RESUMEN
Obesity, a recognized risk factor for breast cancer in postmenopausal women, is associated with higher mortality rates regardless of menopausal status, which could in part be explained by therapeutic escape. Indeed, adipose microenvironment has been described to influence the efficiency of chemo- and hormonal therapies. Residual cancer stem cells could also have a key role in this process. To understand the mechanisms involved in the reduced efficacy of hormonal therapy on breast cancer cells in the presence of adipose secretome, human adipose stem cells (hMAD cell line) differentiated into mature adipocytes were co-cultured with mammary breast cancer cells and treated with hormonal therapies (tamoxifen, fulvestrant). Proliferation and apoptosis were measured (fluorescence test, impedancemetry, cytometry) and the gene expression profile was evaluated. Cancer stem cells were isolated from mammospheres made from MCF-7. The impact of chemo- and hormonal therapies and leptin was evaluated in this population. hMAD-differentiated mature adipocytes and their secretions were able to increase mammary cancer cell proliferation and to suppress the antiproliferative effect of tamoxifen, confirming previous data and validating our model. Apoptosis and cell cycle did not seem to be involved in this process. The evaluation of gene expression profiles suggested that STAT3 could be a possible target. On the contrary, leptin did not seem to be involved. The study of isolated cancer stem cells revealed that their proliferation was stimulated in the presence of anticancer therapies (tamoxifen, fulvestrant, doxorubicine) and leptin. Our study confirmed the role of adipocytes and their secretome, but above all, the role of communication between adipose and cancer cells in interfering with the efficiency of hormonal therapy. Among the pathophysiological mechanisms involved, leptin does not seem to interfere with the estrogenic pathway but seems to promote the proliferation of cancer stem cells.
Asunto(s)
Adipocitos/metabolismo , Antineoplásicos Hormonales/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Resistencia a Antineoplásicos , Fulvestrant/farmacología , Leptina/metabolismo , Células Madre Neoplásicas/efectos de los fármacos , Tamoxifeno/farmacología , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Comunicación Celular , Proliferación Celular/efectos de los fármacos , Técnicas de Cocultivo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Células MCF-7 , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Factor de Transcripción STAT3/metabolismo , Transducción de Señal , Microambiente TumoralRESUMEN
BACKGROUND: Hormonal therapy is an effective treatment for luminal-like breast cancer. Aromatase inhibitor (AI) is widely used for estrogen receptor-positive, postmenopausal breast cancers. However, resistance is occurred and becomes a serious clinical concern. In general, progression of cancer strongly depends on tumor microenvironment, which may, therefore, also contribute to the development of AI resistance. METHODS: We evaluated tumor microenvironment-derived factors with respect to AI resistance using typical estrogen receptor-positive breast cancer cell lines. We established tumor microenvironment-dependent AI-resistant models and elucidated the underlying mechanisms. RESULTS: T-47D cells had a higher dependence on microenvironment-derived factors, such as estrogen or growth factors, for survival than MCF-7 cells. We, therefore, evaluated tumor microenvironment growth factors with respect to AI resistance using T-47D cells. We established three resistant cell lines (V1, V2, and V3) that survived estrogen deprivation and growth factor-supplemented conditions. These cell lines were deficient in estrogen receptor α expression and estrogen-dependent growth. Among six representative growth factors, epidermal growth factor was the most influential. In these models, HER2 protein was overexpressed without gene amplification and intracellular phosphorylation pathways were activated compared to parental cell lines. Molecular targeting inhibitors revealed that V1 and V2 primarily rely on the PI3 K pathway for survival, whereas V3 relies on the MAPK pathway. CONCLUSIONS: This study demonstrates the importance of tumor microenvironment-derived factors for the development of AI resistance. These resistant models did not utilize the same resistance mechanism, suggesting that flexible strategies are essential in conquering resistance.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/metabolismo , Resistencia a Antineoplásicos , Factor de Crecimiento Epidérmico/metabolismo , Receptor alfa de Estrógeno/metabolismo , Microambiente Tumoral , Inhibidores de la Aromatasa/efectos adversos , Inhibidores de la Aromatasa/uso terapéutico , Neoplasias de la Mama/patología , Supervivencia Celular/efectos de los fármacos , Receptor alfa de Estrógeno/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Células MCF-7 , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3/farmacología , Receptor ErbB-2/metabolismo , Tiazoles/farmacología , TransfecciónRESUMEN
BACKGROUND: Presently, hormonal therapy targeting estrogen receptors is the most effective treatment available for luminal breast cancer. However, many patients relapse after the therapy. It has been suggested that cancer stem-like cells are involved with hormonal therapy resistance; in the present study, we evaluated this hypothesis. METHODS: In the present study, we used our previously established hormonal therapy-resistant cell lines, including aromatase inhibitor (AI)-resistant cells (Type 1 and Type 2) and fulvestrant-resistant cells (MFR). RESULTS: AI-resistant cell lines expressing ER (Type 1 V1 and V2) showed high cancer stemness in terms of their CD44/CD24 expression and side populations, which were stimulated by the addition of estrogen and inhibited by fulvestrant. However, ALDH activity was lower than in the ER-negative resistant cells, suggesting that the stemness of luminal cells is distinct from that of basal-like breast cancer cells. The migration and invasion activity of the ER-positive Type 1 V1 and V2 cells were higher than in the ER-negative cell lines, Type 2 and MFR. CONCLUSIONS: Fractionation of parental cells based on CD44/CD24 expression and colony formation assay indicated that CD44+/CD24+ cells might be the origin of hormonal therapy-resistant cells. This population reconstituted various other subpopulations under estrogen deprivation. These results indicate that hormonal therapy resistance is closely related to the cancer stem cell-like properties of luminal breast cancer.
