The search for interacting long non-coding RNAs (lncRNAs), microRNAs (miRNAs), and mRNAs of protein-coding genes through the mechanism of competing endogenous RNAs in tumors of ovarian cancer patients was carried out. The levels of expression of 24 lncRNAs, 20 miRNAs, and 28 mRNAs of protein-coding genes involved in oncogenesis were determined by real-time PCR on a set of representative samples. Correlations between lncRNAs/miRNA and miRNA/mRNA levels in ovarian cancer samples were analyzed. We identified 8 pairs of lncRNAs/miRNA and 17 pairs of miRNA/mRNA, the expression levels of which have a negative correlation. Five triplets of potentially interacting lncRNAs/miRNA/mRNA have been identified, among which the most significant triplet is the OIP5-AS1/miR-203a-3p/ZEB1. The data obtained determine new epigenetic profiles, as well as new potential biomarkers and targets for targeted therapy of ovarian cancer patients.

The role of methylation of 9 miRNA genes in the pathogenesis of metastatic clear cell renal cell carcinoma was determined by quantitative methylation-specific PCR (MS-PCR). For 5 genes (MIR125B-1, MIR137, MIR193A, MIR34B/C, and MIR375), a significant correlation of high methylation level with late (III-IV) stages, large size (T3+T4) of the tumor, and metastasis to lymph nodes and/or distant organs was revealed. For another group of genes (MIR125B-1, MIR1258, MIR193A, MIR34B/C, and MIR375), a statistically significant correlation of high methylation level with loss of differentiation in the tumor (G3-G4) was found, and the opposite pattern was found for MIR203A. A total of 7 microRNA genes (MIR125B-1, MIR1258, MIR137, MIR193A, MIR203A, MIR34B/C, and MIR375) were identified, the methylation of which is associated with the progression of metastatic clear cell renal cell carcinoma. For 6 of them (except MIR34B/C) these data were obtained for the first time. Thus, new factors of the development and progression of clear cell renal cell carcinoma were identified as potential biomarkers for the early diagnosis and prognosis of metastatic clear cell renal cell carcinoma.

To date, certain problems have been identified in cancer immunotherapy using the inhibition of immune checkpoints (ICs). Despite the excellent effect of cancer therapy in some cases when blocking the PD-L1 (programmed death-ligand 1) ligand and the immune cell receptors PD-1 (programmed cell death protein 1) and CTLA4 (cytotoxic T-lymphocyte-associated protein 4) with antibodies, the proportion of patients responding to such therapy is still far from desirable. This situation has stimulated the exploration of additional receptors and ligands as targets for immunotherapy. In our article, based on the analysis of the available data, the TIM-3 (T-cell immunoglobulin and mucin domain-3), LAG-3 (lymphocyte-activation gene 3), TIGIT (T-cell immunoreceptor with Ig and immunoreceptor tyrosine-based inhibitory motif (ITIM) domains), VISTA (V-domain Ig suppressor of T-cell activation), and BTLA (B- and T-lymphocyte attenuator) receptors and their ligands are comprehensively considered. Data on the relationship between receptor expression and the clinical characteristics of tumors are presented and are analyzed together with the results of preclinical and clinical studies on the therapeutic efficacy of their blocking. Such a comprehensive analysis makes it possible to assess the prospects of receptors of this series as targets for anticancer therapy. The expression of the LAG-3 receptor shows the most unambiguous relationship with the clinical characteristics of cancer. Its inhibition is the most effective of the analyzed series in terms of the antitumor response. The expression of TIGIT and BTLA correlates well with clinical characteristics and demonstrates antitumor efficacy in preclinical and clinical studies, which indicates their high promise as targets for anticancer therapy. At the same time, the relationship of VISTA and TIM-3 expression with the clinical characteristics of the tumor is contradictory, and the results on the antitumor effectiveness of their inhibition are inconsistent.

