EXPRESSION OF HEPATOCYTE GROWTH FACTOR AND C-MET RECEPTOR IN STROMAL FIBROBLASTS AND TUMOR CELLS OF ENDOMETRIAL CARCINOMA.

BACKGROUND: HGF/c-Met is one of the main signaling pathways that ensure communication between epithelial cells and components of the tumor microenvironment determining the invasive and metastatic potential of many cancers. However, the significance of HGF and c-Met in endometrial carcinoma (ECa) progression remains unclear. AIM: To evaluate copy number variations as well as expression of the c-Met receptor and its ligand HGF in endometrial carcinomas considering the clinical and morphological characteristics of ECa. MATERIALS AND METHODS: The study was conducted on ECa samples of 57 patients, among which 32 had lymph nodes and/or distant metastasis. The copy number of c-MET gene was estimated by qPCR. The expression of HGF and c-Met in tissue samples was determined by the immunohistochemical method. RESULTS: Amplification of c-MET gene was detected in 10.5% of the ECa cases. In most carcinomas, a combined expression pattern of HGF and c-Met was established, in which co-expression of these markers was observed in tumor cells, and the content of HGF+ fibroblasts increased in the stroma. The expression of HGF in tumor cells was associated with the tumor differentiation grade and was higher in G3 ECa (p = 0.041). The number of HGF+ fibroblasts in the stromal component increased in the ECa cases with metastasis compared to the cases without metastasis (p = 0.032). The content of stromal c-Met+ fibroblasts was higher in deeply invasive carcinomas of patients with metastases than in tumors with invasion of < 1/2 myometrium (p = 0.035). CONCLUSION: Increased expression of HGF and c-Met in stromal fibroblasts of endometrial carcinomas is associated with metastasis in patients with ECa and deep invasion of the tumor into the myometrium, and can contribute to the aggressive course of the disease.

ROLE OF STROMAL MICROENVIRONMENT IN THE FORMATION OF INVASIVE, ANGIOGENIC, AND METASTATIC POTENTIAL OF ENDOMETRIOID CARCINOMA OF ENDOMETRIUM.

The aim of the study was to determine the association of indicators of the progression of endometrioid carcinoma of the endometrium (ECE) with the type of stromal microenvironment, the counts of CXCL12+ fibroblasts and CD163+ macrophages, and the expression of the chemokine CXCL12 and its receptor CXCR4 in tumor cells. MATERIALS AND METHODS: Histological preparations of ECE samples (n = 51) were analyzed. Expression of CXCL2 and CXCR4 antigens in tumor cells, the content of CXCL12+ fibroblasts and CD163+ macrophages, and the density of microvessels were determined by the immunohistochemical method. RESULTS: Groups of ECE with desmoplastic and inflammatory stromal reactions were delineated. The majority (80.0%) of tumors with desmoplasia were of low differentiation grade, deeply invading the myometrium; 65.0% of patients with these tumors were at stage III of the disease. In ECE cases of stages I-II, 77.4% of ECE showed an inflammatory type of stroma. The high angiogenic and invasive potential of EC of stages I-II was associated with an inflammatory stromal type, high counts of CD163+ macrophages and CXCL12+ fibroblasts in the tumor microenvironment, high expression of the chemokine receptor CXCR4, and reduced expression of its ligand CXCL12 in tumor cells. In the majority of EC of stage III, the increase in angiogenic, invasive, and metastatic potential was accompanied by the presence of desmoplastic stroma, increased expression of CXCR4 in tumor cells, and a high count of CXCL12+ fibroblasts. CONCLUSIONS: The obtained results showed that the morphological architecture of the stromal ECE component is related to the molecular features of its constituents and tumor cells. Their interaction modulates the phenotypic characteristics of ECE associated with the degree of malignancy.

VALIDATION OF A PANEL OF BIOMARKERS ASSOCIATED WITH AGGRESSIVE PHENOTYPE OF ENDOMETRIOID CARCINOMA OF ENDOMETRIUM.

