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1.
Prostate ; 80(3): 256-266, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31856338

RESUMO

BACKGROUND: Because androgen receptor (AR) signaling is essential for prostate cancer (PCa) initiation and progression, castration is the main approach for treatment. Unfortunately, patients tend to enter a stage called castration-resistant prostate cancer (CRPC) despite the initial response to castration. For various reasons, AR signaling is reactivated in CRPC. As such, AR signaling inhibitors, such as enzalutamide, has been approved by the Food and Drug Administration to treat CRPC in the clinic. However, the limited success of these new drugs suggests an immediate unmet need to understand the underlying mechanisms for resistance so novel targets can be identified to enhance their efficacy. METHODS: An unbiased bioinformatics analysis was performed with the existing human patient dataset and RNA-seq results of in-house PCa cell lines to identify new targets to overcome enzalutamide resistance. Cell viability and growth were detected by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide and colony formation assay. Cell invasion and migration were detected by transwell assay. Protein levels were detected by Western blot or immunofluorescence. RESULTS: We found that the noncanonical Wnt signaling was activated in enzalutamide-resistant PCa cells and that the activation of noncanonical Wnt signaling was correlated with AR expression and disease progression. This was validated by the elevated expression of noncanonical Wnt pathway members such as Wnt5a, RhoA, and ROCK in enzalutamide-resistant PCa cells in comparison to their enzalutamide-sensitive counterparts. And, both Y27632, an inhibitor of ROCK, and depletion of ROCK enhanced the efficacy of enzalutamide in enzalutamide-resistant PCa cells. Of significance, a combination of Y27632 and enzalutamide inhibited 22RV1-derived xenograft tumor growth synergistically. Finally, ROCK depletion plus enzalutamide treatment inhibited invasion and migration of enzalutamide-resistant PCa cells via inhibition of epithelial-mesenchymal transition. CONCLUSIONS: The noncanonical Wnt pathway is activated in enzalutamide-resistant PCa and inhibition of noncanonical Wnt pathway overcomes enzalutamide resistance and enhances its efficacy in CRPC.


Assuntos
Amidas/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Feniltioidantoína/análogos & derivados , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Piridinas/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , Amidas/administração & dosagem , Animais , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos , Sinergismo Farmacológico , Humanos , Masculino , Camundongos , Feniltioidantoína/administração & dosagem , Feniltioidantoína/farmacologia , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Piridinas/administração & dosagem , Distribuição Aleatória , Receptores Androgênicos/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Quinases Associadas a rho/antagonistas & inibidores , Quinases Associadas a rho/metabolismo
2.
Mol Carcinog ; 59(1): 62-72, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31674708

RESUMO

Prostate cancer (PCa) deaths are typically the result of metastatic castration-resistant PCa (mCRPC). Recently, enzalutamide (Enz), an oral androgen receptor inhibitor, was approved for treating patients with mCRPC. Invariably, all PCa patients eventually develop resistance against Enz. Therefore, novel strategies aimed at overcoming Enz resistance are needed to improve the survival of PCa patients. The role of exosomes in drug resistance has not been fully elucidated in PCa. Therefore, we set out to better understand the exosome's role in the mechanism underlying Enz-resistant PCa. Results showed that Enz-resistant PCa cells (C4-2B, CWR-R1, and LNCaP) secreted significantly higher amounts of exosomes (2-4 folds) compared to Enz-sensitive counterparts. Inhibition of exosome biogenesis in resistant cells by GW4869 and dimethyl amiloride strongly decreased their cell viability. Mechanistic studies revealed upregulation of syntaxin 6 as well as its increased colocalization with CD63 in Enz-resistant PCa cells compared to Enz-sensitive cells. Syntaxin 6 knockdown by specific small interfering RNAs in Enz-resistant PCa cells (C4-2B and CWR-R1) resulted in reduced cell number and increased cell death in the presence of Enz. Furthermore, syntaxin 6 knockdown significantly reduced the exosome secretion in both Enz-resistant C4-2B and CWR-R1 cells. The Cancer Genome Atlas analysis showed increased syntaxin 6 expressions associated with higher Gleason score and decreased progression-free survival in PCa patients. Importantly, IHC analysis showed higher syntaxin 6 expression in cancer tissues from Enz-treated patients compared to Enz naïve patients. Overall, syntaxin 6 plays an important role in the secretion of exosomes and increased survival of Enz-resistant PCa cells.


Assuntos
Antineoplásicos/farmacologia , Exossomos/metabolismo , Feniltioidantoína/análogos & derivados , Neoplasias da Próstata/tratamento farmacológico , Proteínas Qa-SNARE/metabolismo , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Exossomos/efeitos dos fármacos , Humanos , Masculino , Feniltioidantoína/farmacologia , Neoplasias da Próstata/metabolismo
3.
Prostate ; 79(16): 1837-1851, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31524299