In the immunotherapy based on immune checkpoint inhibition (IC), additional ICs are being studied to increase its effectiveness. An almost unstudied feature is the possible co-expression of ICs, which can determine the therapeutic efficacy of their inhibition. For the selection of promising ICs, information on the association of their expression with cancer development may be essential. We have obtained data on the expression correlation of ADAM17, PVR, TDO2, CD274, CD276, CEACAM1, IDO1, LGALS3, LGALS9, and HHLA2 genes in gastric cancer (GC). All but one, TDO2, have other IC genes with co-expression at some stage. At the metastatic stage, the expression of the IDO1 does not correlate with any other gene. The correlations are positive, but the expressions of the CD276 and CEACAM1 genes are negatively correlated. The expression of TDO2 and LGALS3 is associated with GC metastasis. The expression of TDO2 four-fold higher in metastatic tumors than in non-metastatic tumors, but LGALS3 was two-fold lower. The differentiation is associated with IDO1. The revealed features of TDO2, with a significant increase in expression at the metastatic stage and the absence of other IC genes with correlated expression indicates that the prospect of inhibiting TDO2 in metastatic GC. IDO1 may be considered for inhibition in low-differentiated tumors.

Our work aimed to differentiate 20 aberrantly methylated miRNA genes that participate at different stages of development and metastasis of ovarian carcinoma (OvCa) using methylation-specific qPCR in a representative set of clinical samples: 102 primary tumors without and with metastases (to lymph nodes, peritoneum, or distant organs) and 30 peritoneal macroscopic metastases (PMM). Thirteen miRNA genes (MIR107, MIR124-2, MIR124-3, MIR125B-1, MIR127, MIR129-2, MIR130B, MIR132, MIR193A, MIR339, MIR34B/C, MIR9-1, and MIR9-3) were hypermethylated already at the early stages of OvCa, while hypermethylation of MIR1258, MIR137, MIR203A, and MIR375 was pronounced in metastatic tumors, and MIR148A showed high methylation levels specifically in PMM. We confirmed the significant relationship between methylation and expression levels for 11 out of 12 miRNAs analyzed by qRT-PCR. Moreover, expression levels of six miRNAs were significantly decreased in metastatic tumors in comparison with nonmetastatic ones, and downregulation of miR-203a-3p was the most significant. We revealed an inverse relationship between expression levels of miR-203a-3p and those of ZEB1 and ZEB2 genes, which are EMT drivers. We also identified three miRNA genes (MIR148A, MIR9-1, and MIR193A) that likely regulate EMT–MET reversion in the colonization of PMM. According to the Kaplan–Meier analysis, hypermethylation of several examined miRNA genes was associated with poorer overall survival of OvCa patients, and high methylation levels of MIR130B and MIR9-1 were related to the greatest relative risk of death.

Currently, the search for new promising tools of immunotherapy continues. In this regard, microRNAs (miRNAs) that influence immune checkpoint (IC) gene expression in tumor and T-cells and may be important regulators of immune cells are considered. MiRNAs regulate gene expression by blocking mRNA translation. An important feature of miRNA is its ability to affect the expression of several genes simultaneously, which corresponds to the trend toward the use of combination therapy. The article provides a list of miRNAs acting simultaneously on several ICs and miRNAs that, in addition to IC, can regulate the expression of targeted therapy genes. There is dependence of miRNA interactions with IC genes on the type of cancer. The analysis of the accumulated data demonstrates that only about 14% (95% CI: 9.8–20.1%) of the studied miRNAs regulate the expression of specific IC in more than one type of cancer. That is, there is tumor specificity in the miRNA action on ICs. A number of miRNAs demonstrated high efficiency in vitro and in vivo. This indicates the potential of miRNAs as promising agents for cancer immunotherapy. Additional studies of the miRNA–gene interaction features and the search for an optimal miRNA mimic structure are necessary.

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.

