RESUMO
On the basis of deguelin, a series of the B,C-ring truncated surrogates with N-substituted amide linkers were investigated as HSP90 inhibitors. The structure activity relationship of the template was studied by incorporating various substitutions on the nitrogen of the amide linker and examining their HIF-1α inhibition. Among them, compound 57 showed potent HIF-1α inhibition and cytotoxicity in triple-negative breast cancer lines in a dose-dependent manner. Compound 57 downregulated expression and phosphorylation of major client proteins of HSP90 including AKT, ERK and STAT3, indicating that its antitumor activity was derived from the inhibition of HSP90 function. The molecular modeling of 57 demonstrated that 57 bound well to the C-terminal ATP-binding pocket in the open conformation of the hHSP90 homodimer with hydrogen bonding and pi-cation interactions. Overall, compound 57 is a potential antitumor agent for triple-negative breast cancer as a HSP90 C-terminal inhibitor.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Rotenona/análogos & derivados , Antineoplásicos/síntese química , Antineoplásicos/química , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Modelos Moleculares , Estrutura Molecular , Rotenona/síntese química , Rotenona/química , Rotenona/farmacologia , Relação Estrutura-AtividadeRESUMO
Tumor metastasis remains the cause of 90% of cancer-related deaths. Cancer stem cells (CSC) are thought to be responsible for the aggressive and metastatic nature of triple-negative breast cancers (TNBC), and new therapeutic strategies are being devised to target them. Flubendazole (FLU) is a widely used anthelmintic agent that also exhibits anticancer activity in several cancer types. The aim of this study was to characterize the mechanism of action of FLU on breast cancer stem cell (BCSC)-like properties and metastasis in TNBC. FLU treatment caused a significant induction of apoptosis, accompanied by G2/M phase accumulation, caspase-3/-7 activation and the dysregulation of STAT3 activation in TNBC cells. The latter phenomenon was associated with impairment of cancer stem-like traits, concomitant with a reduction in the CD24low /CD44high , CD24high /CD49fhigh subpopulation, ALDH1 activity and mammosphere formation. The BCSC-enriched populations exhibited enhanced metastasis with higher STAT3 activation, while FLU administration inhibited tumor growth, angiogenesis and lung and liver metastasis, coinciding with decreased MMP-2 and MMP-9 levels in circulating blood. FLU kills not only rapid proliferating tumor cells but also effectively eradicates BCSC-like cells in vitro and in vivo. Our findings warrant further investigation of FLU as a treatment for metastatic TNBC.
Assuntos
Mebendazol/análogos & derivados , Metástase Neoplásica/tratamento farmacológico , Fator de Transcrição STAT3/antagonistas & inibidores , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/metabolismo , Antígenos CD/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/prevenção & controle , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/prevenção & controle , Mebendazol/farmacologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismoRESUMO
In the presence of copper (Cu), disulfiram (DSF) suppresses properties associated with cancer stem cells (CSCs) in breast cancer, but the mechanism of action is poorly understood. In the present study, we observed that DSF/Cu treatment induced apoptosis, mediated by caspase-3 activation in triple-negative breast cancer (TNBC) cells. DSF/Cu treatment also specifically targeted CSC-like cell populations, marked by the inhibition of ALDH1 activity, the suppression of CD44+/CD24-and CD49f+/CD24 + subpopulations, and the subsequent impairment of mammosphere formation. These effects were functionally associated with a significant impact on the STAT3 signaling pathway, characterized by the downregulation of phospho-STAT3, cyclin D1 and survivin. In an MDA-MB-231-derived xenograft model, DSF administration significantly downregulated ALDH1A1, CD44 and phospho-STAT3 levels. These findings show for the first time that DSF suppresses stem-like properties in TNBC by targeting the STAT3 signaling pathway.