RESUMO

BACKGROUND: Advanced prostate cancers depend on protein synthesis for continued survival and accelerated rates of metabolism for growth. RNA polymerase I (Pol I) is the enzyme responsible for ribosomal RNA (rRNA) transcription and a rate-limiting step for ribosome biogenesis. We have shown using a specific and sensitive RNA probe for the 45S rRNA precursor that rRNA synthesis is increased in prostate adenocarcinoma compared to nonmalignant epithelium. We have introduced a first-in-class Pol I inhibitor, BMH-21, that targets cancer cells of multiple origins, and holds potential for clinical translation. METHODS: The effect of BMH-21 was tested in prostate cancer cell lines and in prostate cancer xenograft and mouse genetic models. RESULTS: We show that BMH-21 inhibits Pol I transcription in metastatic, castration-resistant, and enzalutamide treatment-resistant prostate cancer cell lines. The genetic abrogation of Pol I effectively blocks the growth of prostate cancer cells. Silencing of p53, a pathway activated downstream of Pol I, does not diminish this effect. We find that BMH-21 significantly inhibited tumor growth and reduced the Ki67 proliferation index in an enzalutamide-resistant xenograft tumor model. A decrease in 45S rRNA synthesis demonstrated on-target activity. Furthermore, the Pol I inhibitor significantly inhibited tumor growth and pathology in an aggressive genetically modified Hoxb13-MYC|Hoxb13-Cre|Ptenfl/fl (BMPC) mouse prostate cancer model. CONCLUSION: Taken together, BMH-21 is a novel promising molecule for the treatment of castration-resistant prostate cancer.


Assuntos
Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias da Próstata/tratamento farmacológico , RNA Polimerase I/antagonistas & inibidores , Animais , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Inibidores Enzimáticos/farmacologia , Humanos , Masculino , Camundongos , Camundongos Nus , Terapia de Alvo Molecular , Células PC-3 , Feniltioidantoína/análogos & derivados , Feniltioidantoína/farmacologia , Neoplasias da Próstata/enzimologia , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Neoplasias de Próstata Resistentes à Castração/enzimologia , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , RNA Polimerase I/genética , RNA Polimerase I/metabolismo , RNA Ribossômico/genética , Distribuição Aleatória , Transcrição Genética/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
4.
Nucleic Acids Res ; 47(19): 10104-10114, 2019 11 04.
Artigo em Inglês | MEDLINE | ID: mdl-31501863

RESUMO

Enzalutamide, a second-generation androgen receptor (AR) antagonist, has demonstrated clinical benefit in men with prostate cancer. However, it only provides a temporary response and modest increase in survival, indicating a rapid evolution of resistance. Previous studies suggest that enzalutamide may function as a partial transcriptional agonist, but the underlying mechanisms for enzalutamide-induced transcription remain poorly understood. Here, we show that enzalutamide stimulates expression of a novel subset of genes distinct from androgen-responsive genes. Treatment of prostate cancer cells with enzalutamide enhances recruitment of pioneer factor GATA2, AR, Mediator subunits MED1 and MED14, and RNA Pol II to regulatory elements of enzalutamide-responsive genes. Mechanistically, GATA2 globally directs enzalutamide-induced transcription by facilitating AR, Mediator and Pol II loading to enzalutamide-responsive gene loci. Importantly, the GATA2 inhibitor K7174 inhibits enzalutamide-induced transcription by decreasing binding of the GATA2/AR/Mediator/Pol II transcriptional complex, contributing to sensitization of prostate cancer cells to enzalutamide treatment. Our findings provide mechanistic insight into the future combination of GATA2 inhibitors and enzalutamide for improved AR-targeted therapy.


Assuntos
Fator de Transcrição GATA2/genética , Feniltioidantoína/análogos & derivados , Neoplasias da Próstata/tratamento farmacológico , Receptores Androgênicos/genética , Antagonistas de Receptores de Andrógenos/farmacologia , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Fator de Transcrição GATA2/antagonistas & inibidores , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Complexo Mediador/genética , Subunidade 1 do Complexo Mediador/genética , Feniltioidantoína/farmacologia , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , RNA Polimerase II/genética
5.
Eur J Med Chem ; 180: 1-14, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31288149

RESUMO

SAR studies on bicalutamide, enobosarm and enzalutamide analogues, functionalised with polyfluorinated groups, is presented. Among the novel bicalutamide and enobosarm derivatives synthesised, several displayed significantly improved in vitro anticancer activity, with IC50 values in the low micromolar range against four different prostate cancer cell lines (LNCaP, VCaP, DU-145 and 22Rv1), showing up to 48-fold increase in comparison with the parent structures. In particular, SF5 enobosarm analogues were found to be most potent compounds, full AR antagonists and with favourable ADME properties. The most promising compound (48a) was evaluated for its in vivo efficacy in PC xenograft mouse model (22Rv1) with results comparable to the standard-of-care docetaxel.


