RESUMEN
AIM: To search for additional molecular-biological markers of cancer stem cell (CSC) involved in the development of intra-tumor heterogeneity for the detection of features of the breast cancer (BC) pathogenesis. MATERIALS AND METHODS: Expression of estrogen receptors (ER), progesterone receptors (PR), Her2/neu, E- and N-cadherin, CD24, CD44, Bcl-2, Bax, Slug, P-gp, glutathione-S-transferase (GST) and metallothionein in cell lines was determined by the immunocytochemical method. Expression of ER, PR, Her2/neu, CD24 and CD44 in the surgical material of BC patients were determined by the immunohistochemical method. The levels of the miRNA were determined using real-time polymerase chain reaction. RESULTS: Cells of high-grade malignancy (HGM), MDA-MB-231 and MDA-MB-468 are characterized by high expression of stem cell markers compared to the cells of low-grade malignancy (LGM), T47D and MCF-7: CD44 levels in T47D and MCF-7 cells were in range of 72-79 points, which is significantly lower than in HGM cells (p < 0.05). Also, HGM cells with the properties of CSC were characterized by high expression of antiapoptotic proteins, the transcription factor Slug, and low levels of proapoptotic protein Bax (p < 0.05) compared to LGM cells. In cells with CSC characteristics an increased expression of transferrin and its receptor, ferritin, fentorin and hepcidin was revealed indicating activation of the endogenous iron metabolism. The characteristic feature of HGM cells with CSC phenotype were the increased levels of oncogenic miR-221, -155 and -10b by 60%, 92% and 78%, respectively, and decreased levels of oncosuppressive miR-29b, -34a and -200b by 8.4 ± 0.3, 4.6 ± 0.2, and 3.4 ± 0.6 times compared to MCF-7 line cells. It has been established that the development of resistance to cytostatics is accompanied by increased aggressiveness of tumor cells, loss of expression of hormonal receptors and acquiring of stem phenotype. In particular, increased expression of P-gp was observed in BC cells during the development of resistance to doxorubicin, of GST during the development of resistance to cisplatin along with increased CD44 expression (p < 0.05). We have revealed the relation between the presence of cells with the CSC phenotype (CD44+CD24-/low) and clinical and pathological characteristics of BC patients, their survival and BC sensitivity to neoadjuvant therapy (p > 0.05). CONCLUSIONS: The dependence between the expression of CSC markers and the degree of malignancy of tumor cells, development of resistance to cytostatics in vitro was established as well as the predictive value of the detection of the CSC for the individual prognosis of the BC course and sensitivity of the tumors to the treatment.
Asunto(s)
Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Antígeno CD24/metabolismo , Receptores de Hialuranos/metabolismo , Adulto , Anciano , Antineoplásicos/farmacología , Biomarcadores , Neoplasias de la Mama/genética , Antígeno CD24/genética , Línea Celular Tumoral , Progresión de la Enfermedad , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Femenino , Humanos , Receptores de Hialuranos/genética , Inmunohistoquímica , Inmunofenotipificación , Persona de Mediana Edad , Clasificación del Tumor , Estadificación de Neoplasias , Células Tumorales CultivadasRESUMEN
AIM: To determine frequency of tumors with immunohistochemical markers of cancer stem cells (CSC) CD44+/CD24- in patients with breast cancer (BC) of different molecular subtype and to evaluate their prognostic value. OBJECT: Surgical material of 132 patients with BC stage I-II, age from 23 to 75 years, mean age - 50.2 ± 3.1 years was studied. METHODS: Clinical, immunohistochemical (expression CD44+/CD24-), morphological, statistical. RESULTS: BC is characterized by heterogeneity of molecular subtypes and expression of markers (CD44+/CD24-). Immunohistochemical study of expression of CSC markers in surgical material has detected their expression in 34 (25.4%) patients with BC of different molecular subtypes. The highest frequency of cells with expression of CSC marker was observed in patients with basal molecular subtype (44.8% patients). Most of BC patients with phenotype CD44+/CD24 had stage I of tumor process (34.3%). Statistical processing of data has showen that Yule colligation coefficient equaled 0.28 (Ñ > 0.05) that argues poor correlation between stage of tumor process and number of tumors with positive expression of CSC markers. Statistical processing of data has showen high correlation between presence of cells with expression of CSC markers and metastases of BC in regional lymph nodes (Yule colligation coefficient equals 0.943; Ñ < 0.5). Difference in overall survival of patients with BC of basal molecular subtype depending on expression of CSC CD44+/CD24- markers was detected. Survival of patients with basal BC was reliably higher at lack in tumors of cells with CSC markers CD44+/CD24- and, correspondingly, lower at presence of such cells (Ñ < 0.05). In patients with BC of luminal (A and B), HER-2-positive subtypes, significant change in survival of patients depending on expression of CSC markers was not determined (Ñ > 0.05). CONCLUSION: Significance of tumor cells with markers CD44+/CD24- within the limits of molecular subtype of BC may be additional criterion for advanced biological characteristic of BC, and in patients with BC of basal molecular subtype - for predictive evaluation of individual potential of tumor to aggressive clinical course.