Ранее считалось, что РНК в основном передаёт информацию, закодированную в ДНК, чтобы другие молекулы, такие как рибосомы, могли использовать код для создания белков. Однако за последние 30 лет исследователи обнаружили, что существуют разные типы РНК, среди которых одним из наиболее важных является некодирующая РНК (нкРНК), характеризующаяся отсутствием открытой рамки считывания и не участвующая в производстве белков. Открытие большого количества нкРНК произвело революцию в этой области, изменив представление исследователей о физиологии и развитии. Некодирующие РНК, иногда называемые «тёмной материей» генома, составляют более 90% от всех РНК, кодируемых геномом человека. Но большая часть из известных нкРНК (~50000) обнаружена только за последние 10 лет и остается пока мало изученной. Тем не менее, с тех пор показано, что многие нкРНК играют важную роль как в нормальной физиологии клетки, так и при заболеваниях, включая онкологические. Так, некоторые малые нкРНК настолько стабильны, что могут длительное время существовать в кровотоке и могут стать основой для точного и чувствительного скрининга основных видов рака человека в нескольких каплях крови или плазмы. Кроме того, нкРНК могут стать лекарством, и их доставка может осуществляться на основе тех же принципов, что и доставка синтетической РНК и олигонуклеотидов, нацеленных на мРНК, кодирующие белок. В этом обзоре мы рассматриваем роль некоторых нкРНК - микроРНК (миРНК) и длинных нкРНК (днРНК) в патогенезе рака молочной железы (РМЖ). Большая часть этого обзора, будет посвящена нкРНК, как диагностическим биомаркёрам. Мы обсудим также их потенциальное использование в персонализированном лечении РМЖ. Previously it was believed that RNA mainly conveys the information encoded in DNA, so that other molecules, such as ribosomes, could use that code to create proteins. However, over the past 30 years, different types of RNA have been discovered. One of the most important of them is non-coding RNA (ncRNA) characterized by the absence of the open reading frame and not involved in the protein production. The discovery of numerous ncRNAs has revolutionized this field and changed the researchers’ ideas about physiology and development. Non-coding RNAs, sometimes called “dark matter” of the genome, constitute more than 90% of all RNAs encoded by the human genome. However, most of the known ncRNAs (~50,000) were discovered only during the last decade and remain understudied. Nevertheless since that time, it has been shown that many ncRNAs play an important role both in normal cell physiology and in diseases, including cancer. Thus, some small ncRNAs are so stable that they can exist in the bloodstream for a long time and can become a basis for an accurate and sensitive screening for major human cancers using only a few drops of blood or plasma. In addition, ncRNAs may become medications that could be delivered based on the same principle as the delivery of synthetic RNA and oligonucleotides targeting protein-coding mRNAs. In this review, we addressed the role of some ncRNAs, miRNAs (siRNAs) and long ncRNAs (lncRNAs), in the pathogenesis of breast cancer. Much of this review will focus on ncRNAs as diagnostic biomarkers. We will also discuss their potential use in the personalized treatment of breast cancer.

Background. The introduction of immunotherapy based on immune checkpoint inhibitors has significantly improve the effectiveness of kidney cancer treatment. Nevertheless, not all patients respond to such treatment and there are no reliable predictive markers. Therefore, the development of a model system for assessing the cellular immune response to a tumor seems to be an urgent task.Aim. Development of a model to assess the T-cell immune response was the focus of this study.Materials and methods. Primary tumor cell culture and peripheral blood T-cell fraction were obtained under standard sterile conditions. T-cell activation were perform via anti-CD3 and anti-CD28 antibodies. The cell index was assessed using the RTCA xCELLigence biosensor technology (ACEA Biosciences, USA).Results. Tumor and T-cells from the same patient were cultured together to assess the growth rate of the tumor cell population. Measurements were taken at 30-minute intervals. The duration of observation was 24 hours. It has been shown that non-activated T-cells do not affect the proliferative properties of cultured cancer cells. On the contrary, activated T-cells suppressed the proliferative properties of cancer cells, which was associated with an increase in the proportion of Т-cells carrying the HLA-DR receptor (CD3+HLA–DR+) because of activation. Tumor-specific T-cell activity can lead to three consequences: lack of effect, partial suppression of proliferative properties, and complete death of tumor cells. In the latter case, the absence of such cells was determined by flow cytometry.Conclusion. The developed approach makes it possible to evaluate the cytotoxic properties of T cells in relation to tumor cells in a particular patient. The advantage of this method is that the measurement can be carried out in the presence of immune checkpoint inhibitors. The proposed method may be useful for evaluating the treatment regimen within the framework of personalized therapy.