Asunto(s)
Antineoplásicos Hormonales/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Inhibidores de la Aromatasa/farmacología , Neoplasias de la Mama/metabolismo , Antígeno CD24/metabolismo , Línea Celular Tumoral , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Estradiol/metabolismo , Antagonistas del Receptor de Estrógeno/farmacología , Femenino , Fulvestrant , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Receptores de Hialuranos/metabolismo , Células MCF-7 , Células Madre Neoplásicas/patología , Receptores de Estrógenos/metabolismoRESUMEN
The loss of estrogen receptor α (ERα) expression in breast cancer constitutes a major hallmark of tumor progression to metastasis and is generally correlated to a strong increase in Hyaluronic Acid (HA) turnover. The aim of our study was to search for a putative link between these two major events of breast cancer progression in the estrogen receptor-positive (ER+) MCF7 breast cancer cell line. The increase in HA turnover was performed by stable overexpression of the standard CD44 (CD44S) isoform and also by treatment with exogenous Hyaluronidase (Hyal) to allow an increase in HA catabolism. Stable overexpression of CD44S in MCF7 cells was correlated to a decrease in ESR1 gene expression, which did not lead to alteration of estrogen response. Moreover, our results showed that the exposure to exogenous Hyal stimulates the proliferation and strongly decreases the expression of ERα whatever the expression level of CD44 in the MCF7 cell line. The culture in the presence of Hyal led to the decrease in estrogens responsiveness and to hormonal therapy resistance. The effect on growth is correlated to the activation of MAPK/ERK and PI3K/Akt signaling pathways while the Hyal-induced down-regulation of ESR1 gene expression involves the activation of PI3K/Akt and NF-κB signaling pathways. Many of our data suggest that the effects of Hyal described here could be related to the activation of TLR signaling. Taken together, our results demonstrate that the increase in HA degradation could be involved in breast cancer progression and in resistance to hormonal therapy.
Asunto(s)
Neoplasias de la Mama/metabolismo , Receptor alfa de Estrógeno/metabolismo , Ácido Hialurónico/metabolismo , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Estradiol/farmacología , Receptor alfa de Estrógeno/genética , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Inflamación/patología , Células MCF-7 , FN-kappa B/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transducción de Señal/efectos de los fármacos , Receptores Toll-Like/metabolismoRESUMEN
The relationship between tobacco smoke and breast cancer incidence has been studied for many years, but the effect of smoking on hormonal therapy has not been previously reported. We investigated the effect of smoking on hormonal therapy by performing in vitro experiments. We first prepared tobacco smoke condensate (TSC) and examined its effect on estrogen receptor (ER) activity. The ER activity was analyzed using MCF-7-E10 cells into which the estrogen-responsive element (ERE)-green fluorescent protein (GFP) reporter gene had been stably introduced (GFP assay) and performing an ERE-luciferase assay. TSC significantly activated ERs, and upregulated its endogenous target genes. This activation was inhibited by fulvestrant but more weakly by tamoxifen. These results suggest that the activation mechanism may be different from that for estrogen. Furthermore, using E10 estrogen depletion-resistant cells (EDR cells) established as a hormonal therapy-resistant model showing estrogen-independent ER activity, ER activation and induction of ER target genes were significantly higher following TSC treatment than by estradiol (E2). These responses were much higher than those of the parental E10 cells. In addition, the phosphorylation status of signaling factors (ERK1/2, Akt) and ER in the E10-EDR cells treated with TSC increased. The gene expression profile induced by estrogenic effects of TSC was characterized by microarray analysis. The findings suggested that TSC activates ER by both ligand-dependent and -independent mechanisms. Although TSC constituents will be metabolized in vivo, breast cancer tissues might be exposed for a long period along with hormonal therapy. Tobacco smoke may have a possibility to interfere with hormonal therapy for breast cancer, which may have important implications for the management of therapy.