AIM: To evaluate the prognostic significance of a panel of biomarkers for the identification of a highly malignant molecular subtype of endometrioid carcinoma of the endometrium (ECE). MATERIALS AND METHODS: The expression of a number of markers (CD24, CD44, E2F1, FOXP3, Her2/neu, p21WAF1/CIP1, p53, ß-catenin, vimentin, Е-cadherin, с-Myc, cyclins D1 and Е1) was determined by the immunohistochemical method in the samples of resected tumors of 127 patients with ECE of I-II stage. The Kullback method and the PanelomiX web tool were used to assess the informativeness and identify the aggressive subtype of ECE. The associative relationships of the studied markers were determined using the STRING v11 database. RESULTS: The study of the prognostic significance of a number of biomarkers in ECE has revealed the informativeness, high specificity and sensitivity (> 95%) of the Ñ€53+FOXP3-c-Myc+ phenotype, which is associated with a more aggressive tumor process. Bioinformatics analysis confirmed the correlative relationships between p53, FOXP3 and c-Myc, which are significant prognostic markers associated with cancer progression in ECE patients. CONCLUSIONS: The identified molecular phenotype of ECE (р53+FOXP3-c-Myc+) has differential and prognostic significance and objectively reflects a highly malignant subtype of this form of cancer.

Expression of chemokine receptor CXCR4 in tumor cells and content of CXCL12+-fibroblasts in endometrioid carcinoma of endometrium.

BACKGROUND: The expression of the CXCL12 chemokine and its receptor CXCR4 in the stromal component of the tumor plays an important role in tumor cell migration, proliferation, inhibition of apoptosis and determination of invasive and metastatic potential of malignant neoplasms of various genesis. The significance of CXCL12 and CXCR4 expression in endometrial tumor cells for cancer progression is not fully understood. AIM: To evaluate the content of CXCL12+-fibroblasts and expression of CXCL12 and CXCR4 in endometrial cancer cells, depending on the tumor stage. MATERIALS AND METHODS: Surgical material of 45 patients with endometrioid carcinoma of the endometrium (ECE) of the stages I-II and III was studied using morphological and immunohistochemical methods. RESULTS: In ECE of stage I-II CXCR4 expression was lower (43.3 ± 4.2%) while CXCL12 expression was higher (33.6 ± 2.4%) compared with the corresponding indices​​ in ECE of stage III (63.6 ± 3.5%, 24.5 ± 1.9%, respectively, p < 0.05). In ECE of stage III, high expression of CXCR4 (> Me) and low CXCL12 (< Me) was observed in 80% of samples; these tumors invaded more than 1/2 of the myometrium. There was a positive correlation between the depth of tumor invasion in the myometrium and the presence of metastases and CXCR4 expression in tumor cells (R = 0.5 and R = 0.4, respectively, p < 0.05) and the negative correlation with the expression of CXCL12 (R = -0.6 and R = -0.3, respectively, p < 0.05). In tumors that deeply invaded the myometrium, a high number of the CXCL12+-fibroblasts (> Me) (14.9 ± 1.3%) was detected. CONCLUSION: The obtained data reflect the communication of the immunosuppressive factor of the tumor microenvironment, i.e. CXCL12+-fibroblasts and CXCR4 expressing tumor cells. We suggest that the aggressiveness of ECE is determined by the combined effect of these two factors.

Expression of microRNA in tumor cells of endmetrioid carcinoma of endometrium.

