A study of transferrin and ferritin expression in tumor cells of patients with breast cancer

Aim: to study and to evaluate the clinical significance of the expression of iron-containing proteins ferritin and transferrin in tumor cells of patients with breast cancer (BC). Object and methods: the study included 143 patients with BC stage II–III. Methods: clinical, morphological and immunohistochemical, statistical. Results: it was shown that BC is characterized by intertumor heterogeneity of expression of transferrin and ferritin. The high degree of differentiation of BC correlates with the lack of expression of transferrin and ferritin. Positive expression of these proteins in the cells of primary tumors correlated with the development of metastases in regional lymphatic nodes. Correlation dependences between parameters of expression of transferrin and ferritin in tumor cells and overall survival in patients with BC have been determined. Conclusions: the expression of transferrin and ferritin may be individual predictive markers of clinical course and survival in patients with BC and one more molecular target for the development of new anticancer agents.Мета: вивчення експресії залізовмісних білків феритину (ФЕР) і трансферину (ТРФЕР) у пухлинних клітинах та оцінка її клінічного значення у хворих на рак молочної залози (РМЗ). Об’єкт і методи: у дослідження включено 143 пацієнток із РМЗ ІІ–ІІІ стадії. Методи дослідження: клінічні, морфологічний, імуногістохімічний, статистичний. Результати: показано, що РМЗ характеризується міжпухлинною гетерогенністю експресії ТРФЕР і ФЕР. Високий ступінь диференціювання РМЗ корелює з відсутністю експресії ТРФЕР і ФЕР. Позитивна експресія цих білків у клітинах первинної пухлини пов’язана з розвитком метастазів у регіонарних лімфатичних вузлах. Встановлено кореляційну залежність між експресією ТРФЕР та ФЕР у пухлинних клітинах і загальною виживаністю хворих на РМЗ. Висновки: експресія ТРФЕР і ФЕР може бути індивідуальним предиктивним маркером перебігу хвороби та виживаності пацієнток із РМЗ і ще однією молекулярною мішенню для розробки нових протипухлинних агентів

Significance of ferritin expression in formation of malignant phenotype of human breast cancer cells

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 ± 0.19·1016and 3.0 ± 0.22·1016) 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. Key Words: breast cancer, cell lines, luminal subtype, basal subtype, ferritin, “free iron”, ROS, SOD, miRNA 200b

Peculiarities of antioxidant system and iron metabolism in organism during development of tumor resistance to cisplatin

Aim : To study in vivo the peculiarities of changes of iron metabolism and antioxidant system in dynamics of growth of Guerin carcinoma with different sensitivity to cisplatin. Materials and Methods: In order to evaluate the content of metallothionein-1 (MT-1) in tumor homogenates and blood serum of rats with cisplatin-sensitive and cisplatin-resistant Guerin carcinoma the immunoenzyme method was used. The evaluation of ceruloplasmin activity, content of “free iron” complexes, superoxide and NO-generating acti­vity of NADPH-oxidase and iNOS activity in neutrophils, blood serum and tumor homogenates was measured by EPR-spectro­scopy. Results: Maximal accumulation of MT-1 in blood serum and tumor, more pronounced in resistant strain, at the border of latent and exponential phase of growth has been shown that is the evidence of protective role of this protein in the respect to the generation of free radical compounds. It has been determined that in animals with cisplatin-resistant strain of Guerin carcinoma, increase of “free iron” complexes is more apparent both on the level of tumor and organism on the background on increase of CP/TR ratio that is the consequence of organism antioxidant protection system disorder. Conclutions: Mentioned changes in metabolism of iron with its accumulation in tumor and further reprogramming of mitochondria metabolism and activity of NADPH-oxidase for non-transformed cells are favorable conditions for the formation of oxidative phenotype of tumor. Key Words: Guerin carcinoma, metallothionein-1, antioxidant system, transferrin, ceruloplasmin, “free iron” complexes, activity of NADPH-oxidase and iNOS neutrophils

