RESUMEN
Breast cancer has the highest incidence and mortality in women worldwide. There are 70% of breast cancers considered as estrogen receptor α (ERα) positive. Therefore, the ERα-targeted therapy has become one of the most effective solution for patients with breast cancer. Whereas a better understanding of ERα regulation is critical to shape evolutional treatments for breast cancer. By exploring the regulatory mechanisms of ERα at levels of post-translational modifications, we identified the deubiquitinase USP15 as a novel protector for preventing ERα degradation and a critical driver for breast cancer progression. Specifically, we demonstrated that USP15 promoted the proliferation of ERα+, but not ERα- breast cancer, in vivo and in vitro. Meanwhile, USP15 knockdown notably enhanced the antitumor activities of tamoxifen on breast cancer cells. Importantly, USP15 knockdown induced the downregulation of ERα protein via promoting its K48-linked ubiquitination, which is required for proliferative inhibition of breast cancer cells. These findings not only provide a novel treatment for overcoming resistance to endocrine therapy, but also represent a therapeutic strategy on ERα degradation by targeting USP15-ERα axis.
Asunto(s)
Neoplasias de la Mama/metabolismo , Receptor alfa de Estrógeno/metabolismo , Proteasas Ubiquitina-Específicas/metabolismo , Neoplasias de la Mama/genética , Línea Celular Tumoral , Progresión de la Enfermedad , Receptor alfa de Estrógeno/genética , Femenino , Humanos , Células MCF-7 , Transducción de Señal , Transfección , UbiquitinaciónRESUMEN
Prostate cancer (PC) is the second most common cancer with limited treatment option in males. Although the reactivation of embryonic signals in adult cells is one of the characteristics of cancer, the underlying protein degradation mechanism remains elusive. Here, we show that the molecular chaperone GRP75 is a key player in PC cells by maintaining the protein stability of SIX1, a transcription factor for embryonic development. Mechanistically, GRP75 provides a platform to recruit the deubiquitinating enzyme USP1 to inhibit K48-linked polyubiquitination of SIX1. Structurally, the C-terminus of GRP75 (433-679 aa) contains a peptide binding domain, which is required for the formation of GRP75-USP1-SIX1 protein complex. Functionally, pharmacological or genetic inhibition of the GRP75-USP1-SIX1 protein complex suppresses tumor growth and overcomes the castration resistance of PC cells in vitro and in xenograft mouse models. Clinically, the protein expression of SIX1 in PC tumor tissues is positively correlated with the expression of GRP75 and USP1. These new findings not only enhance our understanding of the protein degradation mechanism, but also may provide a potential way to enhance the anti-cancer activity of androgen suppression therapy.
Asunto(s)
Proteínas HSP70 de Choque Térmico/genética , Proteínas de Homeodominio/genética , Proteínas Mitocondriales/genética , Neoplasias de la Próstata Resistentes a la Castración/genética , Proteasas Ubiquitina-Específicas/genética , Animales , Castración , Línea Celular Tumoral , Proliferación Celular/genética , Progresión de la Enfermedad , Regulación Neoplásica de la Expresión Génica/genética , Células HEK293 , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Células PC-3 , Próstata/patología , Neoplasias de la Próstata Resistentes a la Castración/patología , Proteolisis , Receptores Androgénicos/genética , Transducción de Señal/genética , Factores de Transcripción/genética , Ubiquitinación/genéticaRESUMEN
Foam cell formation process is involved in the pathogenesis of atherosclerosis (AS). Activation of this biological process depends on lipid uptake by scavenger receptors, such as CD36, SR-A and SR-B1. Among these receptors, CD36 is the principal one because it dominates roughly 50% lipid uptake in monocytes. In this study, our western blotting and RT-qPCR assays revealed that USP10 inhibition promotes the degradation of CD36 protein but does not change its mRNA level. In addition, Co-IP results showed that USP10 interacts with CD36 and stabilizes CD36 protein by cleaving poly-ubiquitin on CD36. Significantly, USP10 promotes foam cell formation. Immunofluorescence and Oil red O staining assays show that inhibition or knockdown of USP10 suppresses lipid uptake and foam cell formation by macrophages. In conclusion, USP10 promotes the development and progression of atherosclerosis through stabilizing CD36 protein expression. The regulation of USP10-CD36 may provide a significant therapeutic scheme in atherosclerosis.
