[Molecular Diagnostics of the Involvement of Visually Normal Mucosa in the Malignancy Process in Urothelial Bladder Cancer].

Immunofluorescent method by flow cytometry was used to quantify the expression of the tumor-associated protein ßIII-tubulin (TUBB3) in the tissue of urothelial bladder cancer and visually normal mucosa (56 samples in total). The expression of the marker was detected in 100% of cases, and heterogeneity of the TUBB3 expression level both in tumor tissue and in "normal" mucosa was revealed. The level of TUBB3 in the "normal" mucosa did not depend on the distance from the tumor (1 cm or more than 3 cm) and, on average, it was lower than in the tumor tissue (21.8 ± 10.8% and 24.9 ± 13.2% vs 35.2 ± 12.4%; p = 0.04 and 0.005, respectively). An increase of the TUBB3 expression in the tumor and in the "normal" mucosa was revealed in muscle invasive bladder cancer compared to non-muscle invasive bladder cancer. Therefore, in urothelial bladder cancer, the tumor-associated protein TUBB3 is a molecular marker of bladder mucosa involvement in the malignancy process and predicts the risk of tumor muscle invasion, which may influence indications for early cystectomy.

Panel of Candidate Genes to Predict the Survival of Patients with Clear Cell Renal Cancer on the Basis of Gene Expression Regulated by HIF1α/HIF2α.

The detection of genes related to the lifetime of patients with clear cell renal cancer provides information on the mechanisms of the tumor development and can be the basis for creating approaches to predict patient survival. In this paper, the expression of genes regulated by the HIF2α transcriptional factor was studied. Based on the results obtained here and previously identified genes regulated by the transcriptional factor HIF1α, a new panel of 6 genes, including the BAP1 gene, was proposed. Expression of genes of this panel allows predicting the survival of patients with clear cell renal cancer with high sensitivity (93%), specificity (96%), and relative risk (21.5). After verification, the application of this panel can be useful for personalized treatment of patients with clear cell renal cancer, which will increase the effectiveness of therapy.

[BRCA1 and Estrogen Receptor α Expression Regulation in Breast Cancer Cells].

BRCA1 (breast cancer 1) protein is involved in the genome stability maintenance participating in homologous recombination-dependent DNA repair. Disruption of BRCA1 functioning is associated with breast and ovarian cancer. Despite the important role of BRCA1 in DNA repair in all cell types, the development of BRCA1-associated cancer takes place mainly in estrogen-dependent tissues such as breast and ovarian ones. Using breast cancer cell line MCF-7 it was demonstrated in in vitro experiments that the estrogen 17ß-estradiol (E2), phytoestrogens (genistein and apigenin) and antiestrogens (tamoxifen and fulvestrant) inhibited estrogen receptor (ERα) expression while only genistein influenced BRCA1 increasing its expression. In hypoxia, that is an important factor of solid tumors progression, the decrease of BRCA1 and ERα expression was demonstrated in MCF-7 cells. Therefore, hypoxia influences both BRCA1-dependent DNA repair and hormonal regulation of breast cancer cell growth. Taken together, obtained results demonstrate a relationship between BRCA1 and steroid hormones signal transduction pathways in breast cancer cells and point out to the importance of complex BRCA1 and ERa expression regulation mechanisms studies including epigenetic gene expression regulation.

Estrogen Receptors alpha and beta in Ovarian Cancer: Expression Level and Prognosis.

The quantitative immunofluorescence assay of serous ovarian cancer tissue for the expression of estrogen receptors (ERα and ERß) revealed a higher expression level of ERß in comparison with ERα in all surgical tumor samples investigated. Significant differences in the expression level of the markers were detected "from tumor to tumor." A high expression level of both ERα (≥ 25%) and ERß (≥ 44%) in the tumor predicts a significantly longer progression-free survival time (p < 0.01) in the patients after the first line of platinum and taxane-based adjuvant chemotherapy.

Flow cytometric analysis of the ERCC1, marker of DNA damage repair, in the tumor specimens embedded into paraffin blocks.

The differences in expression of ERCC1 were estimated between tumor specimens embedded into paraffin blocks and surgical biopsy specimens of non-small cell lung cancer as well as breast and ovarian cancers. Concordance or differences not higher than 20% were observed in 73% of the cases. The number of the cases with more significant differences in ERCC1 expression was less than 17%. The results show that ERCC1 detection in surgical biopsy specimens by flow cytometry is the more preferable method due to reduced preanalytical phase of the analysis.

