RESUMEN
Overexpression of ATP-binding cassette transporter (ABC) subfamily G2 in cancer cells is known to elicit a MDR phenotype, ultimately resulting in cancer chemotherapy failure. Here, we report, for the first time, the effect of eight novel pyrimido[1â³,2â³:1,5]pyrazolo[3,4-b]quinoline (IND) derivatives that inhibit ABCG2 transporter restoring cancer cell chemosensitivity. IND -4, -5, -6, -7, and -8, at 10 µM, and nilotinib at 5 µM, significantly potentiated (8-10 fold) the cytotoxicity of the ABCG2 substrates mitoxantrone (MX) and doxorubicin in HEK293 cells overexpressing ABCG2 transporter, MX (~14 fold) in MX-resistant NCI-H460/MX-20 small cell lung cancer, and of topotecan (~7 fold) in S1-M1-80 colon cancer cells which all stably expressing ABCG2. In contrast, cytotoxicity of cisplatin, which is not an ABCG2 substrate, was not altered. IND-5,-6,-7, and -8 significantly increased the accumulation of rhodamine-123 in multidrug resistant NCI-H460/MX-20 cells overexpressing ABCG2. Both IND-7 and -8, the most potent ABCG2 inhibitors, had the highest affinities for the binding sites of ABCG2 in modeling studies. In conclusion, the beneficial actions of new class of agents warrant further development as potential MDR reversal agents for clinical anticancer agents that suffer from ABCG2-mediated MDR insensitivity.
Asunto(s)
Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/antagonistas & inhibidores , Antineoplásicos/farmacología , Resistencia a Antineoplásicos/efectos de los fármacos , Proteínas de Neoplasias/antagonistas & inhibidores , Neoplasias/tratamiento farmacológico , Pirazoles/farmacología , Quinolinas/farmacología , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/química , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/genética , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/metabolismo , Antineoplásicos/síntesis química , Antineoplásicos/metabolismo , Sitios de Unión , Supervivencia Celular/efectos de los fármacos , Cisplatino/farmacología , Citocromo P-450 CYP3A/biosíntesis , Citocromo P-450 CYP3A/genética , Relación Dosis-Respuesta a Droga , Doxorrubicina/farmacología , Inducción Enzimática , Células HEK293 , Células Hep G2 , Humanos , Concentración 50 Inhibidora , Mitoxantrona/farmacología , Simulación del Acoplamiento Molecular , Proteínas de Neoplasias/química , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Receptor X de Pregnano , Regiones Promotoras Genéticas , Unión Proteica , Conformación Proteica , Pirazoles/síntesis química , Pirazoles/metabolismo , Pirimidinas/farmacología , Quinolinas/síntesis química , Quinolinas/metabolismo , Receptores de Esteroides/agonistas , Receptores de Esteroides/genética , Relación Estructura-Actividad , TransfecciónRESUMEN
Tamoxifen is widely used as an adjuvant therapy for patients with estrogen receptor (ERα)-positive tumors. However, the clinical benefit is often limited because of the emergence of drug resistance. In this study, overexpression of ribonucleotide reductase M2 (RRM2) in MCF-7 breast cancer cells resulted in a reduction in the effectiveness of tamoxifen, through downregulation of ERα66 and upregulation of the 36-kDa variant of ER (ERα36). We identified that NF-κB, HIF1α, and MAPK/JNK are the major pathways that are affected by RRM2 overexpression and result in increased NF-κB activity and increased protein levels of EGFR, HER2, IKKs, Bcl-2, RelB, and p50. RRM2-overexpressing cells also exhibited higher migratory and invasive properties. Through time-lapse microscopy and protein profiling studies of tamoxifen-treated MCF-7 and T-47D cells, we have identified that RRM2, along with other key proteins, is altered during the emergence of acquired tamoxifen resistance. Inhibition of RRM2 using siRRM2 or the ribonucleotide reductase (RR) inhibitor didox not only eradicated and effectively prevented the emergence of tamoxifen-resistant populations but also led to the reversal of many of the proteins altered during the process of acquired tamoxifen resistance. Because didox also appears to be a potent inhibitor of NF-κB activation, combining didox with tamoxifen treatment cooperatively reverses ER-α alterations and inhibits NF-κB activation. Finally, inhibition of RRM2 by didox reversed tamoxifen-resistant in vivo tumor growth and decreased in vitro migratory and invasive properties, revealing a beneficial effect of combination therapy that includes RRM2 inhibition to delay or abrogate tamoxifen resistance.