The data of a complex immunoassay comparative study of the content of soluble forms of sPD-1, sPD-L1, sNKG2D, sNKG2DL1, sB7-H3 and sHLA-G in the blood plasma of 75 patients with epithelial ovarian cancer and 20 healthy donors of the control group are presented. The diagnostic significance of the studied proteins was determined. The study showed that the profile of soluble immunity checkpoints differs when malignant ovarian pathology occurs. There was a statistically significant decrease in the content of sPD-L1, sNKG2DL1, sB7-H3, and sHLA-G in the blood plasma of patients compared with the control group. Differences were found in the content of the studied markers depending on the histological type of tumors. Correlations between the soluble forms of some of the studied proteins are shown, indicating the presence of independent mechanisms of immune regulation in ovarian cancer, which may explain the insufficient effectiveness of the existing immunotherapy for this type of tumor. The results obtained will undoubtedly facilitate the development of new effective methods for the diagnostics and therapy of ovarian cancer.

Aim: To perform a comparative analysis with simultaneous measurement of vascular endothelial growth factor (VEGF), insulin-like growth factor 1 (IGF1) and matrix metalloproteinase 7 (MMP7) in serum samples taken from healthy women and ovarian cancer patients; to perform association of these markers with their expression in primary tumors depending on clinical, morphological and biochemical characteristics of the disease and its prognosis. Materials and methods: We assessed 54 treatment-naïve patients with ovarian cancer aged from 23 to 74 years (mean ± SD, 53.2 ± 1.9), being at various FIGO stages of the disease. The control group consisted of 120 healthy women of matched age and reproductive status, in whom serum biomarker levels were studied. Patient survival was assessed by the Kaplan-Meier method, with survival curves compared with log-rank test. All analyses were done with “STATISTICA” and SPSS software. Results: Serum VEGF levels in ovarian cancer patients were significantly (p < 0.0001) higher compared those in the control. The most informative cut-off values differentiating the groups studied were serum VEGF values of < 350 pg/ml (median value in the control) and > 505 pg/ml (upper quartile in the control). With 505 pg/ml taken as a threshold, the test had sensitivity of 79.6% and specificity of 75%. Another cut-off value of serum VEGF level between the patients with ovarian cancer and the control group (510 pg/ml) was derived from ROC curves and 75% sensitivity and 78.2% specificity. No acceptable cut-off value for serum IGF1 to differentiate between the patients with ovarian cancer and the controls could be obtained from the ROC curves. Serum MMP7 levels in the patients with ovarian cancer were significantly higher than those in the control group (Mann-Whitney test p < 0.0001). With ROC curves, the best sensitivity to specificity ratio for MMP7 value of 4.6 ng/ml was obtained to differentiate between the patients with ovarian cancer and the controls (sensitivity 83.3%, and specificity 81%). The variance analysis did not reveal any association between serum VEGF, IGF1 and MMP7 and age of patients with ovarian cancer, tumor histology, concomitant somatic and gynecological diseases, and CA-125 levels. Serum VEGF and IGF1 levels did not correlate with the stage of ovarian cancer, in contrast to MMP7, whose levels were significantly higher in stages IIIc–IV. The median VEGF level significantly increased as the degree of differentiation decreased from 510 to 622 pg/ml (p < 0.002), while median IGF1, on the contrary, decreased from 219 to 116 pg/ml (p < 0.0001). There was a direct correlation between serum and tumor VEGF levels in ovarian cancer patients (r = 0.65, p < 0.0001). On the contrary, there was an inverse correlation between serum and tumor IGF1 levels (r = -0.68, p < 0.0001). Serum and tumor MMP7 levels remained unrelated to each other. Tumor VEGF, IGF1 and MMP7 content was unrelated to the age of the patients, their reproductive status, presence of concomitant somatic and gynecological diseases, histology of ovarian cancer, and serum CA-125 levels. VEGF levels in the tumor were not associated with the stage of ovarian cancer, but in patients with initial stages Ia and Ib stages MMP7 values significantly lower (2.1 ng/mg protein) compared to those in stages IIIc and IV (6.1 and 4.7 ng/mg protein, respectively, p < 0.05). Similar pattern was noted for IGF1: tumor IGF1 values in the patients with stages Ia–Ib were significantly lower (0.5 ng/mg protein) than those with stages IIIc–IV (median, 1.3–1.4 ng/mg protein). A significant increase in both serum and tumor VEGF levels was detected in the patients with ovarian cancer with decreased degree of differentiation. On the contrary, tumor IGF1 levels, but not serum ones, were significantly increased from 0.6 to 1.4 ng/ml in the patients with poorly differentiated ovarian cancer. MMP7 tumor expression did not depend on the degree of its differentiation. Serum VEGF levels above 700 pg/ml and tumor levels of above 590 ng/mg protein should be considered as unfavorable prognostic factors in patients with ovarian cancer.