Assuntos
Anilidas/farmacologia , Antineoplásicos/farmacologia , Nitrilos/farmacologia , Feniltioidantoína/análogos & derivados , Neoplasias da Próstata/tratamento farmacológico , Compostos de Tosil/farmacologia , Anilidas/síntese química , Anilidas/química , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Células CHO , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cricetulus , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Canais de Potássio Éter-A-Go-Go/metabolismo , Humanos , Hidrocarbonetos Fluorados/química , Hidrocarbonetos Fluorados/farmacologia , Masculino , Camundongos , Camundongos Nus , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/patologia , Nitrilos/síntese química , Nitrilos/química , Feniltioidantoína/síntese química , Feniltioidantoína/química , Feniltioidantoína/farmacologia , Neoplasias da Próstata/patologia , Relação Estrutura-Atividade , Compostos de Sulfidrila/química , Compostos de Sulfidrila/farmacologia , Compostos de Tosil/síntese química , Compostos de Tosil/química
6.
Prostate ; 79(11): 1347-1359, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31228231

RESUMO

BACKGROUND: Prostate cancer that recurs after initial treatment inevitably progresses to castration-resistant prostate cancer (CRPC), the lethal stage of the disease. Despite improvements in outcomes from next generation androgen receptor (AR)-axis inhibitors, CRPC remains incurable. Therapeutic strategies to target AR antagonist resistance are urgently needed to improve outcomes for men with this lethal form of prostate cancer. METHODS: Apoptosis and BCL2 family signaling were characterized in cell line models of CRPC. Quantitative real-time polymerase chain reaction and Western blot analysis were used to determine BCL2 expression levels. Drug sensitivity was determined by proliferation, survival and apoptosis analysis. Protein-protein interactions were evaluated by coimmunoprecipitation followed by Western blot detection. RESULTS: In the present study, we identify antiapoptotic BCL2 protein signaling as a mechanism of resistance to AR antagonist enzalutamide. In CRPC cell line models, we found that BCL-xL and MCL-1 proteins block apoptosis through binding and sequestering proapoptotic proteins BIM and BAX, resulting in cell survival in response to enzalutamide. Treatment with BH3-mimetics targeting BCL-xL or MCL-1 disrupts these interactions and activates apoptosis, sensitizing CRPC cells to enzalutamide. Importantly, we demonstrate that PI3K/Akt signaling is activated in response to enzalutamide and mediates apoptosis evasion through inactivation of BAD, a BH3-only protein that activates proapoptotic signlaing through inhbition of BCL-xL. Inhibition of Akt activates BAD, resulting in increased apoptosis and sensitivity to enzalutamide, demonstrating an alternative therapeutic strategy to target drug resistance. CONCLUSIONS: These results demonstrate that CRPC cells employ multiple mechanisms to mediate apoptosis evasion through BCL2 signaling, suggesting this pathway is critical for survival. This study provides a strong preclinical rationale for developing therapeutic strategies to target antiapoptotic BCL2 signaling in combination with AR antagonists to improve treatment options for patients with advanced prostate cancer.


Assuntos
Antagonistas de Androgênios/uso terapêutico , Feniltioidantoína/análogos & derivados , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Antagonistas de Androgênios/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Humanos , Masculino , Feniltioidantoína/farmacologia , Feniltioidantoína/uso terapêutico , Fosfatidilinositol 3-Quinases/metabolismo , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo
7.
Prostate ; 79(10): 1166-1179, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31135075

RESUMO

BACKGROUND: The progression of castration-resistant prostate cancer (CRPC) still relies on the function of androgen receptor (AR), achieved by evolving mechanisms to reactivate AR signaling under hormonal therapy. Histone deacetylase inhibitors (HDACis) disrupt cytoplasmic AR chaperone heat shock protein 90 (Hsp90) via HDAC6 inhibition, leading to AR degradation and growth suppression of prostate cancer (PCa) cells. However, current HDACis are not effective in clinical trials treating CRPC. METHODS: We designed hybrid molecules containing partial chemical scaffolds of AR antagonist enzalutamide (Enz) and HDACi suberoylanilide hydroxamic acid (SAHA) as new anti-PCa agents. We previously demonstrated that Enz-HDACi hybrid drug 2-75 targets both AR and Hsp90, which inhibits the growth of Enz-resistant C4-2 cells. In the current study, we further investigate the molecular and cellular actions of 2-75 and test its anti-PCa effects in vivo. RESULTS: Compared with Enz, 2-75 had greater AR antagonistic effects by decreasing the stability, transcriptional activity, and nuclear translocation of intracellular AR. In addition to inhibition of full-length AR (FL AR), 2-75 downregulated the AR-V7 variant in multiple PCa cell lines. Mechanistic studies indicated that the AR affinity of 2-75 retains the drug in the cytoplasm of AR + PCa cells and further directs 2-75 to the AR-associated protein complex, which permits localized effects on AR-associated Hsp90. Further, unlike pan-HDACi SAHA, the cytoplasm-retaining property allows 2-75 to significantly inhibit cytoplasmic HDAC6 with limited impact on nuclear HDACs. These selective cytoplasmic actions of 2-75 overcome the unfavorable resistance and toxicity properties associated with classical AR antagonists, HDACis, and Hsp90 inhibitors. Finally, 2-75 showed greater antitumor activities than Enz in vivo on SQ xenografts derived from LNCaP cells. CONCLUSIONS: Novel therapeutic strategy using newly designed 2-75 and related AR antagonist-HDACi hybrid drugs has great potential for effective treatment of CRPC.