Assuntos
Dissulfiram/administração & dosagem , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fator de Transcrição STAT3/metabolismo , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/metabolismo , Antineoplásicos/administração & dosagem , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Células-Tronco Neoplásicas/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Resultado do Tratamento , Neoplasias de Mama Triplo Negativas/patologiaRESUMO
Despite recent advances, there remains a significant unmet need for the development of new targeted therapies for triple-negative breast cancer (TNBC). Although the heat shock protein HSP90 is a promising target, previous inhibitors have had issues during development including undesirable induction of the heat shock response (HSR) and off-target effects leading to toxicity. SL-145 is a novel, rationally-designed C-terminal HSP90 inhibitor that induces apoptosis in TNBC cells via the suppression of oncogenic AKT, MEK/ERK, and JAK2/STAT3 signaling and does not trigger the HSR, in contrast to other inhibitors. In an orthotopic allograft model incorporating breast cancer stem cell-enriched TNBC tumors, SL-145 potently suppressed tumor growth, angiogenesis, and metastases concomitant with dysregulation of the JAK2/STAT3 signaling pathway. Our findings highlight the potential of SL-145 in suppressing metastatic TNBC independent of the HSR.
Assuntos
Antineoplásicos , Neoplasias de Mama Triplo Negativas , Antineoplásicos/farmacologia , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Proteínas de Choque Térmico HSP90/metabolismo , Resposta ao Choque Térmico , Humanos , Neoplasias de Mama Triplo Negativas/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Triple-negative breast cancer (TNBC) is an aggressive heterogeneous disease with a divergent profile. It has an earlier tendency to form metastases and is associated with poor clinical outcomes due to the limited treatment options available. Heat-shock protein (HSP90) represents a potential treatment target as it promotes tumor progression and metastasis by modulating the maturation and stabilization of signal transduction proteins. We sought to investigate the efficacy of the C-terminal HSP90 inhibitor L80 on cell proliferation, breast cancer stem cell (BCSC)-like properties, tumor growth and metastasis. L80 suppressed cell viability and concomitantly inhibited AKT/MEK/ERK/JAK2/STAT3 signaling in TNBC cells but did not induce cytotoxicity in normal cells. L80 effectively targeted BCSC-like traits, together with significant reductions in the CD44high/CD24low-population, ALDH1 activity and mammosphere forming-ability. In support of the in vitro observations, L80 administration caused significant impairment in tumor growth, angiogenesis and distant metastases in an orthotopic allograft model with BCSC-enriched cells in vivo. These phenomena were associated with the suppression of BCSC-like characteristics and STAT3 dysfunction. Our findings highlight properties of the L80 compound that may be useful in suppressing metastatic TNBC.
Assuntos
Antineoplásicos/farmacologia , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Metástase Neoplásica/tratamento farmacológico , Fator de Transcrição STAT3/antagonistas & inibidores , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Neovascularização Patológica/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Although trastuzumab provides significant clinical benefit for HER2-positive breast cancers, responses are limited by the emergence of resistance. Trastuzumab resistance is a multi-factorial phenomenon thought to arise from the presence of cancer stem cells and interactions between truncated p95HER2 and HER family members. Flubendazole (FLU) is a potent anthelmintic agent with an exceptional safety profile. Evidence also suggests that it can act as an anticancer agent in several cancer cell types. We sought to investigate the effect of FLU on apoptosis, HER2/Akt signaling, breast cancer stem cell (BCSC)-like properties and trastuzumab resistance in HER2-positive breast cancer cells. FLU treatment induced apoptosis, associated with a significant downregulation of truncated p95HER2, phospho-HER2, phospho-HER3 and phospho-Akt levels, as well as suppression of HER2/HER3 hetero-dimerization in both trastuzumab-sensitive and -resistant lines. FLU effectively targeted BCSC-like properties including aldehyde dehydrogenase 1 (ALDH1) expression and the CD44high/CD24low phenotype, concomitant with a suppression of mammosphere-forming ability. FLU administration also caused significant tumor suppression in trastuzumab-resistant xenografts, coinciding with the downregulation of BCSC-related markers and intracellular HER2. These findings highlight the mechanisms of action of FLU in overcoming trastuzumab resistance in breast cancer.