Asunto(s)
Antígenos CD36/metabolismo , Células Espumosas/enzimología , Lipoproteínas LDL/metabolismo , Macrófagos/enzimología , Ubiquitina Tiolesterasa/metabolismo , Línea Celular , Humanos , Ubiquitina Tiolesterasa/genéticaRESUMEN
Rationale: Androgen receptor splice variant 7 (AR-V7) is a leading cause of the development of castration-resistant prostate cancer (CRPC). However, the regulation and function of AR-V7 at levels of post-translational modifications in prostate cancer therapy remain poorly understood. Here, we conducted a library screen of natural products to identify potential small molecules responsible for AR-V7 protein degradation in human prostate cancer cell lines. Methods: A natural product library was used to screen the inhibitor of AR-V7. Co-IP and biomass spectrum assays were used to identify the AR-V7-interacting proteins, whereas western blot, confocal microscopy, RNA interfering, and gene transfection were used to validate these interactions. Cell viability, EDU staining, and colony formation assays were employed to detect cell growth and proliferation. Flowcytometry assays were used to detect the distribution of cell cycle. Mouse xenograft models were used to study the anti-CRPC effects in vivo. Results: This screen identified rutaecarpine, one of the major components of the Chinese medicine Evodia rutaecarpa, as a novel chemical that selectively induces AR-V7 protein degradation via K48-linked ubiquitination. Mechanically, this effect relies on rutaecarpine inducing the formation of a GRP78-AR-V7 protein complex, which further recruits the E3 ligase SIAH2 to directly promote the ubiquitination of AR-V7. Consequently, the genetic and pharmacological activation of the GRP78-dependent AR-V7 protein degradation restores the sensitivity of castration-resistant prostate cancer to anti-androgen therapy in cell culture and animal models. Conclusions: These findings not only provide a new approach for overcoming castration-resistance in prostate cancer therapy, but also increase our understanding about the interplay between molecular chaperones and ubiquitin ligase in shaping protein stability.
Asunto(s)
Alcaloides/uso terapéutico , Antagonistas de Receptores Androgénicos/uso terapéutico , Antineoplásicos Fitogénicos/uso terapéutico , Proteínas de Choque Térmico/antagonistas & inhibidores , Neoplasias de la Próstata Resistentes a la Castración/tratamiento farmacológico , Receptores Androgénicos/metabolismo , Animales , Castración , Línea Celular Tumoral , Chaperón BiP del Retículo Endoplásmico , Células HEK293 , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Proteínas Nucleares/metabolismo , Isoformas de Proteínas/antagonistas & inhibidores , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteolisis , Receptores Androgénicos/genética , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitinación , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Bcr-Abl is the primary cause as well as currently key therapeutic target of chronic myeloid leukemia (CML). SKP2, an E3 ligase, is a downstream factor of Bcr-Abl to motivate the cell cycle transition of CML and also found to bind and activate Bcr-Abl in reverse. Therefore, SKP2/Bcr-Abl pathway is an attractive target for CML treatment. This study aims to identify an inhibitor of the SKP2/Bcr-Abl pathway based on a large screening of the natural products. We demonstrate that Diosmetin, a kind of phytoestrogens, notably downregulates the expression of SKP2, Bcr-Abl phosphorylation, and moderately downregulates the Bcr-Abl level. Furthermore, Diosmetin displays a favorable anti-tumor activity in CML cells and xenograft models. Collectively, our study reveals a natural compound in the treatment of CML on the basis of SKP2/Bcr-Abl signaling pathway.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Proliferación Celular/efectos de los fármacos , Flavonoides/farmacología , Proteínas de Fusión bcr-abl/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Quinasas Asociadas a Fase-S/antagonistas & inhibidores , Animales , Apoptosis/efectos de los fármacos , Regulación hacia Abajo , Resistencia a Antineoplásicos , Regulación Neoplásica de la Expresión Génica , Humanos , Mesilato de Imatinib/farmacología , Células K562 , Leucemia Mielógena Crónica BCR-ABL Positiva/enzimología , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , Fosforilación , Proteínas Quinasas Asociadas a Fase-S/genética , Proteínas Quinasas Asociadas a Fase-S/metabolismo , Transducción de Señal , Carga Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Breast cancer (BCa) has the highest incidence and mortality among malignant diseases in female worldwide. BCa is frequently caused by estrogen receptor α (ERα), a ligand-dependent receptor that highly expressed in about 70% of breast tumors. Therefore, ERα has become a well-characterized and the most effective target for treating ERα-expressing BCa (ERα+ BCa). However, the acquire resistance was somehow developed in patients who received current ERα signaling-targeted endocrine therapies. Hence, discovery of novel anti-estrogen/ERα strategies is urgent. In the present study, we identified butein as a potential agent for breast cancer treatment by the use of a natural product library. We showed that butein inhibits the growth of ERα+ BCa both in vitro and in vivo which is associated with cell cycle arrest that partially triggered by butein-induced ERα downregulation. Mechanically, butein binds to a specific pocket of ERα and promotes proteasome-mediated degradation of the receptor. Collectively, this work reveals that butein is a candidate to diminish ERα signaling which represents a potentially novel strategy for treating BCa.