Assuntos
Antineoplásicos/uso terapêutico , Proliferação de Células/efeitos dos fármacos , Inibidores de Histona Desacetilases/uso terapêutico , Feniltioidantoína/análogos & derivados , Próstata/efeitos dos fármacos , Neoplasias da Próstata/tratamento farmacológico , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Regulação para Baixo , Inibidores de Histona Desacetilases/farmacologia , Humanos , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Feniltioidantoína/farmacologia , Feniltioidantoína/uso terapêutico , Próstata/patologia , Neoplasias da Próstata/patologia
8.
Prostate ; 79(10): 1147-1155, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31077419

RESUMO

BACKGROUND: Cigarette smoking is associated with worse outcomes in prostate cancer, whose growth is dependent on androgen receptor (AR) signaling. We aimed to elucidate the biological effect of cigarette smoking on AR signaling and its clinical influence on oncological outcome. METHODS: Gene expression levels after exposure to tobacco smoke condensate (TSC) were evaluated by quantitative real-time polymerase chain reaction and Western blot analysis in prostate cancer cells. Cellular sensitivities to enzalutamide and docetaxel after TSC exposure were evaluated using a prostate cancer cell proliferation assay. Prognosis was compared between current smokers and nonsmokers when treated with AR-axis-targeting (ARAT) agent enzalutamide and docetaxel. RESULTS: Expression of AR variants as well as prostate-specific antigen was augmented after TSC exposure, which occurred after Akt phosphorylation. These inductions were suppressed by Akt inhibitor LY294002 as well as antioxidant N-acetylcysteine. Consistently, TSC exposure augmented cellular resistance to enzalutamide. In clinical data, cigarette smoking was associated with worse progression-free survival and cancer-specific survival when patients with prostate cancer were treated with ARAT agents but not docetaxel. CONCLUSIONS: It was suggested that cigarette smoking leads to detrimental oncological outcome when prostate cancer patients are treated with ARAT agents through induction of aberrant AR signaling. Accordingly, we recommend that patients with advanced prostate cancer should refrain from cigarette smoking.


Assuntos
Antagonistas de Androgênios/farmacologia , Fumar Cigarros/patologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias da Próstata/patologia , Receptores Androgênicos/metabolismo , Antagonistas de Androgênios/uso terapêutico , Androstenos/farmacologia , Androstenos/uso terapêutico , Linhagem Celular Tumoral , Fumar Cigarros/metabolismo , Humanos , Masculino , Feniltioidantoína/análogos & derivados , Feniltioidantoína/farmacologia , Feniltioidantoína/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/metabolismo
9.
J Exp Clin Cancer Res ; 38(1): 220, 2019 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-31126320

RESUMO

BACKGROUND: Androgen receptor (AR) is expressed in approximately 70% of breast tumors. Recent studies increasingly support AR as a potential therapeutic target of AR-positive breast cancer. We have previously reported that deubiquitinase USP14 stabilizes AR proteins by deubiquitination and USP14 inhibition results in inhibition of cell growth and tumor progression in AR-positive prostate cancer and breast cancer. The current study aims to explore the anticancer effect of a treatment combining AR antagonist enzalutamide with USP14 inhibition on breast cancer cells. METHODS: The combining effects of enzalutamide and USP14 inhibition on breast cancer cell proliferation and apoptosis and associated cell signaling were evaluated in vitro and in vivo. RESULTS: USP14 inhibition via administration of IU1 or USP14-specific siRNA/shRNA enhanced cell growth inhibition and apoptosis induction by enzalutamide in breast cancer cell lines in vitro and in vivo. Additionally, the combination of enzalutamide with USP14 inhibition/knockdown induced significant downregulation of AR proteins and suppression of AR-related signaling pathways, including Wnt/ß-catenin and PI3K/AKT pathways. Moreover, AKT inhibition via MK2206 increased the antiproliferative and proapoptotic effects of enzalutamide+IU1 combined treatment. CONCLUSION: Collectively, our data suggest that USP14 inhibition in combination with enzalutamide represents a potentially new therapeutic strategy for breast cancer.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feniltioidantoína/análogos & derivados , Pirróis/administração & dosagem , Pirrolidinas/administração & dosagem , Ubiquitina Tiolesterase/metabolismo , Animais , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Regulação para Baixo , Sinergismo Farmacológico , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Compostos Heterocíclicos com 3 Anéis/administração & dosagem , Compostos Heterocíclicos com 3 Anéis/farmacologia , Humanos , Células MCF-7 , Camundongos , Feniltioidantoína/administração & dosagem , Feniltioidantoína/farmacologia , Pirróis/farmacologia , Pirrolidinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Ubiquitina Tiolesterase/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Oncogene ; 38(28): 5700-5724, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31043708

RESUMO

Androgen receptor (AR) signalling is a key prostate cancer (PC) driver, even in advanced 'castrate-resistant' disease (CRPC). To systematically identify microRNAs (miRs) modulating AR activity in lethal disease, hormone-responsive and -resistant PC cells expressing a luciferase-based AR reporter were transfected with a miR inhibitor library; 78 inhibitors significantly altered AR activity. Upon validation, miR-346, miR-361-3p and miR-197 inhibitors markedly reduced AR transcriptional activity, mRNA and protein levels, increased apoptosis, reduced proliferation, repressed EMT, and inhibited PC migration and invasion, demonstrating additive effects with AR inhibition. Corresponding miRs increased AR activity through a novel and anti-dogmatic mechanism of direct association with AR 6.9 kb 3'UTR and transcript stabilisation. In addition, miR-346 and miR-361-3p modulation altered levels of constitutively active AR variants, and inhibited variant-driven PC cell proliferation, so may contribute to persistent AR signalling in CRPC in the absence of circulating androgens. Pathway analysis of AGO-PAR-CLIP-identified miR targets revealed roles in DNA replication and repair, cell cycle, signal transduction and immune function. Silencing these targets, including tumour suppressors ARHGDIA and TAGLN2, phenocopied miR effects, demonstrating physiological relevance. MiR-346 additionally upregulated the oncogene, YWHAZ, which correlated with grade, biochemical relapse and metastasis in patients. These AR-modulatory miRs and targets correlated with AR activity in patient biopsies, and were elevated in response to long-term enzalutamide treatment of patient-derived CRPC xenografts. In summary, we identified miRs that modulate AR activity in PC and CRPC, via novel mechanisms, and may represent novel PC therapeutic targets.


