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1.
Oncogene ; 36(24): 3441-3449, 2017 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-28114285

RESUMO

Chronic lymphocytic leukemia (CLL) is characterized by the accumulation of B cells in the hematopoietic system and lymphoid tissues. Although inhibitors targeting the B-cell receptor (BCR) pathway have been successful in the treatment of the disease, the underlying mechanisms leading to BCR over-activity in CLL are not fully understood. In this study, we found that HSP90, a highly conserved molecular chaperone, is overexpressed in CLL compared with resting B cells. HSP90 overexpression is accompanied by the overexpression of several BCR kinases including LYN, spleen tyrosine kinase, Bruton tyrosine kinase and AKT. Chemical and immune-precipitation demonstrated that these BCR constituents are present in a multi-client chaperone complex with HSP90. Inhibition of HSP90 with PU-H71 destabilized the BCR kinases and caused apoptosis of CLL cells through the mitochondrial apoptotic pathway. Further, PU-H71 induced apoptosis in the presence of stromal co-culture or cytoprotective survival signals. Finally, genetic knockdown of HSP90 and its client AKT, but not BTK, reduced CLL viability. Overall, our study suggests that the chaperone function of HSP90 contributes to the over-activity of the BCR signaling in CLL and inhibition of HSP90 has the potential to achieve a multi-targeting effect. Thus, HSP90 inhibition may be explored to prevent or overcome drug resistance to single targeting agents.


Assuntos
Proteínas de Choque Térmico HSP90/metabolismo , Leucemia Linfocítica Crônica de Células B/metabolismo , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Quinase Syk/metabolismo , Quinases da Família src/metabolismo , Tirosina Quinase da Agamaglobulinemia , Benzodioxóis/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Cocultura , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Purinas/farmacologia
2.
Cancer Res ; 74(24): 7475-86, 2014 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-25320008

RESUMO

Histone deacetylases (HDAC) that regulate gene expression are being explored as cancer therapeutic targets. In this study, we focused on HDAC6 based on its ability to inhibit cancerous Hsp90 chaperone activities by disrupting Hsp90/p23 interactions. To identify novel HDAC6 inhibitors, we used a dual-luciferase reporter system in cell culture and living mice by bioluminescence imaging (BLI). On the basis of existing knowledge, a library of hydrazone compounds was generated for screening by coupling cinnamic hydroxamates with aldehydes and ketones. Potency and selectivity were determined by in vitro HDAC profiling assays, with further evaluation to inhibit Hsp90(α/ß)/p23 interactions by BLI. In this manner, we identified compound 1A12 as a dose-dependent inhibitor of Hsp90(α/ß)/p23 interactions, UKE-1 myeloid cell proliferation, p21(waf1) upregulation, and acetylated histone H3 levels. 1A12 was efficacious in tumor xenografts expressing Hsp90(α)/p23 reporters relative to carrier control-treated mice as determined by BLI. Small animal (18)F-FDG PET/CT imaging on the same cohort showed that 1A12 also inhibited glucose metabolism relative to control subjects. Ex vivo analyses of tumor lysates showed that 1A12 administration upregulated acetylated-H3 by approximately 3.5-fold. Taken together, our results describe the discovery and initial preclinical validation of a novel selective HDAC inhibitor.


Assuntos
Inibidores de Histona Desacetilases/isolamento & purificação , Ácidos Hidroxâmicos/isolamento & purificação , Imagem Molecular , Imagem Multimodal , Acetilação , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cinamatos/síntese química , Cinamatos/isolamento & purificação , Cinamatos/farmacologia , Fluordesoxiglucose F18 , Desacetilase 6 de Histona , Inibidores de Histona Desacetilases/síntese química , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/química , Humanos , Ácidos Hidroxâmicos/síntese química , Camundongos , Células Mieloides/efeitos dos fármacos
3.
Cell Death Differ ; 15(5): 859-66, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18239673

RESUMO

Members of the inhibitor of apoptosis protein (IAP) family have demonstrated functions in cell death, cell signalling, cell migration and mitosis. Several of them are E3 enzymes in the ubiquitination of proteins that leads to their degradation by the proteosomal machinery. We previously reported that one of them, cellular inhibitor of apoptosis protein-1 (c-IAP1), migrated from the nucleus to the surface of the Golgi apparatus in cells undergoing differentiation. Here, we show that c-IAP1 is a client protein of the stress protein HSP90 beta. In three distinct cellular models, the two proteins interact and migrate from the nucleus to the cytoplasm along the differentiation process through a leptomycin B-sensitive pathway. Inhibition of HSP90 proteins by small chemical molecules and specific depletion of HSP90 beta isoform by siRNA both lead to auto-ubiquitination of c-IAP1 and its degradation by the proteasome machinery. This chaperone function of HSP90 towards c-IAP1 is specific of its beta isoform as specific depletion of HSP90alpha does not affect c-IAP1 content. Chemical inhibition of HSP90 or siRNA-mediated depletion of HSP90 beta both inhibit cell differentiation, which can be reproduced by siRNA-mediated depletion of c-IAP1. Altogether, these results suggest that HSP90 beta prevents auto-ubiquitination and degradation of its client protein c-IAP1, whose depletion would be sufficient to inhibit cell differentiation.