Digoxin is a selective modifier increasing platinum drug anticancer activity.

Using the model of breast cancer Ehrlich ascites tumor in mice, we showed that a sigle intraperitoneal injection of cardiac glycoside digoxin 1 h before the intraperitoneal injection of cisplatin increased the anticancer effect of the cytostatic drug more than twice when recalculated for the dose. It is assumed that the modifying effect of digoxin is determined by the direct inhibition of glycolysis in tumor cells. Taking into account the design of the study, we consider promising the clinical evaluation of the effectiveness of digoxin as a modifier of cisplatin efficiency in intracavitary therapy of ascites cancers with pleural and abdominal dissenmination.

New Data on Molecular Targets of Tamoxifen Besides Estrogen Receptors, Their Clinical Significancy.

Tamoxifen is the first target agent with a high-end position in breast cancer therapy till now. In recent years experimental researches revealed new biological effects of tamoxifen on tumor cells. The present study continues the theme of the review published in 2012, where a plenty of tamoxifen effects besides interaction with estrogen receptors was discussed. Thus, there is described a wide range of the drug targets which are the key points of signal cascades activating the cell proliferation and determining the course of the growth of the cancer and its sensitivity to chemotherapy. Also clinical trials of tamoxifen based on existing of targets besides the estrogen receptors are reviewed. Furthermore, the data on the antiviral, antibacterial, antifungal and antiparasitic activities of tamoxifen are indicated.

Analysis of informativeness of immunohistochemical and flow cytometric methods for estrogen receptor α assessment.

Informative capacity analysis of immunohistochemistry (IHC) and flow cytometry (FCM) in the assessment of estrogen receptor α (ERα) expression in breast cancer tissue was performed. Similar frequencies of expression were shown by both methods: 27% of ERα-negative and 73% ERα-positive cases. However, IHC evaluation detected low levels in only 20% of ERα-positive cases, whereas low levels of ERα detected by FCM were 2 times more often (48%). Moreover, FCM revealed positive expression (23-60%) in 33% of IHC ERα-negative cases. Among IHC ER-positive cases, zero ERα expression was detected by FCM in 12.5%. The approaches to minimize errors in routine clinical determination of the estrogen receptor status were proposed.

[ERCC1 as a Marker of Ovarian Cancer Resistance to Platinum Drugs].

The review is concerned with the crucial marker of nucleotide excision repair ERCC1 and its contribution to platinum resistance of ovarian cancer. All the variants of the laboratory and clinical ERCC1 assessment in the ovarian cancer tissue (single nucleotide polymorphisms of the ERCC1 gene, levels of mRNA or protein) are considered. Data on the prognostic and predictive value of ERCC1 as a marker of the response to platinum-based therapy in ovarian cancer are systematized. The authors discuss the possible causes of heterogeneity of the results and emphasize the necessity of a unified and integrated approach to evaluation of ERCC1 in the tumor. The publications cited in the Search Engine Pub Med up to January 2015 were analyzed.

[Discordance of estrogen receptor status between primary and metastatic breast cancer--possible reasons and prognostic value].

The aim of the review was systematization of the data on discordance in expression of estrogen receptors between primary and metastatic breast cancer, different metastases and repeated analyses of the same tissue. The possible reasons for the phenomenon are discussed. The authors emphasize the need to analyze estrogen receptors in breast cancer metastases, regardless of the receptor status of the primary tumor, for predicting the course of the metastatic disease and providing an adequate treatment of the metastatic tumor in strict accordance with its receptor status during drug therapy. The works cited in the search engine Pub Med to May 2013 were analyzed.

[Estrogen receptors beta--new target in cellular lung cancer treatment].

Literature review upon various types of estrogen receptors expression (type alpha and beta) in the cells of cellular lung cancer, their participation in estrogen and antiestrogen effects implementation, influence of estrogens and antiestrogens on occurrence and progression of malignant lung tumors in animals and humans. Were analyzed reasons of data ambiguity on type beta estrogen receptors (ERbeta) expression frequency. The results of authors own research in quantitative assessment of ERbeta expression in tumor tissue of patients with cellular lung cancer (79 male and 22 female patients are presented in this article. An increase in expression rate and incidence of tumors with high ERbeta level has been shown in patients with lung adenocarcinoma regardless of smoking status or gender. A new strategy of antiestrogen use, especially tamoxifen, has been formulated for cellular lung cancer treatment. Authors believe in a positive effect of adjuvant treatment with tamoxifen in patients with ERbeta-positive cellular lung cancer used independently or during and after the chemotherapy, by analogy with breast cancer patients.