Assuntos
Antineoplásicos Imunológicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Mebendazol/análogos & derivados , Células-Tronco Neoplásicas/efeitos dos fármacos , Receptor ErbB-2/fisiologia , Transdução de Sinais/efeitos dos fármacos , Trastuzumab/farmacologia , Animais , Apoptose/efeitos dos fármacos , Neoplasias da Mama/química , Neoplasias da Mama/patologia , Caspases/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Feminino , Humanos , Mebendazol/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Receptor ErbB-2/análise , Transdução de Sinais/fisiologiaRESUMO
Triple-negative breast cancers (TNBC) often exhibit an aggressive phenotype. Disulfiram (DSF) is an approved drug for the treatment of alcohol dependence, but has also been shown to kill TNBC cells in a copper (Cu)-dependent manner. Exactly how this occurs has not been clearly elucidated. We sought to investigate the mechanisms responsible for DSF/Cu-dependent induction of apoptosis and suppression of lung colonization by TNBC cells. DSF/Cu induced anoikis and significantly suppressed cell migration and invasion with negative effects on focal adhesions, coinciding with vimentin breakdown and calpain activation in TNBC cells. In a xenograft tumor model, DSF suppressed tumor growth and lung nodule growth, which was also associated with calpain activation. These findings warrant further investigation of disulfiram as a potential treatment for metastatic TNBC.
Assuntos
Anoikis/efeitos dos fármacos , Antineoplásicos/farmacologia , Calpaína/metabolismo , Movimento Celular/efeitos dos fármacos , Dissulfiram/farmacologia , Neoplasias Pulmonares/prevenção & controle , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cobre/farmacologia , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/enzimologia , Citoesqueleto/patologia , Relação Dose-Resposta a Droga , Ativação Enzimática , Feminino , Adesões Focais/efeitos dos fármacos , Adesões Focais/enzimologia , Adesões Focais/patologia , Humanos , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/secundário , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica , Proteólise , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Neoplasias de Mama Triplo Negativas/enzimologia , Neoplasias de Mama Triplo Negativas/patologia , Carga Tumoral/efeitos dos fármacos , Vimentina/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Ubiquitin-specific protease 34 (USP34) is a deubiquitinating enzyme that regulates Axin stability and plays a critical role in Wnt/ß-catenin signaling. We sought to investigate the role of USP34 on epithelial-mesenchymal (EMT) induction and its effects on mammary epithelial stem cells. USP34 expression levels were relatively lower in MDA-MB-231 and 4T1 mesenchymal-like cells when compared to epithelial-like cells. Inhibition of USP34 in NMuMG cells induced EMT, as evidenced by the upregulation of EMT markers including N-cadherin, phospho-Smad3, Snail and active-ß-catenin, as well as the downregulation of Axin 1 and E-cadherin. USP34 knockdown (KD) in these cells also resulted in the acquisition of invasive behavior, and promoted stemness as indicated by enhanced mammosphere-forming ability, concomitant with the upregulation of Nanog, Oct4 and Sox2 mRNA expression. Endogenous USP34 expression was observed to be at low levels in virgin mouse mammary glands in vivo. When USP34-KD cells were transplanted into the cleared mammary fat pads (CFP) of mice, these cells reconstituted the mammary gland with ductal tree development within 3months. Our findings suggest a previously unknown role for USP34 in mammary gland development.
Assuntos
Células Epiteliais/metabolismo , Transição Epitelial-Mesenquimal , Glândulas Mamárias Animais/citologia , Glândulas Mamárias Humanas/citologia , Células-Tronco/metabolismo , Proteases Específicas de Ubiquitina/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Autorrenovação Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Feminino , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Invasividade Neoplásica , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Esferoides Celulares/efeitos dos fármacos , Esferoides Celulares/metabolismo , Células-Tronco/efeitos dos fármacos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/farmacologia , Proteases Específicas de Ubiquitina/metabolismoRESUMO
HER2-positive breast tumors are known to harbor cancer stem-like cell populations and are associated with an aggressive tumor phenotype and poor clinical outcomes. Disulfiram (DSF), an anti-alcoholism drug, is known to elicit cytotoxicity in many cancer cell types in the presence of copper (Cu). The objective of the present study was to investigate the mechanism of action responsible for the induction of apoptosis by DSF/Cu and its effect on cancer stem cell properties in HER2-positive breast cancers in vitro and in vivo. DSF/Cu treatment induced apoptosis, associated with a marked decrease in HER2, truncated p95HER2, phospho-HER2, HER3, phospho-HER3 and phospho-Akt levels, and p27 nuclear accumulation. This was accompanied by the eradication of cancer stem-like populations, concomitant with the suppression of aldehyde dehydrogenase 1 (ALDH1) activity and mammosphere formation. DSF administration resulted in a significant reduction in tumor growth and an enhancement of apoptosis, as well as HER2 intracellular domain (ICD) and ALDH1A1 downregulation. Our results demonstrate that DSF/Cu induces apoptosis and eliminates cancer stem-like cells via the suppression of HER2/Akt signaling, suggesting that DSF may be potentially effective for the treatment of HER2-positive cancers.