BACKGROUND: It is known that more than half of the genes encoding human proteins are regulated by various microRNAs (miRNAs, miR), the expression of which may be associated with various pathological conditions. At the same time, the question of assessing the relationship between the expression of particular miRNAs and the aggressive molecular subtype of endometrial cancer remains open. Aim of the study was to determine the relationship between the expression of miR-34a, miR-125b, miR-142 and miR-101 in endometrioid carcinomas of the endometrium (ECE) and the features of the disease course. MATERIALS AND METHODS: The samples of surgical material of 51 patients with ECE (mean age 59.8 ± 7.1 years), I-III stage were investigated using morphological, immunohistochemical methods, real time polymerase chain reaction (PCR), cytofluorometry. RESULTS: In endometrial tumors with high proliferation index (< Me), the expression of miR-34a, miR-142 and miR-125b significantly decreased (1.8, 2.7 and 1.5 times, respectively) compared with those in ECE with low proliferation index (> Me). The expression of all studied miRNAs was lower in G3 tumors and those that deeply invaded the myometrium compared to G2 carcinomas and tumors with an invasion of > 1/2 myometrium and significantly decreased in tumors of patients with low stage III compared with stage I-II. The high (< Me) microvessel density in ECE was associated with a significant decrease of miR-125b and miR-101 expression, and the presence of signs of epithelial-mesenchymal transition - with a decreased expression of miR-34a and miR-101. CONCLUSIONS: The study revealed a significant heterogeneity of expression of miR-34a, miR-125b, miR-142 and miR-101 in ECE, which is associated with changes in morphofunctional characteristics of endometrial carcinoma.

Molecular phenotype of high-grade endometrioid carcinoma of the endometrium.

BACKGROUND: Prognosis of the course of tumor progression is one of urgent problems of clinical oncology. A relevant specificity of endometrial cancer is its clinical polymorphism within the same histological type of the disease. The search for molecular-biological features associated with the aggressive phenotype of endometrioid carcinomas is indisputably urgent. AIM: To study molecular-biological features of endometrioid carcinoma of the endometrium (ECE) and to identify the molecular subtype of tumors with high potential of malignancy. MATERIALS AND METHODS: Surgical specimens of 127 patients with EC, stages I-II, aged 36-72 (the average age - 59.3 ± 3.2) were studied using morphological and immunohistochemical methods. The multivariant analysis with the Kullback's informative measure and PanelomiX were used to estimate the significance of the expression of specific biomarkers. RESULTS: The expression of a complex of multifunctional markers was evaluated in ECE cells of different malignancy stage: p53, FOXP3, p21WAF1/CIP1, р16INK4a, E2F1, cyclins Е and D1, Her2/neu, с-Myc, Е-cadherin, ß-catenin, vimentin, CD44, CD24. A triad of biomarkers with threshold expression levels was determined (р53 < 45%; FOXP3 > 14%; с-Myc < 10%). The high expression of oncogene c-Myc and oncosuppressor p53 along with the low level of FOXP3 in tumor cells of ECE was associated with high proliferative potential, low differentiation grade, and deep invasion of a tumor into the myometrium. CONCLUSIONS: The molecular phenotype of ECE, most informative in terms of specificity and sensitivity (95%) - р5highFOXP3lowc-Mychigh, was first characterized, which would help identify a high-grade subtype of this cancer form.

Morphological characteristics and expression of adhesion markers in cells of low differentiated endometrial carcinoma.

The aim of the study was to evaluate the morphological features of endometrioid carcinoma of the endometrium (ECE) of low differentiation grade with different invasive potential and to characterize their molecular phenotype by the expression of a number of adhesion markers. MATERIALS AND METHODS: We have studied the samples of operation material of 37 patients with ECE of low differentiation grade with deep invasion (> ½ myometrium), n = 26, and with invasion < ½ myometrium, n = 11, with the use of morphological and immunohistochemical methods, and flow cytometry. RESULTS: In the morphological study of tumors with deep invasion in the myometrium, we have detected pronounced structural heterogeneity, which became the basis for the discretion of two groups of tumors with different characteristics of morphological phenotypes. In the majority of cases, solid layers and glandular-like structures are detected, and the similarity of the tumor epithelium with the elements of the endometrium is completely lost. In such tumors high expression of adhesion molecules - E-cadherin, CD44, CD24, and ß-catenin and low expression of the marker of mesenchymal tissues - vimentin were determined. Other tumors were characterized by morphological features of the epithelial-mesenchymal transition (EMT), with the decrease of the expression of E-cadherin, ß-catenin, CD24, CD44, and a significant increase in vimentin expression in comparison with these indices in tumors without signs of EMT. In ECEs that invade < ½ myometrium, the morphological indices of malignancy were less pronounced, which was associated by the changes in the expression of the molecular markers. CONCLUSION: This comprehensive study has established associations between the morphological heterogeneity of ECE and the expression of adhesion markers and vimentin, which is important for understanding the mechanisms of tumor cell migration.