Asunto(s)
Biomarcadores de Tumor/análisis , Neoplasias de la Mama/diagnóstico , Mama/patología , Antígeno CD24/análisis , Receptores de Hialuranos/análisis , Células Madre Neoplásicas/patología , Adulto , Anciano , Neoplasias de la Mama/patología , Femenino , Humanos , Persona de Mediana Edad , Pronóstico , Estudios Retrospectivos , Adulto JovenRESUMEN
AIM: The aim of our study is to investigate the disorders of ferritin functioning in breast cancer (BC) cells of different molecular subtype. MATERIALS AND METHODS: The cell lines used in the analysis include T47D, MCF-7, MDA-MB-231, MDA-MB-468, MCF-10A, and 184A1. Ferritin heavy chains (FTH) expression was studied by immunohistochemical method. "Free iron" content and superoxide dismutase (SOD) activity were determined by means of EPR spectroscopy. Reactive oxygen species (ROS) level and peculiarities of microRNA expression in studied cell lines were evaluated using flow cytometry and PCR analysis, respectively. RESULTS: It has been demonstrated that FTH expression directly correlates with proliferative activity of cells of both luminal (r = 0.51) and basal subtypes (r = 0.25), inversely correlates with expression of steroid hormones in cells of basal subtype (ER: r = -0.46; PR: r = -0.44) and does not depend on tumorigenic activity of both subtypes of studied cells (r = 0.12 and r = 0.9). Obtained data are the evidence that cells of luminal subtype B (MCF-7 cell line) and basal subtype (MDA-MB-231 and MDA-MB-468 cell lines) with high proliferative activity contain the highest level of free iron (2.9 ± 0.19·10(16)and 3.0 ± 0.22·10(16)) that can be consequence of intensive use of this element by cells, which actively divide and grow. Along with it, in cell of lines of basal subtype MDA-MB-231 and MDA-MB-468, high level of FTH (254 ± 2.3 and 270 ± 1.9) is being detected in consequence of increase of level of free iron, ROS (11.3 ± 1.05 and 7.27 ± 0.26) and SOD (9.4 ± 0.24 and 8.5 ± 0.18) as well as decrease of expression of microRNA 200b. In contrast, cells of luminal subtype B of MCF-7 line were distinguished by high expression of microRNA 200b and low ferritin level (125 ± 2.7). CONCLUSION: Obtained data demonstrate that tumor cells, which are referred to different molecular subtypes, are characterized by changes in system of support of balance of intercellular iron and certain associations of studied factors.
Asunto(s)
Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Ferritinas/metabolismo , Regulación Neoplásica de la Expresión Génica , MicroARNs/genética , Neoplasias de la Mama/genética , Femenino , Ferritinas/genética , Humanos , Técnicas para Inmunoenzimas , Oxidorreductasas , Fenotipo , ARN Mensajero/genética , Especies Reactivas de Oxígeno/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Superóxido Dismutasa/metabolismo , Células Tumorales CultivadasRESUMEN
AIM: To study the expression of estrogen receptors (ER) and progesterone receptors (PR) and proliferation marker Ki-67 in ovarian tumors using immunohistochemistry, and evaluate possible prognostic significance of these markers. METHODS: Immunohistochemical evaluation of Ki-67, ER and PR expression was performed on serous ovarian cancer (OC) tissue samples from 81 OC patients. RESULTS: Serous OC is characterised by high proliferative activity and increased expression of steroid hormone receptors compared to nontransfomed ovarian surface epithelium. It has been shown that ER and PR expression levels depend on tumor histologic grade and the stage of the disease, and are variable between tumors of the same grade. The ER and PR expression levels correlate with OC patients' survival. CONCLUSION: Proliferative activity and steroid hormone receptor status along with clinical and morphological characteristics of serous OC possess prognostic significance and may be used for evaluation of the disease course.