[Expression of estrogen receptors beta and beta1 in tissue of human non-small cell lung cancer].

Comparability of the level and intensity of estrogen receptors beta (ERbeta) expression in non-small cell lung cancer tissue of 32 patients was analyzed by flow cytometry using various antibodies--to the total fraction of ERbeta (clone 14C8) as well to the full-length ERbeta1 isoform (clone EMRO2). The differences in the ER expression indexes detected by anti-ERbeta or anti-ERbeta1 antibodies were revealed in some patients, but it had no influence on average indexes of the ERbeta expression in the patient groups investigated. It was confirmed by the findings on more frequent and more intensive expression of ERbeta in the non-small cell lung cancer tissue of female patients vs. the males irrespective of antibody type - anti-ER/ or anti-ERbeta1. Therefore, in comparative analysis of ERbeta expression in the groups of the patients with different clinicomorphologic characteristics of the disease it is possible to use both the antibodies. For individual disease prognosis in the routine clinical practice it is recommended to use the antibodies to the total fraction of ERbeta, since there are individual differences between the ERbeta expression indexes revealing by various types of antibodies.

[Tamoxifen molecular targets different from estrogen receptors].

Experimental studies showing ever new biological effects of tamoxifen on tumor cells, both expressing and nonexpressing estrogen receptors, are providing a novel conception of the drug, likely well known at present. The review describes tamoxifen targets, whose blocking induces inhibition of tumor cell growth and angiogenesis, stimulation of the programmed cell death (apoptosis, autophagia and necrosis), inhibition of multiple drug resistance mechanism and inhibition of invasion and metastasizing. In all the events, the results of the tamoxifen interaction with the cells are prognostically favourable from the viewpoint of both the inhibition of the tumor growth and metastasizing and the susceptibility to the medicinal therapy, that is considered by some authors as an extremely important addition to the tamoxifen antiestrogenic effect. The strategy of long-term tamoxifen adjuvant therapy of breast cancer with positive status of the estrogen reseptors was developed by Craig V. Jordan as far back as in the seventies of the XXth century, however there are arguments allowing to consider it also useful for the treatment of other tumors. First of all it is the fact described lately in regard to expression of estrogen beta-reseptors in solid tumors of practically all known localization and histological types, that are also the targets of tamoxifen. Apart from estimation of estrogen receptors, it is believed by some authors that molecular and biological choice of patients is necessary with an account of expression of other cell targets of antiestrogen for complete realization of all the aspects of tamoxifen biological activity in long-term adjuvant therapy of malignant tumors of various localization.

Estrogen receptors, antiestrogens, and non-small cell lung cancer.

This review considers data on expression of different types of estrogen receptors (ERα and ERß) in in vitro cultured cells of non-small cell lung cancer and also in human and animal lung tumors. Estrogens are shown to play an important role in genesis and development of non-small cell lung cancer because the estrogen-stimulated cell proliferation as well as antiestrogen-caused inhibition of proliferation occurred only in the cells expressing different types of estrogen receptors. In general, the situation is similar to that observed in breast cancer, but in the cells of non-small cell lung cancer not ERα are expressed in more than half of cases but ERß. Just estrogen receptors ß play the crucial role in inducing cell proliferation in response to estrogens, and ERß is a prognostic marker of a favorable course of non-small cell lung cancer. Data on the interactions between ER and EGFR signaling pathways, as well as on the additive antitumor effect of antiestrogens (tamoxifen and fulvestrant) combined with tyrosine kinase inhibitors (gefitinib, erlotinib, and vandetanib) are considered. The review also includes data on the influence of estrogens on genesis and development of lung cancer in humans and animals and the frequency of ERα and ERß expression in non-small cell lung cancer in tissues from patients of the two sexes. Problems of quantitative determination of α and ß estrogen receptors in the tumor cells are also discussed.

[Glutoxime--an inhibitor of multiple drug resistance phenotype associated with Pgp expression].