Assuntos
Inibidores de Acetaldeído Desidrogenases/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Cobre/farmacologia , Dissulfiram/farmacologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor ErbB-2/metabolismo , Aldeído Desidrogenase/antagonistas & inibidores , Aldeído Desidrogenase/metabolismo , Família Aldeído Desidrogenase 1 , Animais , Neoplasias da Mama/enzimologia , Neoplasias da Mama/patologia , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Relação Dose-Resposta a Droga , Feminino , Humanos , Células MCF-7 , Camundongos Endogâmicos BALB C , Camundongos Nus , Células-Tronco Neoplásicas/enzimologia , Células-Tronco Neoplásicas/patologia , Fenótipo , Fosforilação , Retinal Desidrogenase , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
MicroRNAs (miRs) serve either as oncogenes or tumor-suppressor genes in tumor progression. MicroRNA-20b (miR20b) is known to be involved with the oncomirs of several types of cancers. However, in the present study we describe how miR-20b inhibits the proliferation, migration and invasion of bladder cancer EJ cells. In the present study, miR-20b was downregulated in bladder cancer cell lines, and its overexpression resulted in a significant reduction in the proliferation of EJ cells. In addition, via a bioinformatics approach, we identified cell cycle-regulated genes that are the putative targets of miR-20b. The transfection of miR-20b into EJ cells induced G1 phase cell cycle arrest via the decreased expression of cyclin D1, CDK2 and CDK6 without affecting another G1 phase cell cycle regulator, cyclin E. The cell cycle inhibitor p21WAF1 was upregulated in the miR-20b transfected cells. Moreover, the enforced expression of miR-20b resulted in impaired wound-healing migration and invasion in the EJ cells. Based on our target prediction analysis of miRs, we confirmed that miR-20b overexpression strongly impedes MMP-2 expression via suppressive activation of the Sp-1 binding motif, an important transcription factor present in the MMP-2 promoter. Herein, we report the novel concept that miR-20b exerts a suppressive effect on both cell cycle-modulated proliferation and MMP-2-mediated migration and invasion in bladder cancer EJ cells.
Assuntos
Pontos de Checagem do Ciclo Celular/genética , Metaloproteinase 2 da Matriz/genética , MicroRNAs/genética , Neoplasias da Bexiga Urinária/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Ciclina D1/biossíntese , Quinase 6 Dependente de Ciclina/biossíntese , Regulação Neoplásica da Expressão Gênica , Humanos , Invasividade Neoplásica/genética , Transfecção , Neoplasias da Bexiga Urinária/patologiaRESUMO
We identified a novel mechanism of AKT signaling in the fucoidan-induced proliferation and migration of human urinary 5637 cancer cells. Fucoidan treatment showed a significant growth inhibition followed by G1-phase-associated up-regulation of p21WAF1 expression and suppression of cyclins and CDK expression in 5637 cells. Also, fucoidan treatment induced the activation of AKT signaling, which was inhibited by treatment with wortmannin, a PI3K-specific inhibitor. Blockade of the AKT function reversed the fucoidan-mediated inhibition of cell proliferation, the increased G1-phase-associated p21WAF1 expression, and the reduction of cell-cycle proteins. Moreover, treatment with fucoidan blocked migration and invasion of 5637 cells. This inhibition was attributed to decreased expression of MMP-9, which was mediated by down-regulation of AP-1 and NF-κB binding activity. Furthermore, wortmannin treatment abolished the decreased cell migration and invasion and the inhibition of MMP-9 expression via the suppression of NF-κB and AP-1 in fucoidan-treated cells. Similar results were observed in another bladder cancer T-24 cells treated with fucoidan. Finally, overexpression of the AKT gene inhibited the proliferation, migration and invasion of bladder cancer cells. These data suggest that the activation of AKT signaling is involved in growth inhibition and suppression of the migration and invasion of bladder cancer cells treated with fucoidan.