Assuntos
Diferenciação Celular/fisiologia , Proteínas de Choque Térmico HSP90/metabolismo , Proteínas Inibidoras de Apoptose/metabolismo , Isoformas de Proteínas/metabolismo , Animais , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Células Epiteliais/citologia , Células Epiteliais/fisiologia , Proteínas de Choque Térmico HSP90/genética , Humanos , Proteínas Inibidoras de Apoptose/genética , Macrófagos/citologia , Macrófagos/fisiologia , Isoformas de Proteínas/genética , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo
4.
Cell Death Differ ; 2008 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-25361076

RESUMO

Members of the inhibitor of apoptosis protein (IAP) family have demonstrated functions in cell death, cell signalling, cell migration and mitosis. Several of them are E3 enzymes in the ubiquitination of proteins that leads to their degradation by the proteosomal machinery. We previously reported that one of them, cellular inhibitor of apoptosis protein-1 (c-IAP1), migrated from the nucleus to the surface of the Golgi apparatus in cells undergoing differentiation. Here, we show that c-IAP1 is a client protein of the stress protein HSP90ß. In three distinct cellular models, the two proteins interact and migrate from the nucleus to the cytoplasm along the differentiation process through a leptomycin B-sensitive pathway. Inhibition of HSP90 proteins by small chemical molecules and specific depletion of HSP90ß isoform by siRNA both lead to auto-ubiquitination of c-IAP1 and its degradation by the proteasome machinery. This chaperone function of HSP90 towards c-IAP1 is specific of its ß isoform as specific depletion of HSP90α does not affect c-IAP1 content. Chemical inhibition of HSP90 or siRNA-mediated depletion of HSP90ß both inhibit cell differentiation, which can be reproduced by siRNA-mediated depletion of c-IAP1. Altogether, these results suggest that HSP90ß prevents auto-ubiquitination and degradation of its client protein c-IAP1, whose depletion would be sufficient to inhibit cell differentiation.Cell Death and Differentiation advance online publication, 1 February 2008; doi:10.1038/sj.cdd.4402320.

5.
Anticancer Agents Med Chem ; 6(1): 1-8, 2006 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-16475922

RESUMO

Hsp90 is a chaperone with important roles in maintaining transformation and in elevating the survival and growth potential of cancer cells. Activation of signaling pathways mediated by Hsp90 protein clients is necessary for cell proliferation, regulation of cell cycle progression and apoptosis. Additionally, gain-of-function mutations responsible for transformation often require Hsp90 for the maintenance of their folded, functionally active conformations. These characteristics promise Hsp90 as an important target in cancer therapy and prompt for the identification, development and clinical translation of small molecule inhibitors of the chaperone. This review intends to update the reader on the status of several existing and emerging classes of direct inhibitors of Hsp90 ATPase activity.


Assuntos
Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Difosfato de Adenosina/metabolismo , Adenosina Trifosfatases/antagonistas & inibidores , Adenosina Trifosfatases/metabolismo , Trifosfato de Adenosina/metabolismo , Antineoplásicos/classificação , Antineoplásicos/farmacologia , Desenho de Fármacos , Proteínas de Choque Térmico HSP90/metabolismo , Ligação Proteica
7.
Science ; 293(5534): 1484-7, 2001 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-11520986

RESUMO

Pathogenic enterococci are becoming resistant to currently available antibiotics, including vancomycin, the drug of last resort for Gram-positive infections. Enterococci pose a significant public health threat, not least because of the risk of transferring vancomycin resistance to the ubiquitous Staphylococcus aureus. Vancomycin resistance is manifested by cell wall peptidoglycan precursors with altered termini that cannot bind the antibiotic. Small molecules with well-oriented nucleophile-electrophile assembly and complementary chirality to the peptidoglycan termini were identified as catalytic and selective cleavers of the peptidoglycan precursor depsipeptide. These molecules were tested in combination with vancomycin and were found to re-sensitize vancomycin-resistant bacteria to the antibiotic.


