown to sustain signal-dependent or pathogenic gene expression programs. Here, the hypothesis was tested that the transcription factor drivers of castration-resistant prostate cancer (CRPC) clinical progression, including the androgen receptor (AR), are critically dependent on BRD4 and thus represent a sensitive solid tumor indication for the BET inhibitor ABBV-075. DHT-stimulated transcription of AR target genes was inhibited by ABBV-075 without significant effect on AR protein expression. Furthermore, ABBV-075 disrupted DHT-stimulated recruitment of BET family member BRD4 to gene-regulatory regions cooccupied by AR, including the well-established PSA and TMPRSS2 enhancers. Persistent BET inhibition disrupted the composition and function of AR-occupied enhancers as measured by a reduction in AR and H3K27Ac ChIP signal and inhibition of enhancer RNA transcription. ABBV-075 displayed potent antiproliferative activity in multiple models of resistance to second-generation antiandrogens and inhibited the activity of the AR splice variant AR-V7 and ligand-binding domain gain-of-function mutations, F877L and L702H. ABBV-075 was also a potent inhibitor of MYC and the TMPRSS2-ETS fusion protein, important parallel transcription factor drivers of CRPC. IMPLICATIONS: The ability of BET family inhibitor ABBV-075 to inhibit transcription activation downstream of the initiating events of transcription factors like AR and TMPRSS2:ETS fusion proteins provides a promising therapeutic option for CRPC patients who have developed resistance to second-generation antiandrogens. Mol Cancer Res; 15(1); 35-44. ©2016 AACR.
Asunto(s)
Neoplasias de la Próstata Resistentes a la Castración/patología , Piridonas/farmacología , Sulfonamidas/farmacología , Factores de Transcripción/antagonistas & inhibidores , Antagonistas de Andrógenos/farmacología , Animales , Antineoplásicos/farmacología , Benzamidas , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cromatina/metabolismo , Dihidrotestosterona/farmacología , Resistencia a Antineoplásicos/efectos de los fármacos , Elementos de Facilitación Genéticos/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Concentración 50 Inhibidora , Ligandos , Masculino , Ratones , Nitrilos , Fenotipo , Feniltiohidantoína/análogos & derivados , Feniltiohidantoína/farmacología , Neoplasias de la Próstata Resistentes a la Castración/genética , Dominios Proteicos , Receptores Androgénicos/química , Receptores Androgénicos/genética , Receptores Androgénicos/metabolismo , Transducción de Señal/efectos de los fármacos , Factores de Transcripción/metabolismo , Transcripción Genética/efectos de los fármacosRESUMEN
An increasing number of BET family protein inhibitors have recently entered clinical trials. It has been reported that attempts of monitoring target engagement of the BET bromodomain inhibitor OTX015 using literature-described putative pharmacodynamic markers, such as c-Myc, BRD2, etc., failed to detect pharmacodynamic marker responses in AML patients treated at active dose and those with clinical responses. Here, we report the identification and characterization of HEXIM1 and other genes as robust pharmacodynamic markers for BET inhibitors. Global gene expression profiling studies were carried out using cancer cells and surrogate tissues, such as whole blood and skin, to identify genes that are modulated by BET family proteins. Candidate markers were further characterized for concentration- and time-dependent responses to the BET inhibitor ABBV-075 in vitro and in vivo HEXIM1 was found to be the only gene that exhibited robust and consistent modulation by BET inhibitors across multiple cancer indications and surrogate tissues. Markers such as SERPINI1, ZCCHC24, and ZMYND8 were modulated by ABBV-075 and other BET inhibitors across cancer cell lines and xenograft tumors but not in blood and skin. Significant downregulation of c-Myc, a well-publicized target of BET inhibitors, was largely restricted to hematologic cancer cell lines. Incorporating well-characterized pharmacodynamic markers, such as HEXIM1 and other genes described here, can provide a better understanding of potential efficacy and toxicity associated with inhibiting BET family proteins and informs early clinical decisions on BET inhibitor development programs. Mol Cancer Ther; 16(2); 388-96. ©2016 AACR.