Матриксные металлопротеиназы (ММП) - ферменты класса гидролаз, осуществляющие ферментативный катализ с помощью связанного в активном центре иона цинка. Функции ММП разнообразны, и нарушение баланса их активности может быть одним из этиологических факторов различных заболеваний. В данном обзоре рассмотрена классификация ММП человека, особенности их структуры и регуляции, а также роль в физиологических и патологических процессах в организме человека. Приведен перечень наиболее изученных на настоящий момент полиморфных вариантов генов MMП, описаны их функциональные эффекты и представлены результаты ассоциативных исследований. Matrix metalloproteinases (MMPs) are enzymes of the hydrolase class that carry out enzymatic catalysis with the help of a zinc ion bound in the active center. MMP functions are diverse, and a disturbance in the balance of their activity may be one of the etiological factors of various diseases. In this review, the classification of human MMP, the features of their structure and regulation, as well as the role in physiological and pathological processes in the human body are considered. A list of the most studied polymorphic versions of MMP genes has been given, their functional effects have been described, and the results of associative studies have been presented.

IGF-I, II, IGFBP-1, 2 and 3 levels were measured with standard ELISA kits (Mediagnost) in blood serum of 44 ovarian cancer, 12 benign and 11 borderline ovarian tumorpatients. Control group comprised 33 practically healthy women. Serum IGF-1 content in ovarian cancer patients was significantly lower, and IGFBP-1 content higher than in all other groups. IGFBP-2 level was increased both in ovarian cancer and borderline tumor groups as compared to control and benign ovarian tumor patients. No significant associations were found between the majority ofparameters studied and main clinico-pathologic characteristics of ovarian cancer. Thus, disturbances in IGFs/IGBPs balance were revealed in blood serum of ovarian cancer patients, and IGFBP-2 proved to be a potential diagnostic serological marker with 90% specificity and 90% sensitivity.

К настоящему времени сформулированы основные и дополнительные признаки, отличающие опухолевую клетку от клетки нормальной ткани, однако эти характеристики изменяются и дополняются в результате значительного прогресса, достигнутого в последние годы в области экспериментальной онкологии, молекулярной генетики и биохимии [1—3]. Одним из наиболее выдающихся открытий в биологии последнего десятилетия следует считать обнаружение системного уровня регуляции активности генов с помощью малых некодирующих молекул — микроРНК [4]. Подавление экспрессии генов с участием микроРНК считают важным механизмом, вовлеченным в большинство внутриклеточных сигнальных путей у многих эукариот. Нарушение этого механизма обнаружено при различных патологиях, в том числе и при развитии опухоли [4]. МикроРНК представляют собой небольшие молекулы, транскрибируются с геномной ДНК, подвергаются процессингу и экспорту в цитоплазму. Они могут входить в состав транскриптов, кодирующих белки, либо транскрибироваться с белок-некодирующих участков. Первый процессинг происходит с участием специализированного ферментного комплекса либо в ходе стандартного сплайсинга мРНК. После экспорта в цитоплазму промежуточный продукт подвергается окончательному процессингу с образованием активного РНК-белкового комплекса, способного связываться с комплементарными участками мРНК-«мишеней». Результатом такого связывания является подавление трансляции с данной мРНК. Сама мРНК может быть расщеплена за счет РНКазной активности комплекса. Известdoi: 10.17116/repro201521330-37

The comparative analysis of the quantity of matrix metalloproteinases (MMP) 2,7 and 9 in breast cancer is carried out; the concentration of sex hormones receptors and protein HER-2/neu are determined. Authentically high values MMP-2 and MMP-7 are found in a tumor tissue in contrast to the normal one. The elevated concentration of ММР-2 in a tumor is connected with receptor status (p < 0,05). The highest values of concentration ММР-7 and ММP- 9 are found at RE-, RP- and HER-2/neu + tumors, and also in the tumors with the size more than 4,0 cm.

Enzyme immunoassay showed that the content of matrix metalloproteinases (MMP) 2 and 7 in tumors was higher than in the adjacent histologically intact tissue in 91 and 76% patients with breast cancer, respectively, while MMP-9 levels in the tumor and intact tissue were virtually the same. Serum concentrations of MMP-2 and MMP-7 did not correlate with their levels in the tumors, were within the normal range, and virtually did not decrease after removal of the primary tumor. Serum levels of MMP-9 in patients were significantly lower than in the control and increased after surgery in 85% patients. No clear-cut relationship between the studied parameters and clinical morphological prognostic factors of breast cancer was detected.

The content of vascular endothelium growth factor is significantly increased, while the level of matrix metalloproteinase-2 is 2-fold reduced in ovarian cancer tissue compared to benign tumors. A trend to an increase in the levels of matrix metalloproteinases 7 and 9 and reduction of vascular endothelial growth factor type 2 receptors in tumor tissue was also detected. A highly significant negative correlation between the levels of vascular endothelial growth factor and matrix metalloproteinase 2 and positive correlations between vascular endothelial growth factor and matrix metalloproteinase 7, vascular endothelial growth factor and matrix metalloproteinase 9, matrix metalloproteinase 2 and vascular endothelial growth factor type 2 receptors were revealed. In the tumors assayed after preoperative therapy, relative normalization of the studied parameters was observed: the level of vascular endothelial growth factor decreased significantly, while the levels of matrix metalloproteinase 2 and vascular endothelial growth factor type 2 receptors increased. The levels of the markers differed significantly in ovarian tumors of different histological types, and the levels of vascular endothelial growth factor type 2 receptors were higher in patients with stage III compared to stage I and the content of matrix metalloproteinase 7 was higher in stage III compared to stage II cancer.