Assuntos
MicroRNAs/fisiologia , Neoplasias da Próstata/tratamento farmacológico , Receptores Androgênicos/fisiologia , Regiões 3' não Traduzidas , Elementos Antissenso (Genética) , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Transição Epitelial-Mesenquimal , Humanos , Masculino , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Invasividade Neoplásica , Metástase Neoplásica , Feniltioidantoína/análogos & derivados , Feniltioidantoína/farmacologia , Neoplasias da Próstata/patologia , Transdução de Sinais
12.
J Biol Chem ; 294(21): 8543-8554, 2019 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-30940724

RESUMO

Prostate cancer is the second leading cause of cancer death among men in the United States. The androgen receptor (AR) antagonist enzalutamide is a Food and Drug Administration-approved drug for treatment of patients with late-stage prostate cancer and is currently under clinical study for early-stage prostate cancer treatment. After a short positive response period, tumors will develop drug resistance. In this study using RNA-Seq and bioinformatics analyses, we observed that NOTCH signaling is a deregulated pathway in enzalutamide-resistant cells. NOTCH2 and c-MYC gene expression positively correlated with AR expression in samples from patient with hormone refractory disease in which AR expression levels correspond to those typically observed in enzalutamide resistance. Cleaved NOTCH1, HES1 (Hes family BHLH transcription factor 1), and c-MYC protein expression levels are elevated in two enzalutamide-resistant cell lines, MR49F and C4-2R, indicating NOTCH signaling activation. Moreover, inhibition of the overexpressed ADAM metallopeptidase domain 10 (ADAM10) in the resistant cells induces an exclusive reduction in cleaved NOTCH1 expression. Furthermore, exposure of enzalutamide-resistant cells to both PF-03084014 and enzalutamide increased cell death, decreased colony formation ability, and resensitized cells to enzalutamide. Knockdown of NOTCH1 in C4-2R increased enzalutamide sensitivity by decreasing cell proliferation and increasing cleaved PARP expression. In a 22RV1 xenograft model, PF-03084014 and enzalutamide decreased tumor growth through reducing cell proliferation and increasing apoptosis. These results indicate that NOTCH1 signaling may contribute to enzalutamide resistance in prostate cancer, and inhibition of NOTCH signaling can resensitize resistant cells to enzalutamide.


Assuntos
Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feniltioidantoína/análogos & derivados , Receptor Notch1/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tetra-Hidronaftalenos/farmacologia , Valina/análogos & derivados , Animais , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Masculino , Camundongos , Camundongos Nus , Feniltioidantoína/farmacologia , Neoplasias da Próstata , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Receptor Notch1/genética , Receptor Notch2/genética , Receptor Notch2/metabolismo , Transdução de Sinais/genética , Fatores de Transcrição HES-1/genética , Fatores de Transcrição HES-1/metabolismo , Valina/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
13.
J Exp Clin Cancer Res ; 38(1): 157, 2019 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-30975171

RESUMO

BACKGROUND: Prostate cancer (PCa) remains a challenge worldwide. Due to the development of castration-resistance, traditional first-line androgen deprivation therapy (ADT) became powerlessness. Epidermal growth factor receptor (EGFR) is a well characterized therapeutic target to treat colorectal carcinoma and non-small cell lung cancer. Increasing studies have unraveled the significance of EGFR and its downstream signaling in the progression of castration-resistant PCa. METHOD: MTS, colony formation and Edu staining assays were used to analyze the cell proliferation of PCa cells. Flow cytometry was used to analyze PCa cell cycle distribution and cell apoptosis. Western blot was used to measure the expression of key proteins associated with cell cycle progression, apoptosis and EGFR signaling pathways. Transfection of exogenous small interfering RNA (siRNA) or plasmid was used to intervene specific gene expression. Nude mouse model was employed to test the in vivo effect of Spautin-1. RESULTS: The current study reveals that Spautin-1, a known inhibitor of ubiquitin-specific peptidase 10 (USP10) and USP13, inhibits EGFR phosphorylation and the activation of its downstream signaling. Inhibition of EGFR signaling induced by Spautin-1 leads to cell cycle arrest and apoptosis of PCa in a USP10/USP13 independent manner. The application of Spautin-1 reduces the expression of glucose transporter 1 (Glut1) and dramatically induces cell death under glucose deprivation condition. In vivo experiments show a potent anti-tumor effect of Spautin-1 alone and in combination with Enzalutamide. CONCLUSION: This study demonstrates the therapeutic potential of EGFR signaling inhibition by the use of Spautin-1 for PCa treatment.