Таргетные препараты для лечения онкологических заболеваний, в частности, мультикиназные ингибиторы сунитиниб и сорафениб, широко применяются в онкологической практике, и изучение особенностей их применения весьма актуально. На модели культивируемых раковых клеток in vitro показано, что ингибиторы киназ способны усиливать экспрессию генов опухолевой прогрессии (SNAI1, SNAI2, CD44, CDH1, CLDN1), связанных с эпителиально-мезенхимальным переходом (ЭМП). Показана активация таких генов при действии низких доз тирозинкиназных ингибиторов - сунитиниба и сорафениба. Активация изученных генов была различной в разных культурах и зависела от примененного препарата. Предположено, что в силу локального неравномерного распределения лекарственных средств в тканях опухоли, ингибиторы киназ могут являться одной из причин активации экспрессии генов, ассоциированных с ЭМП, и, таким образом, способствовать ее прогрессии. Targeted drugs for the treatment of oncological diseases, in particular, the inhibitors of Sunitinib and Sorafenib, are widely used in oncological practice and the study of the peculiarities of their application is very relevant. On the models of cultured cancer cells in vitro it was shown that kinase inhibitors are able to strengthen the expression of tumor progression genes (SNAI1, SNAI2, CD44, CDH1, CLDN1) associated with the epithelial-mesenchymal transition (EMT). The activation of such genes is shown under the action of low doses of tyrosine kinase inhibitors - Sunitinib and Sorafenib. The activation of the studied genes was different in different cultures and depended on the applied drug. It was assumed that due to the local uneven distribution of drugs in tissues of the tumor, kinase inhibitors may be one of the reasons for activating the expression of genes associated with the epithelial- mesenchymal transition, and thus contribute to its progression.

To increase the effectiveness of anticancer therapy based on immune checkpoint (IC) inhibition, some ICs are being investigated in addition to those used in clinic. We reviewed data on the relationship between PD-L1, B7-H3, B7-H4, IDO1, Galectin-3 and -9, CEACAM1, CD155, Siglec-15 and ADAM17 expression with cancer development in complex with the results of clinical trials on their inhibition. Increased expression of the most studied ICs-PD-L1, B7-H3, and B7-H4-is associated with poor survival; their inhibition is clinically significant. Expression of IDO1, CD155, and ADAM17 is also associated with poor survival, including gastric cancer (GC). The available data indicate that CD155 and ADAM17 are promising targets for immune therapy. However, the clinical trials of anti-IDO1 antibodies have been unsatisfactory. Expression of Galectin-3 and -9, CEACAM1 and Siglec-15 demonstrates a contradictory relationship with patient survival. The lack of satisfactory results of these IC inhibitor clinical trials additionally indicates the complex nature of their functioning. In conclusion, in many cases it is important to analyze the expression of other participants of the immune response besides target IC. The PD-L1, B7-H3, B7-H4, IDO1 and ADAM17 may be considered as candidates for prognosis markers for GC patient survival.

Background. Metastasing and degree of differentiation refer to the main clinical characteristics of malignant tumors. Both listed features need an in-depth study that can lead to an understanding of the mechanisms for the occurrence of certain state of cancer cells.Objective. Studying the processes of metastasis and differentiation of the clear cell renal cell carcinoma (ccRCC) on gene expression.Materials and methods. The levels of expression of ten genes in 65 paired samples were studied (ccRCC tumor tissue and the normal kidney tissue) by the real-time polymerase chain reaction.Results. It is shown that the expression of CA9, NDUFA4L2, VWF, IGFBP3, BHLHE41, ANGPTL4 and EGLN3 genes is associated both with the degree of differentiation of the ccRCC and with the metastasis of this tumor. C1QA expression is connected only with metastasis, but does not participate in the process of differentiation of tumor cells. An ambiguous situation with FN1 and CSF1R gene expression is not essential for ccRCC metastasis processes, but may have a certain value for differentiation of cells of this tumor. Low-differentiated tumors have about five times an increased metastasis frequency during the year relative to highly differentiated tumors (odds ratio 4.94). A low correlation of gene expression in tumors with a low degree of differentiation is revealed, as opposed to their high co-expression during tumor progression by TNM classifications.Conclusion. A significant part of genes substantial for the development of ccRCC is associated with both metastasis and the degree of differentiation of the ccRCC, which is due to the similarity of functional changes that stimulate both of these processes. For low-differentiated tumors the number of genes with correlated expression is less than in high-differentiated tumors. This may be due to disorganization of gene expression.

