RESUMEN
Triple negative breast cancer (TNBC) is a breast cancer subtype associated with high rates of metastasis, heterogeneity, drug resistance and a poor prognosis. Extracellular vesicles (EVs) are vesicles of endosomal and plasma membrane origin, and are secreted by healthy and cancer cells. In cancer, EVs contribute to tumor progression by mediating escape from the immune system surveillance, and are involved in extracellular matrix degradation, invasion, angiogenesis, migration and metastasis. Furthermore, EVs have been identified in several human fluids. However, the role of EVs from patients with breast cancer in the migration and invasion of human breast cancer cells is not fully understood. The present study investigated whether EVs isolated from Mexican patients with breast cancer can induce cellular processes related to invasion in breast cancer. Moreover, plasma fractions enriched in EVs and deprived of plateletderived EVs obtained from blood samples of 32 Mexican patients with biopsydiagnosed breast cancer at different clinical stages who had not received treatment were analyzed. Furthermore, one control group was included, which consisted of 20 Mexican healthy females. The present results demonstrated that EVs from women with breast cancer promote migration and invasion, and increase matrix metalloproteinase (MMP)2 and MMP9 secretion in TNBC MDAMB231 cells. In addition, it was found that EVs from patients with breast cancer induced Src and focal adhesion kinase activation, and focal adhesions assembly with an increase in focal adhesions number, while the migration and invasion was dependent on Src activity. Collectively, EVs from Mexican patients with breast cancer induce migration and invasion via a Srcdependent pathway in TNBC MDAMB231 cells.
Asunto(s)
Vesículas Extracelulares/metabolismo , Transducción de Señal , Neoplasias de la Mama Triple Negativas/patología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Movimiento Celular , Transición Epitelial-Mesenquimal , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Células MCF-7 , Metaloproteinasa 2 de la Matriz/metabolismo , Metaloproteinasa 9 de la Matriz/metabolismo , México , Persona de Mediana Edad , Invasividad Neoplásica , Estadificación de Neoplasias , Neoplasias de la Mama Triple Negativas/sangre , Neoplasias de la Mama Triple Negativas/metabolismo , Adulto Joven , Familia-src Quinasas/metabolismoRESUMEN
MicroRNAs derived from extracellular vesicles (EV-miRNAs) are circulating miRNAs considered as potential new diagnostic markers for cancer that can be easily detected in liquid biopsies. In this study, we performed RNA sequencing analysis as a screening strategy to identify EV-miRNAs derived from serum of clinically well-annotated breast cancer (BC) patients from the south of Brazil. EVs from three groups of samples (healthy controls (CT), luminal A (LA), and triple-negative (TNBC)) were isolated from serum using a precipitation method and analyzed by RNA-seq (screening phase). Subsequently, four EV-miRNAs (miR-142-5p, miR-150-5p, miR-320a, and miR-4433b-5p) were selected to be quantified by quantitative real-time PCR (RT-qPCR) in individual samples (test phase). A panel composed of miR-142-5p, miR-320a, and miR-4433b-5p distinguished BC patients from CT with an area under the curve (AUC) of 0.8387 (93.33% sensitivity, 68.75% specificity). The combination of miR-142-5p and miR-320a distinguished LA patients from CT with an AUC of 0.9410 (100% sensitivity, 93.80% specificity). Interestingly, decreased expression of miR-142-5p and miR-150-5p were significantly associated with more advanced tumor grades (grade III), while the decreased expression of miR-142-5p and miR-320a was associated with a larger tumor size. These results provide insights into the potential application of EVs-miRNAs from serum as novel specific markers for early diagnosis of BC.
Asunto(s)
Neoplasias de la Mama/genética , Vesículas Extracelulares/genética , Regulación Neoplásica de la Expresión Génica , MicroARNs/genética , Adulto , Anciano , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/genética , Neoplasias de la Mama/sangre , Femenino , Perfilación de la Expresión Génica , Humanos , MicroARNs/sangre , Persona de Mediana Edad , Neoplasias de la Mama Triple Negativas/sangre , Neoplasias de la Mama Triple Negativas/genéticaRESUMEN
Triple negative breast cancer (TNBC) is a major global public health problem. The lack of targeted therapy and the elevated mortality evidence the need for better knowledge of the tumor biology. Hypoxia and aberrant glycosylation are associated with advanced stages of malignancy, tumor progression and treatment resistance. Importantly, serum deprivation regulates the invasive phenotype and favors TNBC cell survival. However, in TNBC, the role of hypoxia and serum deprivation in the regulation of glycosylation remains largely unknown. The effects of hypoxia and serum deprivation on the expression of glycosyltransferases and glycan profile were evaluated in the MDA-MB-231 cell line. We showed that the overexpression of HIF-1α was accompanied by acquisition of epithelial-mesenchimal transition features. Significant upregulation of fucosyl- and sialyltransferases involved in the synthesis of tumor-associated carbohydrate antigens was observed together with changes in fucosylation and sialylation detected by Aleuria aurantia lectin and Sambucus nigra agglutinin lectin blots. Bioinformatic analysis further indicated a mechanism by which HIF-1α can regulate ST3GAL6 expression and the relationship within the intrinsic characteristics of TNBC tumors. In conclusion, our results showed the involvement of hypoxia and serum deprivation in glycosylation profile regulation of TNBC cells triggering breast cancer aggressive features and suggesting glycosylation as a potential diagnostic and therapeutic target.
Asunto(s)
Hipoxia/metabolismo , Polisacáridos/metabolismo , Neoplasias de la Mama Triple Negativas/metabolismo , Supervivencia Celular , Humanos , Hipoxia/sangre , Polisacáridos/sangre , Factores de Transcripción/metabolismo , Neoplasias de la Mama Triple Negativas/sangre , Neoplasias de la Mama Triple Negativas/patología , Células Tumorales CultivadasRESUMEN
BACKGROUND: Calcitriol (D) or 1,25(OH)2D3 inhibits the growth of several tumor cells including breast cancer cells, by activating cell death pathways. Menadione (MEN), a glutathione-depleting compound, may be used to potentiate the antiproliferative actions of D on cancer cells. We have previously shown in vitro that MEN improved D-induced growth arrest on breast cancer cell lines, inducing oxidative stress and DNA damage via ROS generation. Treatment with MEN+D resulted more effective than D or MEN alone. OBJECTIVE: To study the in vivo effect of calcitriol, MEN or their combination on the development of murine transplantable triple negative breast tumor M-406 in its syngeneic host. METHODS: Tumor M-406 was inoculated s.c., and when tumors reached the desired size, animals were randomly assigned to one of four groups receiving daily i.p. injections of either sterile saline solution (controls, C), MEN, D, or both (MEN+D). Body weight and tumor volume were recorded three times a week. Serum calcium was determined before and at the end of the treatment, at which time tumor samples were obtained for histological examination. RESULTS: None of the drugs, alone or in combination, affected mice body weight in the period studied. The combined treatment reduced tumor growth rate (C vs. MEN+D, P<0.05) and the corresponding histological sections exhibited small remaining areas of viable tumor only in the periphery. A concomitant DNA fragmentation was observed in all treated groups and MEN potentiated the calcitriol effect on tumor growth. CONCLUSIONS: As previously observed in vitro, treatment with MEN and D delayed tumor growth in vivo more efficiently than the individual drugs, with evident signals of apoptosis induction. Our results propose an alternative protocol to treat triple negative breast cancer, using GSH depleting drugs together with calcitriol, which would allow lower doses of the steroid to maintain the antitumor effect while diminishing its adverse pharmacological effects.