Assuntos
Benzilaminas/farmacologia , Neoplasias da Próstata/metabolismo , Quinazolinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Biomarcadores Tumorais , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ciclina D1/genética , Modelos Animais de Doenças , Receptores ErbB/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glucose/metabolismo , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Masculino , Camundongos , Modelos Biológicos , Terapia de Alvo Molecular , Feniltioidantoína/análogos & derivados , Feniltioidantoína/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
14.
PLoS One ; 14(4): e0214670, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30933998

RESUMO

Radiation therapy is often combined with androgen deprivation therapy in the treatment of aggressive localized prostate cancer. However, castration-resistant disease may not respond to testosterone deprivation approaches. Enzalutamide is a second-generation anti-androgen with high affinity and activity that is used for the treatment of metastatic disease. Although radiosensitization mechanisms are known to be mediated through androgen receptor activity, this project aims to uncover the detailed DNA damage repair factors influenced by enzalutamide using multiple models of androgen-sensitive (LNCaP) and castration-resistant human prostate cancer (22Rv1 and DU145). Enzalutamide is able to radiosensitize both androgen-dependent and androgen-independent human prostate cancer models in cell culture and xenografts in mice, as well as a treatment-resistant patient-derived xenograft. The enzalutamide-mediated mechanism of radiosensitization includes delay of DNA repair through temporal prolongation of the repair factor complexes and halting the cell cycle, which results in decreased colony survival. Altogether, these findings support the use of enzalutamide concurrently with radiotherapy to enhance the treatment efficacy for prostate cancer.


Assuntos
Reparo do DNA/efeitos dos fármacos , Feniltioidantoína/análogos & derivados , Neoplasias da Próstata , Tolerância a Radiação , Radiossensibilizantes/farmacologia , Idoso , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/genética , Reparo do DNA/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Resistencia a Medicamentos Antineoplásicos/efeitos da radiação , Humanos , Masculino , Camundongos , Camundongos Nus , Camundongos Transgênicos , Feniltioidantoína/farmacologia , Feniltioidantoína/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Neoplasias da Próstata/radioterapia , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Neoplasias de Próstata Resistentes à Castração/radioterapia , Tolerância a Radiação/efeitos dos fármacos , Tolerância a Radiação/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Oncogene ; 38(28): 5658-5669, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30996246

RESUMO

BET bromodomain inhibitors block prostate cancer cell growth at least in part through c-Myc and androgen receptor (AR) suppression. However, little is known about other transcriptional regulators whose suppression contributes to BET bromodomain inhibitor anti-tumor activity. Moreover, the anti-tumor activity of BET bromodomain inhibition in AR-independent castration-resistant prostate cancers (CRPC), whose frequency is increasing, is also unknown. Herein, we demonstrate that BET bromodomain inhibition blocks growth of a diverse set of CRPC cell models, including those that are AR-independent or in which c-Myc is not suppressed. To identify transcriptional regulators whose suppression accounts for these effects, we treated multiple CRPC cell lines with the BET bromodomain inhibitor JQ1 and then performed RNA-sequencing followed by Master Regulator computational analysis. This approach identified several previously unappreciated transcriptional regulators that are highly expressed in CRPC and whose suppression, via both transcriptional or post-translational mechanisms, contributes to the anti-tumor activity of BET bromodomain inhibitors.


Assuntos
Proteínas de Ciclo Celular/antagonistas & inibidores , Neoplasias de Próstata Resistentes à Castração/metabolismo , Receptores Androgênicos/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Animais , Azepinas/farmacologia , Proteínas de Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Proteínas Cromossômicas não Histona/fisiologia , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Masculino , Camundongos , Camundongos SCID , Feniltioidantoína/análogos & derivados , Feniltioidantoína/farmacologia , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Biossíntese de Proteínas , Fatores de Transcrição/fisiologia , Transcrição Genética , Triazóis/farmacologia
16.
Int J Mol Sci ; 20(8)2019 Apr 13.
Artigo em Inglês | MEDLINE | ID: mdl-31013891

RESUMO

Currently, there are two Food and Drug Administration (FDA)-approved drugs for androgen deprivation therapy (ADT) of metastatic castration-resistant prostate cancer (mCRPC) patients: abiraterone and enzalutamide. However, our understanding of the effect of these therapies on the immune system in mCRPC patients remains limited. Here, we examined how abiraterone and enzalutamide treatment affects levels of soluble immune mediators in plasma and in circulating immune cells of 44 mCRPC patients. We found that the baseline levels of cytokines fibroblast growth factor (FGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin 10 (IL-10), and IL-6 were significantly lower in ADT-sensitive compared to de novo resistant patients. In addition, resistant patients showed significantly lower T cell frequencies. When comparing the levels of cytokines over the course of treatment, we observed that the levels of proinflammatory mediators, such as interferon-γ (IFN-γ), IL-5, macrophage inflammatory protein 1 alpha (MIP-1α), and tumor necrosis factor alpha (TNFα), were significantly increased in the ADT-sensitive patients. At the same time, the abiraterone/enzalutamide therapy did not reduce the percentage of tolerogenic myeloid cell populations, such as polymorphonuclear myeloid-derived suppressor cells, which retained unaltered expression of programmed death-ligand 1 (PD-L1) and B7-H3. Overall, our results suggest that certain immune markers, such as IL-6 and the frequency of effector T cells, could be predictive of therapeutic response to ADT therapies in mCRPC patients.