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Resistencia a Antineoplásicos/efectos de los fármacos , Ácidos Hidroxámicos/farmacología , FN-kappa B/antagonistas & inhibidores , Ribonucleósido Difosfato Reductasa/antagonistas & inhibidores , Tamoxifeno/farmacología , Animales , Antineoplásicos/farmacología , Western Blotting , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Sinergismo Farmacológico , Receptores ErbB/metabolismo , Receptor alfa de Estrógeno/genética , Receptor alfa de Estrógeno/metabolismo , Humanos , Estimación de Kaplan-Meier , Células MCF-7 , Ratones Desnudos , FN-kappa B/metabolismo , Interferencia de ARN , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ribonucleósido Difosfato Reductasa/genética , Ribonucleósido Difosfato Reductasa/metabolismo , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Alterations in PI3K-AKT-mTOR signaling have been implicated in diabetes. This study assessed whether disruption of PRAS40, a substrate of AKT and component of mTORC1, would alter glucose homeostasis and prevent hyperglycemia in the streptozotocin (STZ)-induced diabetes mouse model. PRAS40 ablation resulted in a mild lowering of blood glucose levels and glycated hemoglobin (HbA1C), a lowered insulin requirement, and improved glucose tolerance in untreated PRAS40 gene knockout (PRAS40(-/-)) as compared to wild-type (PRAS40(+/+)) mice. Diabetes was then induced in these mice using STZ at 50mg/kg/day over five days. Following STZ-treatment, PRAS40(-/-) mice exhibited significantly lower blood glucose and HbA1C levels than PRAS40(+/+) mice. Liver tissue of PRAS40(-/-) mice and shPRAS40 Hep3B cells showed increased activation of AKT (p-AKT T308) and mTORC1 (p-p70S6K) signaling as well as decreased p-AKT (S473) and increased p-IRS1 (S612) protein levels. Altered tissue gene expression of several glucose transporters (GLUT) and increased hepatic GLUT4 protein levels were observed in PRAS40(-/-) as compared to PRAS40(+/+) mice. In summary, PRAS40 deletion significantly attenuates hyperglycemia in STZ-induced PRAS40(-/-) mice through increased hepatic AKT and mTORC1 signaling, a lowered serum insulin requirement, and altered hepatic GLUT4 levels.
Asunto(s)
Glucosa/metabolismo , Homeostasis , Fosfoproteínas/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Animales , Secuencia de Bases , Línea Celular , Cartilla de ADN , Insulina/metabolismo , Ratones , Ratones Noqueados , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Transducción de SeñalRESUMEN
Dysregulation of PI3K-AKT-mTOR pathway has been reported in various pathologies, such as cancer and insulin resistance. The proline-rich AKT substrate of 40-kDa (PRAS40), also known as AKT substrate 1 (AKT1S1), lies at the crossroads of these cascades and inhibits the activity of the mTOR complex 1 (mTORC1) kinase. This review discusses the role of PRAS40 and possible feedback mechanisms, and alterations in AKT/PRAS40/mTOR signaling that have been implicated in the pathogenesis of tumor progression. Additionally, we probed new datasets extracted from Oncomine, a cancer microarray database containing datasets derived from patient samples, to further understand the role of PRAS40 (AKT1S1). These data strongly supports the hypothesis that PRAS40 may serve as a potential therapeutic target for various cancers.