Clinical significance of hormonal receptor status of malignant ovarian tumors

Objectives: To study hormonal receptor status (HRS) of malignant ovarian tumors (MOT) and determine its clinical significance. Patients and Methods: Retrospective analysis of case histories of 284 patients with MOT of different genesis of I–IV stages was carried out; immunohistochemical study of paraffin-embedded tissues. The HRS for serous, mucinous ovarian cancer (OC) and sex cord-stromal tumors (SCST) was studied. The phenotype of tumors by HRS in patients with serous OC was determined; overall and relapse-free survival in these patients was evaluated depending on the tumor HRS. Results: Positive expression of ER has been registered in 66.4% of patients with serous OC, PR — in 63.4%, TR — in 53.0%; in patients with mucinous OC — 88.0; 84.0; 60.0%, respectively. Positive staining of cells of stroma-cellular tumors has been observed in 74.1% of patients for ER and 77.8% — for PR and TR. The highest number of patients with tumor phenotype ER+PR+TR+ has been observed in postmenopause — 52.4%, especially in late postmenopausal period — 39.0%. The lowest percentage of patients with mentioned phenotype has been marked in reproductive age — 20.7%. Most patients of reproductive period had phenotype of tumor ER-PR-TR- (35.1%), in late postmenopause this phenotype has been observed only in 16.2%. The patients with serous OC with the positive tumor HRS demonstrated the low indices of overall and relapse-free survival compared to the patients with receptor-negative tumors con­cerning all steroid hormones (р < 0.05). Conclusions: Positive HRS was registered in serous, mucinous OC and in SCST, high percen­tage of tumors with expression of all receptors of steroid hormones was observed at that. The highest frequency of tumors with positive HRS was recorded in patients with serous OC of late postmenopausal period. The patients with serous OC with receptor-positive tumor phenotype showed the rates of overall and relapse-free survival significantly lower compared to the patients with receptor-negative phenotype of OC. Positive HRS, the same as strong expression of TR in patients with serous OC, is a predictive factor of unfavorable course of tumor process. HRS of MOT can be regarded as the additional criterion for solution of a question concerning application of hormonal therapy as a component of complex treatment for the patients. Key Words: malignant ovarian tumors, serous ovarian cancer, hormonal receptor status, estrogen, progesterone, testosterone receptors, phenotype of tumor

Increase of antitumor activity of cisplatin using agonist of gonadotropin-realising hormone and inhibitor of aromatase on the model of ascites ovarian tumor

Aim : To study antitumor activity of triptorelin — agonist of gonadotropin-releasing hormone and exemestane — inhibitor of aromatase in monotherapy and in combination with cisplatin on the model of receptor-positive for estrogens and progesterone malignant ascites transplantable ovarian tumor (TOT), to assess therapeutic pathomorphosis and level of VEGF expression in tumor cells using diffe­rent combinations of cytostatics and hormonal drugs. Materials and methods. 72 female Wistar rats, which underwent intraperitoneal transplantation of ascitic TOT, by 5·106 cells per animal, have been involved in the study. Rats were divided into 8 groups, 9 rats in each group. Histological study with assessment of therapeutic pathomorphosis in TOT and immunohistochemical study has been carried out. Survival of animals in the studied groups has been evaluated. Results. Among animals treated in regimen of monotherapy, the most pronounced antiangiogenic activity in TOT has been observed on application of hormonal drugs (triptorelin — 39.4 ± 1.9 and exemestane — 33.9 ± 1.4%; р = 0.003), the highest grade of treatment pathomorphosis in TOT has been observed at treatment with cisplatin (11.7%; р = 0.001). Combination of triptorelin and exemestane has amplified antiangiogenic activity in TOT (12.2 ± 0.9%; р = 0.001), but has not significantly changed rates of pathomorphosis (22.1 ± 0.4%; р=0.005) and survival of animals (32.2%; р = 0.007) as compared with the same rates in rats treated with hormonal drugs in monotherapy. Significant correlation between VEGF expression and pathomorphosis has been established (relative part of viable tumor tissue (RPVTT)) in TOT (r = 0.712; р = 0.001), as well as between RPVTT and life-span of animals (r = −0.320; р = 0.007). However, lack of correlation between VEGF expression in cells of TOT and survival of rats has been determined (r = −0.194; р = 0.11). Combination of cytostatic agent with triptorelin or exemestane has demonstrated significantly high rates of therapeutic pathomorphosis (10.1 ± 0.1% and 16.2 ± 0.3%, respectively) and antiangiogenic activity in TOT (21.4 ± 1.4% and 15.0 ± 1.3%, respectively) as well as the highest survival of animals (100.0%, increase of life-span (ILS) = 231.9% and 85.7%, ILS = 205.8%, respectively) as compared with the same one in rats treated in regimen of monotherapy with cisplatin, triptorelin, exemestane or by combination of hormonal drugs. Among animals treated by combination of cytostatic drug with triptorelin, two were cured, and among rats, which received cisplatin and exemestane, one animal was cured. Conclusions. Triptorelin and exemestane increase antitumor activity of cisplatin in respect to the malignant ascitic TOT and significantly increase survival of animals, especially when triptorelin and cisplatin are used in combination. Key Words: ascites transplantable ovarian tumor, rat, cytostatic, agonist of gonadotropin-releasing hormone triptorelin, inhibitor of aromatase exemestane, pathomorphosis, VEGF, survival