Assessment of HER-2/neu, с-MYC and CCNE1 gene copy number variations and protein expression in endometrial carcinomas.

AIM: To analyze copy number variations of HER-2/neu, c-MYC and CCNE1 oncogenes and their protein expression in endometrioid endometrial carcinomas in relation to the degree of tumor progression and presence of a family history of cancer in cancer patients. MATERIALS AND METHODS: The study was conducted on endometrial cancer (EC) samples from 68 patients with I-II FIGO stages of disease. Copy number analysis of HER-2/neu, c-MYC and CCNE1 genes was performed by quantitative PCR. Protein expression was analyzed using immunohistochemistry. RESULTS: Assessment of copy number variations of HER-2/neu, c-MYC and CCNE1 genes revealed their amplification in the tumors of 18.8, 25.0 and 14.3% of EC patients, respectively. High expression of corresponding proteins was detected in 14.6, 23.5 and 65.6% of patients, respectively. It was established that HER-2/neu gene amplification is more common in the group of tumors of low differentiation grade than in moderate grade EC (35.7 and 5.5% of cases, respectively, p < 0.05). Also, high expression of c-Myc protein was more frequently observed in low differentiated tumors compared to the moderately differentiated EC (36.6 and 13.2% of cases, respectively, p < 0.05). Expression of HER-2/neu and cyclin E proteins was found to be dependent on the depth of tumor invasion into the myometrium. High expression of HER-2/neu protein was observed in 25.0 and 4.1% of EC patients with tumor invasion > ½ and < ½ of the myometrium, respectively, and cyclin E - in 86.7 and 46.6% of cases, respectively, p < 0.05. It was shown that among patients with a family history of cancer, a larger proportion of cases with high expression of c-Myc protein was observed compared to the group of patients with sporadic tumors (43.8 and 17.3%, respectively; p < 0.05). CONCLUSIONS: Amplification of HER-2/neu gene, along with high expression of c-Myc, HER-2/neu and cyclin E proteins, are associated with such indices of tumor progression as a low differentiation grade and deep myometrial invasion, suggesting the potential possibility of including these markers in the panel for determining the molecular EC subtype associated with an aggressive course of the disease. In a certain category of EC patients, there is a relationship between a family history of cancer and high expression of c-Myc protein.

Overexpression of the mitochondrial ribosomal protein S18-2 in the invasive breast carcinomas.

BACKGROUND: Recent studies allow to consider the mitochondrial ribosomal protein S18-2 (MRPS18-2, S18-2) as a potential oncoprotein, which suggests the need for further characterization of its expression in tumors of different genesis including breast cancer (BC). The aim of the study was to analyze the expression of the S18-2 protein in BC of luminal A and basal subtypes. MATERIALS AND METHODS: Operational material of BC patients stage І-ІІ (luminal A subtype, n = 30, and basal subtype, n = 10) was studied with the use of morphological, immunohistochemical, statistical and bioinformatic methods. RESULTS: Using the immunohistochemical analysis, we found that the S18-2 protein showed the nuclear signal in 66.7% of luminal A subtype BC samples and 80.0% of basal subtype BC samples. The variability of the S18-2 expression in both the luminal A and basal subtypes of BC was revealed. Noteworthy, the number of cells expressing S18-2 in high-proliferating tumors of luminal A and basal subtype is significantly higher than in tumors with a low proliferative potential (p < 0.05). In 10 samples of luminal A subtype, the nuclear S18-2 signal was higher than median value. Moreover, the S18-2 protein was overexpressed in 4 out of such 10 samples. Metastases in the lymph nodes were found in 3 out of 4 patients with the stage II BC, low differentiation grade of the tumor and high proliferative activity. The bioinformatic analysis confirms our preliminary findings that the trend for increasing expression of the S18-2 protein in tumors correlates with the aggressiveness of malignant BC. CONCLUSION: The S18-2 protein may be a marker of cancer aggressiveness in BC patients.