Assuntos
Alanina/metabolismo , Enterococcus/efeitos dos fármacos , Lactatos/metabolismo , Peptidoglicano/metabolismo , Pirrolidinas/farmacologia , Vancomicina/farmacologia , Alanina/análogos & derivados , Antibacterianos/síntese química , Antibacterianos/química , Antibacterianos/metabolismo , Antibacterianos/farmacologia , Catálise , Técnicas de Química Combinatória , Simulação por Computador , Desenho de Fármacos , Sinergismo Farmacológico , Enterococcus/metabolismo , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/metabolismo , Enterococcus faecium/efeitos dos fármacos , Enterococcus faecium/metabolismo , Hidrólise , Compostos de Metilureia/síntese química , Compostos de Metilureia/metabolismo , Compostos de Metilureia/farmacologia , Testes de Sensibilidade Microbiana , Modelos Moleculares , Oligopeptídeos/síntese química , Oligopeptídeos/metabolismo , Oligopeptídeos/farmacologia , Biblioteca de Peptídeos , Peptidoglicano/química , Precursores de Proteínas/metabolismo , Pirrolidinas/síntese química , Pirrolidinas/metabolismo , Estereoisomerismo , Vancomicina/metabolismo , Resistência a Vancomicina
9.
Chem Biol ; 8(3): 289-99, 2001 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-11306353

RESUMO

BACKGROUND: The Hsp90s contain a conserved pocket that binds ATP/ADP and plays an important role in the regulation of chaperone function. Occupancy of this pocket by several natural products (geldanamycin (GM) and radicicol) alters Hsp90 function and results in the degradation of a subset of proteins (i.e. steroid receptors, Her2, Raf). We have used the structural features of this pocket to design a small molecule inhibitor of Hsp90. RESULTS: The designed small molecule PU3 competes with GM for Hsp90 binding with a relative affinity of 15-20 microM. PU3 induces degradation of proteins, including Her2, in a manner similar to GM. Furthermore, PU3 inhibits the growth of breast cancer cells causing retinoblastoma protein hypophosphorylation, G1 arrest and differentiation. CONCLUSIONS: PU3 is representative of a novel class of synthetic compounds that binds to Hsp90 and inhibits the proliferation of cancer cells. These reagents could provide a new strategy for the treatment of cancers.


Assuntos
Nucleotídeos de Adenina/metabolismo , Neoplasias da Mama/patologia , Ciclo Celular/efeitos dos fármacos , Desenho de Fármacos , Proteínas de Choque Térmico HSP90/metabolismo , Receptor ErbB-2/efeitos dos fármacos , Benzoquinonas , Ligação Competitiva , Diferenciação Celular/efeitos dos fármacos , Feminino , Humanos , Lactamas Macrocíclicas , Ligação Proteica , Quinonas/metabolismo , Receptor ErbB-2/metabolismo , Células Tumorais Cultivadas/efeitos dos fármacos
10.
Cancer Res ; 60(8): 2090-4, 2000 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-10786665

RESUMO

Geldanamycin (GM) is a natural antibiotic that binds Hsp90 and induces the degradation of receptor tyrosine kinases, steroid receptors, and Raf. It is a potent inhibitor of cancer cells that overexpress HER-kinases, but its effects on other important proteins may cause significant toxicity and limit its clinical use. We report the synthesis and identification of a GM dimer, GMD-4c, which had selective activity against HER-kinases. Selectivity was a function of linker length and required two intact GM moieties. GMD-4c is a potent inducer of G1 block and apoptosis of breast cancer cell lines that overexpress HER2, but does not appreciably inhibit the growth of 32D cells that lack HER-kinases. GMD-4c could be useful in the treatment of carcinomas dependent on HER-kinases.


Assuntos
Antibióticos Antineoplásicos/farmacologia , Neoplasias da Mama/enzimologia , Quinonas/farmacologia , Receptor ErbB-2/metabolismo , Antibióticos Antineoplásicos/química , Antibióticos Antineoplásicos/uso terapêutico , Benzoquinonas , Western Blotting , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Divisão Celular/efeitos dos fármacos , Dimerização , Regulação para Baixo/efeitos dos fármacos , Humanos , Imuno-Histoquímica , Concentração Inibidora 50 , Lactamas Macrocíclicas , Proteínas Proto-Oncogênicas c-raf/metabolismo , Quinonas/química , Quinonas/uso terapêutico , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-2/biossíntese , Receptor IGF Tipo 1/metabolismo , Receptores de Estrogênio/metabolismo , Especificidade por Substrato , Células Tumorais Cultivadas
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