RESUMEN
Atherosclerosis-associated cardiovascular diseases are main causes leading to high mortality worldwide. Macrophage-derived foam cell formation via uptaking modified lipoproteins is the initial and core step in the process of atherosclerosis. Meanwhile, scavenger receptor is indispensable for the formation of foam cells. UCHL1, a deubiquitinase, has been widely studied in multiple cancers. UCHL1 could be an oncogene or a tumor suppressor in dependent of tumor types. It remains unknown whether UCHL1 influences cellular oxLDL uptake. Herein we show that UCHL1 deletion significantly inhibits lipid accumulation and foam cell formation. Subsequently, we found that UCHL1 inhibitor or siRNA downregulates the expression of CD36 protein whereas SR-A, ABCA1, ABCG1, Lox-1, and SR-B1 have no significant change. Furthermore, the treatment of UCHL1 inhibition increases the abundance of K48-polyubiquitin on CD36 and the suppression of lipid uptake induced by UCHL1 deficiency is attenuated by blocking CD36 activation. Our study concluded that UCHL1 deletion decreases foam cell formation by promoting the degradation of CD36 protein, indicating UCHL1 may be a potential target for atherosclerosis treatment.
Asunto(s)
Antígenos CD36/metabolismo , Células Espumosas/metabolismo , Ubiquitina Tiolesterasa/metabolismo , Transportador 1 de Casete de Unión a ATP/metabolismo , Transportadoras de Casetes de Unión a ATP/genética , Animales , Aterosclerosis/patología , Transporte Biológico , Línea Celular , Colesterol/metabolismo , Enzimas Desubicuitinizantes/metabolismo , Humanos , Metabolismo de los Lípidos/fisiología , Lípidos , Lipoproteínas LDL/metabolismo , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Receptores Depuradores de Clase A/metabolismo , Ubiquitina Tiolesterasa/genética , UbiquitinaciónRESUMEN
The incidence and mortality of breast cancer (BCa) are the highest among female cancers. There are approximate 70% BCa that are classified as estrogen receptor alpha (ERα) positive. Therefore, targeting ERα is the most significantly therapeutic schedule. However, patients with breast cancer develop resistance to ERα or estrogen (E2) antagonists such as fulvestrant and tamoxifen. In the present study, we found that L-Tetrahydropalmatine (L-THP) significantly suppressed cell proliferation in ERα+ BCa cells via inducing cell cycle arrest rather than apoptosis. Additionally, L-THP enhanced the sensitivity of ERα+ BCa cells to tamoxifen and fulvestrant. Mechanically, the application of L-THP promotes ERα degradation through accumulating ubiquitin chains on ERα. Overexpressing ERα abrogates L-THP induced-antiproliferation in ERα+ BCa cells. Collectively, our work indicates that L-THP may represent a potentially novel therapeutic medicine for ERα+ breast cancer patient.
Asunto(s)
Alcaloides de Berberina/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Supervivencia Celular/efectos de los fármacos , Receptor alfa de Estrógeno/antagonistas & inhibidores , Antineoplásicos Hormonales/farmacología , Línea Celular Tumoral , Antagonistas de Dopamina/farmacología , Resistencia a Antineoplásicos/efectos de los fármacos , Antagonistas de Estrógenos/farmacología , Femenino , Fulvestrant/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Modelos Moleculares , Conformación Proteica , Tamoxifeno/farmacologíaRESUMEN
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.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Resistencia a Antineoplásicos/efectos de los fármacos , Feniltiohidantoína/análogos & derivados , Pirroles/administración & dosificación , Pirrolidinas/administración & dosificación , Ubiquitina Tiolesterasa/metabolismo , Animales , Benzamidas , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Regulación hacia Abajo , Sinergismo Farmacológico , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Compuestos Heterocíclicos con 3 Anillos/administración & dosificación , Compuestos Heterocíclicos con 3 Anillos/farmacología , Humanos , Células MCF-7 , Ratones , Nitrilos , Feniltiohidantoína/administración & dosificación , Feniltiohidantoína/farmacología , Pirroles/farmacología , Pirrolidinas/farmacología , Transducción de Señal/efectos de los fármacos , Ubiquitina Tiolesterasa/antagonistas & inhibidores , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Breast cancer is the most common malignancy in women around the world. Estrogen receptor α (ERα) is expressed in approximately 70% of breast tumors, and considered as one of most effective targets in breast cancer therapy. It has been reported that the degradation of ERα protein is mediated by ubiquitin-proteasome system. However, little is known about the regulation of ERα deubiquitination, a critical constituent of its degradation control. The current study first reports that there is a positive correlation between ERα and ubiquitin specific protease 7 (USP7) protein levels in human breast tumor tissues. Subsequent studies showed that USP7 physically interacted with the ERα, thereby mediating the deubiquitination and stabilization of ERα. In addition, USP7 inhibition or silencing led to growth inhibition and apoptosis of ERα-positive breast cancer cells both in vitro and in vivo. Furthermore, overexpression of ERα rescued the USP7 silencing-induced cell cycle arrest and apoptosis, supporting that ERα status is essential to the function of USP7 in breast carcinogenesis. Overall, this study suggests that targeting USP7-ERα complex could be a potential strategy to treat ERα-positive breast cancer.