Markers of the epithelial-mesenchymal transition in cells of endometrial carcinoma.

AIM: To study the expression of adhesion markers (E-cadherin, ß-catenin and vimentin) associated with epithelial-mesenchymal transition (EMT) and their role in progression of endometrial carcinoma (EC). MATERIALS AND METHODS: Expression of E-cadherin, ß-catenin and vimentin was studied immunohistochemically in the samples of surgical material of 55 EC patients stage I-III. The proliferation index was determined by flow cytometry. RESULTS: In the group of vimentin-negative EC, tumors of low differentiation grade and deep invasion in myometrium as well as high expression of E-cadherin and ß-catenin prevailed compared with the cases with high expression of vimentin. In addition, in EC with high expression of vimentin, an increase in the number of cells with expression of E-cadherin in the cytoplasm (78.9 ± 3.6%) and ß-catenin with cytoplasmic-nuclear localization (73.7 ± 3.2%) was observed compared with these indices in vimentin-negative tumors (45.4 ± 4.2%, p < 0.001 and 54.5 ± 2.6%, respectively, p < 0.005), which may indicate EMT-associated changes in EC with high expression of vimentin. CONCLUSIONS: The progression of the endometrioid carcinoma may occur in the setting of various molecular changes, in particular, with decreased expression of E-cadherin and ß-catenin and high expression of vimentin, or in the absence of vimentin, utilizing other mechanisms of regulation of proliferative and metastatic potential.

Sensitivity to 4-hydroxyestradiol and DNA repair efficiency in peripheral blood lymphocytes of endometrial cancer patients.

BACKGROUND: The development of hormone-dependent cancers, including endometrial carcinomas, in great part may be mediated by the genotoxic effects of estrogen metabolites, among which 4-hydroxyestradiol (4OHE2) is characterized by the most prominent DNA-damaging properties. It is assumed that the individual sensitivity to the 4OHE2 may determine the predisposition to endometrial cancer (EС). AIM: To analyze the sensitivity of peripheral blood lymphocytes (PBLs) of EC patients to the 4OHE2 and to evaluate the repair efficiency of 4OHE2-induced DNA damage. MATERIALS AND METHODS: The study was performed on the PBLs of 53 EC patients and 20 healthy women. The level of DNA damage was measured using the comet assay and was expressed as % tail DNA. The DNA repair efficiency (%) was evaluated by determining the ratio between the amount of repaired DNA damage and the level of 4OHE2-induced damage that appeared after incubation of PBLs with 4OHE2. RESULTS: In PBLs of EC patients, a higher level of 4OHE2-induced DNA damage (32.0 ± 2.2% tail DNA) and lower DNA repair efficiency (34.0 ± 4.5%) was observed compared to PBLs of healthy women (22.3 ± 2.3% tail DNA and 48.8 ± 4.5%, respectively). PBLs of EC patients with deep tumor invasion of myometrium were characterized by more prominent decrease of DNA repair than those with less invasive tumor (< ½ of myometrium) (20.9 ± 7.8 and 43.7 ± 6.7%, respectively). Furthermore, lower DNA repair efficiency was detected in the PBLs of EC patients with a family history of cancer compared to this parameter in patients with sporadic tumors (20.9±7.8 and 47.1 ± 5.5%, respectively). CONCLUSION: The PBLs of EC patients are characterized by increased sensitivity to the genotoxic effect of 4OHE2 and reduced repair efficiency regarding 4OHE2-induced DNA damage. A lower level of DNA repair is observed in EC patients with deep tumor myometrial invasion and a family history of cancer.

Phenotypic features of endometrial tumors in patients with family history of cancer.