Interaction of Glutoxime with P-glucoprotein (Pgp), a multiple drug resistance marker, as well as the Glutoxime impact on doxorubicin intracellular accumulation were investigated. It was shown that the Glutoxime effect on the Pgp expressing tumor cells resulted in a decrease of the cell specific fluorescence intensity, conditioned by binding of the monoclonal antibodies to the transport protein. That was evident of Glutoxime competition with the monoclonal antibodies for binding to Pgp and indicative of the modificator interaction with the transport protein. The effect was proved with the use of two cultures of human tumor cells of different histogenesis, i.e., the cells of Jurkat T-cellular leukemia and nonsmall cell lung cancer A549. Inhibition of the Pgp functional activity by Glutoxime was also demonstrateds. The authors suggested that it could be caused by direct competition of the modificator with the antitumor agent for binding to the precipitation sites on Pgp. Glutoxime could be considered as an inhibitor of multiple drug resistance associated with the Pgp function.

[Parameters of specific antibody interaction with P-gp in T-cell leukemia Jurkat cells].

UNLABELLED: Special features of Pgp expression evaluation by flow cytometry were investigated. Indexes of interaction of FITC-conjugated Becton Dickinson Pharmingen monoclonal antibodies to external Pgp epitope (clone 17F9) were analyzed depending on the cell concentration (400000 to 3000000 cells/ml) and the specific antibody concentration (5, 10 and 20 microl of the market product solution per 300 microl of the cell suspension). RESULTS: 1. Optimal condition of incubation with the antibodies was revealed--after the cell fixation in 4% formaldehyde. 2. Character of the increase of the cell fluorescence average intensity in the suspension totally according to the concentration of the Pgp-specific antibodies did not depend on the number of the cells. 3. Both the absolute value of the average intensity of the cell specific fluorescence as well as cell number out of the isotypic control fluorescence region depended on the ratio of the cell number to monoclonal antibody concentration. CONCLUSION: 1. It was shown that Pgp was practically expressed in all Jurkat cells. 2. By the Pgp expression level, the Jurkat cell culture was sufficiently homogeneous and stable in various passages. 3. Jurkat cells could be used as test culture in estimation of the market antibody activity. 4. For immunofluorescent assay of the Pgp expression in human tumor biopsy specimens, it is necessary to use not less than three concentrations of the specific antibodies, not less than three concentrations of the cells in the suspension as well as concurrent assay of the cell culture characterized previously. In particular, for investigated Pgp monoclonal antibodies, it is possible to use Jurkat cell culture. It allows revealing not only the fact of the Pgp expression but the level of the expression as well, i.e. to estimate severity of multidrug resistance phenotype.

[Estrogen receptor expression in tumors different from breast cancer].

A review of the literature data on expression of estrogen receptor alpha and beta (ERalpha and ERbeta) in tumors different from breast cancer. The results regarding the ERalpha and ERbeta expression frequency in non-small cell and small cell lung cancer, colorectal cancer, esophageal, ovarian, prostate and brain tumors are presented. High frequency of estrogen receptor expression (in up to 50 and more per cent of cases) in various types of tumors, differences between ERalpha and ERbeta in expression frequency, prognostic significance and prediction of the neoplastic process aggressiveness as well as in biological implications of interaction with antiestrogens (antagonistic and/or agonistic effect) are shown. The data on comparative evaluation of ERalpha and ERbeta expression in lung, ovarian, prostate tumor cells and corresponding nonneoplastic tissues are reported. Authors consider necessary to include the ERalpha and ERbeta detection into the routine clinical practice not only in breast cancer but in other tumors as well. Prospects of the clinical application of antiestrogens, in particular tamoxifen, in adjuvant therapy of different tumors with positive ER status are discussed.

[The pharmacological strategy of overcoming the phenotype of the multiple pharmacoresistance of non-small-cell lung cancer].

The authors discuss literature data and the results of their own many-year research into molecular markers of multiple pharmacoresistance associated with the function of ABC-transporters (MDRABC) from the point of view of their clinical significance. They consider the evolution of views on the mechanism of functioning and intracellular localization of ABC-transporters in normal and tumoral cells, in particular, the role of these transport proteins in the protection of nuclear targets from toxic agents, including antitumoral preparations. By the example of non-small-cell lung cancer (NSCLC) the authors critically analyze the reasons why data on the frequency of ABC-transporters expression in human solid tumors and on their prognostic value are inconsistent. The article contains a detailed analysis of the results of the authors' own research into the activity of ABC-transporters in NSCLC, in which the frequency of expression and co-expression of different transport proteins (including Pgp and MRP) is defined and new characteristics and mechanisms of the regulation of the functioning of MDRABC molecular markers are revealed. A methodical approach to routine characterization of MDRABC phenotype in NSCLC is offered. The main directions of pharmacological strategy of overcoming the multiple pharmacoresistance of NSCLC are formulated.