Assuntos
Androstenos/farmacologia , Citocinas/metabolismo , Resistencia a Medicamentos Antineoplásicos , Imunomodulação/efeitos dos fármacos , Neoplasias da Próstata/etiologia , Neoplasias da Próstata/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Idoso , Idoso de 80 Anos ou mais , Antagonistas de Androgênios/uso terapêutico , Biomarcadores Tumorais , Linhagem Celular Tumoral , Citocinas/sangue , Humanos , Contagem de Linfócitos , Masculino , Pessoa de Meia-Idade , Células Supressoras Mieloides/imunologia , Células Supressoras Mieloides/metabolismo , Feniltioidantoína/análogos & derivados , Feniltioidantoína/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia
17.
Oncol Rep ; 41(5): 2689-2702, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30864728

RESUMO

Castration­resistant prostate cancer (CRPC) is a major challenge in the treatment of prostate cancer (PCa). Phospholipase Cε (PLCε), an oncogene, has been found to be involved in the carcinogenesis, tumor proliferation and migration of several types of cancer. The effects, however, of PLCε on CRPC remains unclear. In the present study, the expression of PLCε and glioma­associated homolog (Gli)­1/Gli­2 in benign prostatic hyperplasia (BPH), PCa and CRPC tissues and cells was investigated, and the correlations between PLCε and Gli­1/Gli­2 in CRPC tissues and cell lines were further explored. In addition, the effect of PLCε on cell proliferation and invasion was assessed in CRPC cell lines, and the sensitivity of EN­R and 22RV1 cells to enzalutamide following the downregulation of PLCε expression was determined using lentivirus­mediated shPLCε and/or treatment with specific Gli inhibitor GANT61. It was found that the PLCε expression was excessively upregulated in the majority of CRPC tissues, and PLCε positivity was linked to poor progression­free survival (PFS) and overall survival (OS) in patients with PCa. Furthermore, PLCε knockdown significantly suppressed CRPC cell proliferation and invasion. Of note, it was found that PLCε knockdown increased the sensitivity of CRPC cells to enzalutamide in vitro by suppressing androgen receptor (AR) activities via the non­canonical Hedgehog/Gli­2 and p­STAT3 signaling pathways. PLCε knockdown was shown to increase the sensitivity of CRPC cell xenografts to enzalutamide in vivo. Finally, the combination of PLCε knockdown with GANT61 significantly sensitized CRPC cells to enzalutamide. Collectively, the results of the present study suggest that PLCε is a potential therapeutic target for CRPC.


Assuntos
Feniltioidantoína/análogos & derivados , Fosfoinositídeo Fosfolipase C/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/metabolismo , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Técnicas de Silenciamento de Genes , Humanos , Imunoglobulina G/farmacologia , Imunoglobulina G/uso terapêutico , Masculino , Melfalan/farmacologia , Melfalan/uso terapêutico , Camundongos , Camundongos Nus , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/metabolismo , Feniltioidantoína/farmacologia , Feniltioidantoína/uso terapêutico , Fosfoinositídeo Fosfolipase C/genética , Prognóstico , Intervalo Livre de Progressão , Próstata/patologia , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/mortalidade , Piridinas/farmacologia , Piridinas/uso terapêutico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , RNA Interferente Pequeno/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Análise de Sobrevida , Regulação para Cima , Ensaios Antitumorais Modelo de Xenoenxerto , Proteína GLI1 em Dedos de Zinco/antagonistas & inibidores , Proteína GLI1 em Dedos de Zinco/metabolismo , Proteína Gli2 com Dedos de Zinco/antagonistas & inibidores , Proteína Gli2 com Dedos de Zinco/metabolismo
18.
Prostate ; 79(6): 657-666, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30714180

RESUMO

BACKGROUND: DHX15 is a member of the DEAH-box (DHX) RNA helicase family. Our previous study identified it as an AR coactivator which contributes to prostate cancer progression. METHODS: We investigated DHX15 expression in castration resistant prostate cancer specimens and the influence of DHX15 on the responsiveness of prostate cancer cells to DHT stimulation. We also explored the role DHX15 played in enzalutamide resistance and the interacting domain in DHX15 with AR. DHX15 expression level in human CRPC specimens and prostate cancer specimens was detected by tissue microarray (TMA) immunostaining analysis. Colony formation assay was performed to determine the proliferation of cells treated with enzalutamide or DHT. siRNAs were used to knockdown DHX15. The interactions between DHX15 and AR were detected using co-immunoprecipitation assay. RESULTS: The expression level of DHX15 was upregulated in human CRPC specimens compared with hormone naïve prostate cancer specimens. DHX15 knockdown reduced AR sensitivity to low DHT concentrations in C4-2 cells. Inactivation of DHX15 sensitizes the enzalutamide treatment in C4-2 cells. Deletion mutagenesis indicated that DHX1 5 interacts with AR through its N terminal domain. CONCLUSIONS: These findings suggest that DHX15 contributes to prostate cancer progression. DHX15 is required for androgen receptor sensitivity to low DHT concentrations and contributes to enzalutamide resistance in C4-2 cells. Targeting DHX15 may improve the ADT treatment.