AIM: To determine the peculiarities of expression of a number of proteins-regulators of the cell cycle in endometrial cancer (EC) cells in patients with a family history of oncological pathologies. PATIENTS AND METHODS: 95 EC patients (stage І-ІІ) were included into the study. Clinical-genealogical analysis was performed. 54 patients (group I) had healthy relatives, and in families of 41 patients (group II) an aggregation of malignant tumors of different genesis (mainly tumors of the gastrointestinal tract and the female reproductive system) was recorded. p53, p21WAF1/CIP1, p16INK4a, and Ki-67 were assessed immunohistochemically in the surgical samples. RESULTS: In the majority of patients, both from group I and II, moderately differentiated tumors were observed (in 38.9 and 46.3% of cases, respectively), mainly with deep myometrium invasion (64.8 and 58.5% of cases, respectively). In EC patients from group II, a significantly higher number of p16INK4a-positive cells (17.7 ± 1.7%; p = 0.001) and lower number of p53-positive (30.9 ± 3.2%; p = 0.05) and Ki-67-positive (26.9 ± 2.7%; p = 0.048) cells was observed compared to those in tumors of patients from group I (12.0 ± 1.6; 37.7 ± 2.8 and 36.7 ± 3.4%, respectively). CONCLUSION: Phenotypic features of the EC in the patients with family history of cancer differ from those in tumors of patients without such aggregation. The biological heterogeneity of EC seems to relate to the oncogenealogical history of patients. Also this biological heterogeneity is linked to the molecular features of EC cells, which affects cancer aggressiveness and the course of the disease.

DNA damage in tumor cells and peripheral blood lymphocytes of endometrial cancer patients assessed by the comet assay.

To date, genome instability is considered to be a common feature not only of tumor cells, but also of non-malignant cells of cancer patients, including peripheral blood lymphocytes (PBLs). The issue of the association between genome instability in tumor cells and PBLs, as well as of its relationship with tumor progression remains poorly understood. AIM: To evaluate the level DNA damage in tumor cells and PBLs of endometrial cancer (EC) patients with regard to clinical and morphological characteristics of the patients. MATERIALS AND METHODS: DNA damage was assessed in 106 PBLs samples and 42 samples of tumor cell suspension from EC patients by comet assay. PBLs from 30 healthy women were used as control. The level of DNA damage was expressed as the percentage of DNA in the comet tails (% tail DNA). RESULTS: It was revealed that the amount of DNA damage in PBLs of EC patients was 2.2 times higher in comparison with that of healthy donors (8.3 ± 0.7 and 3.7 ± 0.4% tail DNA, respectively) (p < 0.05). In this study, no association between the levels of DNA damage in endometrial tumor cells and PBLs was observed (r = 0.11; p > 0.05). The amounts of DNA damage both in tumor cells and PBLs were not related to the degree of tumor differentiation as well as the depth of myometrial invasion, but depended on the body mass index (BMI) of EC patients: high level of lesions was observed in patients with elevated BMI values. Furthermore, the level of DNA damage in tumor cells was associated to familial aggregation of cancer and was significantly higher in endometrial cells from patients with family history of cancer vs that from EC patients with sporadic tumors (32.3 ± 2.9 and 22.8 ± 1.8% tail DNA, respectively) (p < 0.05). It was also found that for women who had high level of DNA damage in PBLs, the risk of EC was greater (odds ratio value of 3.5) compared to those with low level of such lesions. CONCLUSION: Genome instability that appears as an increased level of DNA damage in tumor cells and PBLs of EC patients is associated with BMI and family history of cancer and can reflect a predisposition to cancer.

FOXP3 gene promoter methylation in endometrial cancer cells.

AIM: To determine the methylation level of promoter region of the FOXP3 gene promoter depending on the heterogeneity of intracellular localization of its protein product in endometrial cancer (EC) cells and assess its relation to the clinical and morphological features of tumor. MATERIALS AND METHODS: Samples of surgical material of 40 EC patients who have not received any specific treatment before the surgery, were studied. Real time methylation-specific PCR (MSP) as well as morphological and immunohistochemical methods were used in the study. RESULTS: Methylation of promoter region of the FOXP3 gene was determined in all EC cases, but variability of the methylation level in EC cells from 45.0% to 85.0% was observed. With tumor progression and in tumors with deep (≥ 1/2) invasion in myometrium, an increase of the methylation level of the FOXP3 and of cell number with cytoplasmic FOXP3 localization was observed. In EC patients the correlation between of methylation level of the FOXP3 gene and the number of FOXP3(+) tumor cells with cytoplasmic expression (r = 0.41) was determined. CONCLUSION: The methylation level of FOXP3 gene promoter region and intracellular localization of its protein product are associated with tumor differentiation grade and the depth of myometrial invasion.