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Antineoplásicos/farmacología , Biomarcadores , Neoplasias/genética , Proteínas Nucleares/antagonistas & inhibidores , Proteínas de Unión al ARN/genética , Animales , Antineoplásicos/farmacocinética , Biopsia , Línea Celular Tumoral , Análisis por Conglomerados , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Ratones , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Neoplasias/patología , Especificidad de Órganos/genética , Proteínas de Unión al ARN/metabolismo , Factores de Transcripción , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Improving the congruity of preclinical models with cancer as it is manifested in humans is a potential way to mitigate the high attrition rate of new cancer therapies in the clinic. In this regard, three-dimensional (3D) tumor cultures in vitro have recently regained interest as they have been acclaimed to have higher similarity to tumors in vivo than to cells grown in monolayers (2D). To identify cancer functions that are active in 3D rather than in 2D cultures, we compared the transcriptional profiles (TPs) of two non-small cell lung carcinoma cell lines, NCI-H1650 and EBC-1 grown in both conditions to the TP of xenografted tumors. Because confluence, diameter or volume can hypothetically alter TPs, we made intra- and inter-culture comparisons using samples with defined dimensions. As projected by Ingenuity Pathway Analysis (IPA), a limited number of signal transduction pathways operational in vivo were better represented by 3D than by 2D cultures in vitro. Growth of 2D and 3D cultures as well as xenografts induced major changes in the TPs of these 3 modes of culturing. Alterations of transcriptional network activation that were predicted to evolve similarly during progression of 3D cultures and xenografts involved the following functions: hypoxia, proliferation, cell cycle progression, angiogenesis, cell adhesion, and interleukin activation. Direct comparison of TPs of 3D cultures and xenografts to monolayer cultures yielded up-regulation of networks involved in hypoxia, TGF and Wnt signaling as well as regulation of epithelial mesenchymal transition. Differences in TP of 2D and 3D cancer cell cultures are subject to progression of the cultures. The emulation of the predicted cell functions in vivo is therefore not only determined by the type of culture in vitro but also by the confluence or diameter of the 2D or 3D cultures, respectively. Consequently, the successful implementation of 3D models will require phenotypic characterization to verify the relevance of applying these models for drug development.
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Regulación Neoplásica de la Expresión Génica , Transcriptoma , Animales , Técnicas de Cultivo de Célula , Línea Celular Tumoral , Análisis por Conglomerados , Modelos Animales de Enfermedad , Femenino , Perfilación de la Expresión Génica , Xenoinjertos , Humanos , Ratones , Esferoides CelularesRESUMEN
Hyperexpression of antiapoptotic BCL-2 family proteins allows cells to survive despite the receipt of signals that would ordinarily induce their deletion, a facet frequently exploited by tumors. Tumors addicted to the BCL-2 family proteins for survival are now being targeted therapeutically. For example, navitoclax, a BCL-2/BCL-XL/BCL-W inhibitor, is currently in phase I/II clinical trials in numerous malignancies. However, the related family member, MCL-1, limits the efficacy of navitoclax and other chemotherapeutic agents. In the present study, we identify breast cancer cell lines that depend upon MCL-1 for survival and subsequently determine the mechanism of apoptosis mediated by the MCL-1 selective inhibitor A-1210477. We demonstrate that apoptosis resulting from a loss in MCL-1 function requires expression of the proapoptotic protein BAK. However, expression of BCL-XL can limit apoptosis resulting from loss in MCL-1 function through sequestration of free BIM. Finally, we demonstrate substantial synergy between navitoclax and MCL-1 siRNA, the direct MCL-1 inhibitor A-1210477, or the indirect MCL-1 inhibitor flavopiridol, highlighting the therapeutic potential for inhibiting BCL-XL and MCL-1 in breast cancer.
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Neoplasias de la Mama/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Compuestos de Anilina/farmacología , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Neoplasias de la Mama/genética , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Femenino , Expresión Génica , Humanos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/antagonistas & inhibidores , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/genética , Sulfonamidas/farmacología , Proteína Destructora del Antagonista Homólogo bcl-2/genética , Proteína Destructora del Antagonista Homólogo bcl-2/metabolismo , Proteína bcl-X/genética , Proteína bcl-X/metabolismoRESUMEN
Protein tyrosine phosphatase 1B (PTP1B) has recently been implicated in the regulation of body weight. A surprising phenotype of PTP1B-deficient mice is their resistance to diet-induced obesity. Since leptin is one of the primary hormones involved in the regulation of body weight and energy homeostasis, we investigated whether PTP1B affects leptin receptor (lepR) signaling directly. A mouse hypothalamic cell line, GT1-7, was established as a suitable cell model for the study of leptin signaling. Stimulation of GT1-7 cells by leptin caused tyrosine phosphorylation of endogenous STAT3 and activation of a STAT-dependent luciferase reporter gene. Over-expression of PTP1B in GT1-7 cells resulted in a dose-dependent decrease in endogenous JAK2 and STAT3 tyrosine phosphorylation compared with cells transfected with lepR alone. Consistent with inhibition of JAK-STAT signaling, PTP1B over-expression caused a dose-dependent decrease in leptin-induced, STAT-dependent luciferase reporter gene activation in GT1-7 cells. Furthermore, over-expression of PTP1B led to a decrease in mRNA accumulation of suppressor-of-cytokine-signalling-3 (SOCS3) and c-fos, genes that are acutely induced by leptin. Using gene microarray analysis, we confirmed that PTP1B reduces the level of gene expression of SOCS3 and showed that the expression level of other leptin-regulated genes was affected. Genes up-regulated by leptin were decreased in cells over-expressing PTP1B. Conversely, the expression of genes down-regulated by leptin was enhanced by PTP1B over-expression in GT1-7 cells. Our findings indicate that PTP1B is a negative regulator of leptin signaling and suggest that PTP1B inhibitors might be efficacious in the treatment of obesity by increasing leptin sensitivity.