Assuntos
Feniltioidantoína/análogos & derivados , Neoplasias de Próstata Resistentes à Castração , RNA Helicases , Receptores Androgênicos/metabolismo , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Imunoprecipitação/métodos , Masculino , Feniltioidantoína/farmacologia , Próstata/metabolismo , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , RNA Helicases/genética , RNA Helicases/metabolismo , Ativação Transcricional , Regulação para Cima
19.
Int J Cancer ; 145(7): 1970-1981, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-30807643

RESUMO

TMPRSS2-ERG expression in blood has been correlated with low docetaxel benefit in metastatic castration-resistant prostate cancer (mCRPC). This multicenter study aimed to prospectively asses its role as a taxane-resistance biomarker in blood and retrospectively in tumors, exploring also the impact of prior abiraterone/enzalutamide (A/E) in patients and in vitro. TMPRSS2-ERG was tested by quantitative reverse-transcription PCR. We included 204 patients (137 blood and 124 tumor samples) treated with taxanes. TMPRSS2-ERG expression was correlated with prostate-specific antigen (PSA)-progression-free survival (PFS), radiological-PFS (RX-PFS), and overall survival (OS). Independent association with survival was evaluated by multivariate Cox modeling. In vitro ERG knockdown and combinatorial and sequential experiments with enzalutamide and docetaxel were performed in VCaP cells. Prior A/E (HR 1.8, 95% CI 1.2-2.8) and blood TMPRSS2-ERG detection (HR 2, 95% CI 1.1-3.7) were independently associated to lower PSA-PFS. In patients without prior A/E, blood and tumor TMPRSS2-ERG independently predicted lower PSA-PFS (HR 3.3, 95% CI 1.4-7.9 and HR 1.8, 95% CI 1.02-3.3, respectively) to taxanes. When prior A/E was administered, TMPRSS2-ERG was not associated with outcome. There was a significant interaction between blood TMPRSS2-ERG and prior A/E related to PSA-PFS (p = 0.032) and RX-PFS (p = 0.009). In vitro stable ERG inhibition did not sensitize VCaP cells to docetaxel. Concomitant enzalutamide and taxanes were synergistic, but prior enzalutamide reduced docetaxel cytotoxicity in VCaP cells. Enzalutamide induced the expression of neuroendocrine markers and reduced that of E-cadherin. We conclude that prior hormone-therapy may influence taxanes response and TMPRSS2-ERG prognostic value. Thus, multiple and sequential biomarkers are needed in CRPC follow-up evaluation.


Assuntos
Biomarcadores Tumorais/genética , Docetaxel/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteínas de Fusão Oncogênica/genética , Feniltioidantoína/análogos & derivados , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Biomarcadores Tumorais/sangue , Hidrocarbonetos Aromáticos com Pontes , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Intervalo Livre de Doença , Sinergismo Farmacológico , Técnicas de Inativação de Genes , Humanos , Masculino , Proteínas de Fusão Oncogênica/sangue , Feniltioidantoína/farmacologia , Neoplasias de Próstata Resistentes à Castração/sangue , Neoplasias de Próstata Resistentes à Castração/genética , Estudos Retrospectivos , Taxoides , Regulador Transcricional ERG/genética
20.
Int J Biol Sci ; 15(3): 522-532, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30745839

RESUMO

The dysregulation of cyclin D -Cyclin-dependent kinase 4/6 (CDK4/6)-Rb axis has been implicated in breast cancer progression and the selective CDK4/6 inhibitors have shown effective activity in advanced breast cancer, especially in tumors driven by the estrogen receptor (ER). However, resistance to these small molecular inhibitors has become an inevitable clinical issue after their initial use. Here, we investigated the potential mechanism of resistance by establishing a CDK4/6 inhibitor palbociclib-resistant breast cancer cell line (MCF-7pR). After prolonged exposure to palbociclib, we detected the loss of the ER signaling and an increase in androgen receptor (AR). Moreover, we demonstrated more localization of AR in the cell nucleus of MCF-7pR compared to the parental cell (MCF-7). We also reported that AR could promote the progression of the cell cycle. Blockade of AR signaling could reduce the level of the relative G1-S cyclins, abolish Rb phosphorylation and inhibit the activation of transcriptional programs in S phase. Furthermore, dual inhibition of AR and CDK4/6 could reverse the resistance of palbociclib both in vitro and in vivo. In sum, our studies provide evidence that AR activation promotes cell cycle progression and cell proliferation in CDK4/6 inhibitor resistance, and identify AR inhibition as a putative novel therapeutic strategy to treat CDK4/6 inhibitor resistance in cancer.


Assuntos
Neoplasias da Mama/metabolismo , Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Quinase 4 Dependente de Ciclina/metabolismo , Quinase 6 Dependente de Ciclina/metabolismo , Piperazinas/farmacologia , Piridinas/farmacologia , Receptores Androgênicos/isolamento & purificação , Animais , Western Blotting , Quinase 4 Dependente de Ciclina/genética , Quinase 6 Dependente de Ciclina/genética , Feminino , Humanos , Imuno-Histoquímica , Imunoprecipitação , Células MCF-7 , Camundongos , Feniltioidantoína/análogos & derivados , Feniltioidantoína/farmacologia , Receptores Androgênicos/genética
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