The study of mismatch repair in endometrial cancer patients with a family history of cancer.

AIM: To assess the expression of mismatch repair (MMR) proteins MSH2 and MLH1 and carry out microsatellite analysis in patients with endometrial cancer (EC) with regard to the family history of cancer. MATERIALS AND METHODS: Morphological and immunohistochemical study was performed on tumor tissue samples of 49 EC patients. Microsatellite instability was determined using PCR with primers which flank microsatellite region BAT-26. RESULTS: A tendency to a decreased expression of both MSH2 and MLH1 markers in a group of EC patients with a family history of cancer as compared with a group without aggregation of cancer in family history was observed (labeling index - LI - was 36.1 ± 8.1% and LI 20.7 ± 9.1% versus LI 48.0 ± 5.8% and 33.8 ± 5.8%, respectively). It was determined that the number of EC patients with tumors deficient by expression of MMR markers was reliably higher in a group of patients with a family history of cancer than in a group of patients without aggregation of cancer in family history (р < 0.05). It was shown that in a group of EC patients with a family history of cancer, MMR-proficient tumors were detected in 38.5% of cases. Microsatellite instability was determined in 10.7% of EC patients including one patient with aggregation of Lynch-associated tumors in family history. CONCLUSION: Family history of cancer of EC patients is associated with malfunctioning of the MMR system as well as may be related to alternative molecular mechanisms.

Comprehensive analysis of intratumoral lymphocytes and FOXP3 expression in tumor cells of endometrial cancer.

AIM: To study the tumor microenvironment (CD4(+), CD8(+) and FOXP3(+) lymphocytes) and FOXP3 expression by tumor cells and correlation of studied parameters with clinical and morphological characteristics of endometrial adenocarcinomas. MATERIALS AND METHODS: Tumor samples from 40 patients (mean age 56.9 ± 2.8) with endometrial cancer (EC), who did not receive special treatment before surgery (chemotherapy, radiation therapy and hormontherapy), were investigated. Morphological, immunohistochemical methods as well as methods of mathematical statistics were applied in the study. RESULTS: It has been determined that high quantity of FOXP3(+) tumor cells and intratumoral CD4(+) and CD8(+) Т-lymphocytes along with the low content of FOXP3(+)-lymphocytes is typical for the endometrial adenocarcinomas of high differentiation grade (G1). In poorly differentiated (G3) EC an increase of number of FOXP3(+)-lymphocytes and decrease of CD4(+) and CD8(+) lymphocytes in lymphocytic infiltrate have been observed. Moreover, decrease of the content of FOXP3(+) tumor cells has been determined. In EC patients correlation between the following parameters has been detected: proliferative activity and deep invasion of tumor in myometrium (R = 0.74); depth of invasion correlated with the number of the FOXP3(+) tumor cells (R = -0.63) and number of CD4(+) and CD8(+) lymphocytes (R = 0.68 and R = -0.55 respectively) in lymphocytic infiltrate. Thus, results of this study are the evidence of significance of the lymphocytic components of tumor microenvironment and content of FOXP3 expressing tumor cells in EC progression. CONCLUSION: Quantitative changes of tumor microenvironment, such as number of CD4(+), CD8(+) and FOXP3(+) lymphocytes and content of FOXP3(+) tumor cells correlate with biological characteristics of EC.

Analysis of p53, p16(INK4a), pRb and Cyclin D1 expression and human papillomavirus in primary ovarian serous carcinomas.