Here we present the results of evaluation of the expression of neural cell adhesion molecules CD56 (NCAM) in serous ovarian adenocarcinoma. The expression was detected in 48.5% cases. Infiltration of tumor stroma and parenchyma with CD8+ и CD4+ lymphocytes was significantly less pronounced in tumors expressing neural cell adhesion molecules; CD3+CD4+CD25+ predominate among CD4+ lymphocytes in CD56+ tumors. CD56+ tumors were lower in size (5.2±0.6, 7.9±0.8 and 10.3±1.5 cm in monomorphic, mosaic, and negative phenotypes, respectively (p=0.05) and were characterized by the absence of cystic component (p=0.012), larger disseminations in the peritoneum (4.2±1.1 and 2.7±0.5 cm; p=0.05), and larger volume of the residual tumor (p=0.018) after surgical treatment. NCAM phenotype of the tumor does not correlate with the stage and differentiation degree of serous ovarian adenocarcinoma.

The content of matrix metalloproteinases 7 and 9 was significantly increased, while the content of metalloproteinase 2 was reduced in ovarian cancer tissue compared to benign tumors. In blood serum from patients with ovarian cancer, the concentrations of matrix metalloproteinases 7 and 9 and their type 1 tissue inhibitor were significantly elevated, while the concentration of matrix metalloproteinase 2 was reduced compared to the corresponding parameters in healthy women. After chemotherapy, tissue and serum concentrations of metalloproteinases and their inhibitor in patients practically returned to normal. A significant positive correlation between serum levels of matrix metalloproteinases 7 and 9 and tissue inhibitor of metalloproteinases-1 in patients with ovarian cancer and the size of primary tumor (ultrasound examination) and a positive correlation between these parameters and the concentration of classical ovarian cancer marker CA-125 were demonstrated.

Here we present the results of comparative immunoenzyme assay of the initial serum levels of VEGF in breast cancer patients (stages T 1N 0M 0 and T 2N 0M 0) and apparently healthy women (controls). It was found that VEGF concentrations in the serum of patients with breast cancer stages T 1N 0M 0 and T 2N 0M 0 significantly surpassed the control levels. Increased levels of VEGF surpassing the threshold values were more often observed in patients with T 2N 0M 0 breast cancer compared to patients with T 1N 0M 0 tumor. Serum concentration of VEGF in patients with stages T 1N 0M 0 and T 2N 0M 0 breast cancer did not depend on patient’s age and reproductive function and receptor status of the primary tumor (estrogen and progesterone receptors), but was closely associated with tumor histogenesis and differentiation degree. Significantly higher levels of VEGF were observed in patients with lobular infiltrative breast carcinoma compared to patients with ductal tumors and in patients with low-differentiated tumors compared to highly and moderately differentiated tumors. High initial concentrations of VEGF (≥300 pg/ml) were more often detected in patients with T 2N 0M 0 breast cancer developing relapses within the fi rst 3 years of follow-up compared to patients without relapses during the corresponding period (p=0.001). These findings suggest that serum level of VEGF in patients with T 2N 0M 0 breast cancer before treatment can be used as an additional marker in parallel with standard clinical and morphological signs of the disease for more precise prognosis of early relapse.

A significant increase of matrix metalloproteinases’ (MMP) 2 and 7 levels in the tumors as compared to adjacent histologically unchanged mammary gland was demonstrated by enzyme-linked immunosorbent assays in 91 and 76% of breast cancer patients respectively, while MMP-9 levels in these tissues did not differ significantly. The concentrations of MMP-2 and MMP-7 in blood serum were not associated with corresponding tumor values, did not exceed the control levels, and did not decrease after the excision of primary tumor. Serum MMP-9 was significantly decreased in breast cancer patients as compared to control group, and increased post-surgically in 85% of the patients. No unambiguous associations between the parameters studied and breast cancer clinico-morphologic prognostic factors were revealed.

Concentration and DNA-binding activity of nuclear transcription factor NF-κΒ p65 subunit were measured by quantitative immunoenzymatic methods in the tumors and histologically unchanged mammary gland tissues of 119 breast cancer patients. Enhanced as compared to adjacent tissue NF-kBp65 DNA-binding activity was revealed in 97% of the tumors, and in most cases it was associated with an increase of total NF-kBp65 content. No significant associations were found between the parameters studied and disease stage, tumor size and histology, lymph node status, but NF-kBp65 content was significantly increased in grade III as comparedto grade II tumors. HER2 + tumors were characterized by an increase of total NF-kBp65 expression independent on steroid receptor status and not accompanied by enhancement of DNA-binding activity, the latter being the highest in “triple receptor negative” (ER -PR -HER2 -) tumors.