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Hipotálamo/citología , Leptina/fisiología , Proteínas Tirosina Fosfatasas/fisiología , Proteínas Proto-Oncogénicas , Animales , Línea Celular , Proteínas de Unión al ADN/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Hipotálamo/metabolismo , Janus Quinasa 2 , Ratones , Fosforilación , Proteína Tirosina Fosfatasa no Receptora Tipo 1 , Proteínas Tirosina Fosfatasas/genética , Proteínas Tirosina Quinasas/metabolismo , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/fisiología , Receptores de Leptina , Factor de Transcripción STAT3 , Transducción de Señal , Transactivadores/metabolismo , Activación Transcripcional , TransfecciónRESUMEN
ABT-263 inhibits the antiapoptotic proteins Bcl-2, Bcl-x(L), and Bcl-w and has single-agent efficacy in numerous small cell lung carcinoma (SCLC) and leukemia/lymphoma cell lines in vitro and in vivo. It is currently in clinical trials for treating patients with SCLC and various leukemia/lymphomas. Identification of predictive markers for response will benefit the clinical development of ABT-263. We identified the expression of Bcl-2 family genes that correlated best with sensitivity to ABT-263 in a panel of 36 SCLC and 31 leukemia/lymphoma cell lines. In cells sensitive to ABT-263, expression of Bcl-2 and Noxa is elevated, whereas expression of Mcl-1 is higher in resistant cells. We also examined global expression differences to identify gene signature sets that correlated with sensitivity to ABT-263 to generate optimal signature sets predictive of sensitivity to ABT-263. Independent cell lines were used to verify the predictive power of the gene sets and to refine the optimal gene signatures. When comparing normal lung tissue and SCLC primary tumors, the expression pattern of these genes in the tumor tissue is most similar to sensitive SCLC lines, whereas normal tissue is most similar to resistant SCLC lines. Most of the genes identified using global expression patterns are related to the apoptotic pathway; however, all but Bcl-rambo are distinct from the Bcl-2 family. This study leverages global expression data to identify key gene expression patterns for sensitivity to ABT-263 in SCLC and leukemia/lymphoma and may provide guidance in the selection of patients in future clinical trials.
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Compuestos de Anilina/farmacología , Biomarcadores Farmacológicos/análisis , Perfilación de la Expresión Génica , Leucemia/genética , Neoplasias Pulmonares/genética , Linfoma/genética , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Carcinoma Pulmonar de Células Pequeñas/genética , Sulfonamidas/farmacología , Compuestos de Anilina/uso terapéutico , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Biomarcadores Farmacológicos/metabolismo , Línea Celular Tumoral , Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Redes Reguladoras de Genes/efectos de los fármacos , Humanos , Leucemia/diagnóstico , Leucemia/metabolismo , Leucemia/patología , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Linfoma/diagnóstico , Linfoma/metabolismo , Linfoma/patología , Familia de Multigenes/efectos de los fármacos , Análisis de Secuencia por Matrices de Oligonucleótidos , Pronóstico , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Carcinoma Pulmonar de Células Pequeñas/diagnóstico , Carcinoma Pulmonar de Células Pequeñas/metabolismo , Carcinoma Pulmonar de Células Pequeñas/patología , Sulfonamidas/uso terapéuticoRESUMEN
Both Akt and Aurora A kinase have been shown to be important targets for intervention for cancer therapy. We report here that Compound A (A-443654), a specific Akt inhibitor, interferes with mitotic progression and bipolar spindle formation. Compound A induces G(2)/M accumulation, defects in centrosome separation, and formation of either monopolar arrays or disorganized spindles. On the basis of gene expression array studies, we identified Aurora A as one of the genes regulated transcriptionally by Akt inhibitors including Compound A. Inhibition of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway, either by PI3K inhibitor LY294002 or by Compound A, dramatically inhibits the promoter activity of Aurora A, whereas the mammalian target of rapamycin inhibitor has little effect, suggesting that Akt might be responsible for up-regulating Aurora A for mitotic progression. Further analysis of the Aurora A promoter region indicates that the Ets element but not the Sp1 element is required for Compound A-sensitive transcriptional control of Aurora A. Overexpression of Aurora A in cells treated with Compound A attenuates the mitotic arrest and the defects in bipolar spindle formation induced by Akt inhibition. Our studies suggest that that Akt may promote mitotic progression through the transcriptional regulation of Aurora A.