AIM: To evaluate the prognostic relevance of key cell cycle regulatory proteins p53, p16(INK4a), pRb and Cyclin D1 expression, the presence of high risk HPVs and their association with clinicopathological parameters and the clinical follow up in ovarian cancer patients. METHODS: 53 cases of primary ovarian serous carcinomas were immunohistochemically examined for the expression of p53, p16(INK4a), pRb and Cyclin D1 proteins. Tumor DNA was extracted from paraffin blocks and subjected to HPV 16 and 18 testing. The association between HPV 16 and 18 E6 oncoprotein and cell cycle proteins expression in ovarian carcinomas also was evaluated by immunohistochemistry. RESULTS: We demonstrated that a majority of moderately and poorly differentiated ovarian carcinomas are characterized by strong expression of p53 and p16(INK4a) proteins. In contrast, strong staining with cyclin D1 antibody was observed in well differentiated tumors. The correlation between strong p53, pRb, Cyclin D1 and clinical stages of disease was also observed. We show that patients with high positivity for p53, p16(INK4a) and Cyclin D1 had a poor prognosis and reduced overall survival. The presence of HPV 16/18 DNA was detected in 17% of ovarian carcinomas. The tumor tissues that reacted positively to HPV E6 antibody in focal and diffuse manners had also significantly low p53 expression profile. CONCLUSION: These findings suggest that p53, p16(INK4a) and Cyclin D1 expression and HPV infection may represent a promising tool toward the identification of ovarian cancer patients with poorer prognosis and shorter survival who might therefore need a more aggressive therapy and HPV screening.

Expression of the estrogen and progesterone receptors as prognostic factor in serous ovarian cancers.

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.

Expression of p53, p21WAF1/CIP1, p16INK4A and Ki-67 proteins in serous ovarian tumors.

AIM: The analysis of p53, p21(WAF1/CIP1), p16(INK4A) and Ki-67 expression in serous ovarian carcinomas of different grade. MATERIALS AND METHODS: In total, 43 ovarian adenocarcinomas and 8 non-altered ovarian epithelial tissues were immunohistochemically investigated for expression of Ki-67, p53, p21(WAF1/CIP1) and p16(INK4A). RESULTS: It has been shown that expression of Ki-67, p53, p21(WAF1/CIP1) and p16(INK4A) in non-altered ovarian epithelial tissue is absent. Serous ovarian carcinomas are characterized by high proliferative activity (PI Ki-67 = 30.0 +/- 0.3%), p53 and p16(INK4A) overexpression (LI is 40.3 +/- 0.3% and 31.1 +/- 0.6% respectively) and low expression of p21(WAF1/CIP1) (LI = 6.8 +/- 0.3%). The association between expression of these markers and ovarian tumor grade was defined: the maximal level of Ki-67, p53 and p16/(INK4A) and minimal of p21(WAF1/CIP1) expression were observed in G3 tumors. So, low p21(WAF1/CIP1) expression (LI < 7.0%) combined with p16(INK4A) overexpression is considered to be the factor for a poor prognosis in serous ovarian cancer. CONCLUSIONS: The present study has indicated that biomolecular markers of cell proliferation along with traditional clinical and morphologic characteristics can be used for differential diagnostics of ovarian tumors.

Different trends of p53, MDM2 and p14 ARF expression patterns in endometrial adenocarcinomas versus hyperplasia.

AIM: To study the expression of p53, MDM2, and p14 ARF in the highly, moderately and low differentiated endometrial adenocarcinomas, compared to hyperplasia. MATERIAL AND METHODS: Surgical material and the scrapes of endometrial cancer, glandular and atypical hyperplasia patients. Expression of 53, MDM2 and p14 ARF was evaluated by immunohistochemical method using respective monoclonal antibodies. RESULTS: High p53 expression level is accompanied by decreased level of MDM2 expression in endometrial cancers. On contrary, in endometrial hyperplasia, there was clear connection between the expression levels of p53 and MDM2. We hypothesize that the high p53 and low MDM2 levels in endometrial cancers could arise due to the inhibition of transcriptional activity of p53 by its binding to estrogen receptors. CONCLUSION: High p53 expression level with low MDM2 and p14 ARF levels may be the characteristic features of low differentiated endometrial carcinoma.