Background. Renal cell carcinoma (RCC) is asymptomatic up to severe stages and is characterized by a high mortality rate, reaching 90 % with the development of a metastatic process.Objective: to determine the group of microRNA genes, the methylation of which is associated with the progression of the disease, in particular, with metastasis.Materials and methods. Methylation-specific polymerase chain reaction in a representative sample of RCC patients (98 cases) showed an increase in the methylation status of 6 microRNA genes (MIR9-1, MIR9-3, MIR34b/c, MIR130b, MIR1258, MIR107) in tumor DNA samples relative to matched samples of histologically unchanged tissue.Results. For 4 genes (MIR9-1, MIR107, MIR130b, MIR1258), a significant association of methylation with late (III-IV) stages, tumor size, loss of differentiation, and metastasis to lymph nodes or distant organs was shown. These 4 genes were used to compose a potential metastatic prognosis marker system with a clinical sensitivity of 68 % and a specificity of 84 % (area under curve 0.83), which will be applied in the final development of a system for personalized therapy of RCC patients.Conclusion. The association of methylation of the MIR1258 with RCC metastasis has been shown for the first time and is of independent interest as a new promising marker for the prognosis of metastatic relapses.

Neuroendocrine neoplasms (NEN) are infrequent malignant tumors of a neuroendocrine nature that arise in various organs. They occur most frequently in the lungs, intestines, stomach and pancreas. Molecular diagnostics and prognosis of NEN development are highly relevant. The role of clinical biomarkers can be played by microRNAs (miRNAs). This work is devoted to the analysis of data on miRNA expression in NENs. For the first time, a search for specificity or a community of their functional characteristics in different types of NEN was carried out. Their properties as biomarkers were also analyzed. To date, more than 100 miRNAs have been characterized as differentially expressed and significant for the development of NEN tumors. Only about 10% of the studied miRNAs are expressed in several types of NEN; differential expression of the remaining 90% was found only in tumors of specific localizations. A significant number of miRNAs have been identified as potential biomarkers. However, only a few miRNAs have values that characterized their quality as markers. The analysis demonstrates the predominant specific expression of miRNA in each studied type of NEN. This indicates that miRNA’s functional features are predominantly influenced by the tissue in which they are formed.

The fundamental novelty in the pathogenesis of renal cell carcinoma (RCC) was discovered as a result of the recent identification of the role of long non-coding RNAs (lncRNAs). Here, we discuss several mechanisms for the dysregulation of the expression of protein-coding genes initiated by lncRNAs in the most common and aggressive type of kidney cancer-clear cell RCC (ccRCC). A model of competitive endogenous RNA (ceRNA) is considered, in which lncRNA acts on genes through the lncRNA/miRNA/mRNA axis. For the most studied oncogenic lncRNAs, such as HOTAIR, MALAT1, and TUG1, several regulatory axes were identified in ccRCC, demonstrating a number of sites for various miRNAs. Interestingly, the LINC00973/miR-7109/Siglec-15 axis represents a novel agent that can suppress the immune response in patients with ccRCC, serving as a valuable target in addition to the PD1/PD-L1 pathway. Other mechanisms of action of lncRNAs in ccRCC, involving direct binding with proteins, mRNAs, and genes/DNA, are also considered. Our review briefly highlights methods by which various mechanisms of action of lncRNAs were verified. We pay special attention to protein targets and signaling pathways with which lncRNAs are associated in ccRCC. Thus, these new data on the different mechanisms of lncRNA functioning provide a novel basis for understanding the pathogenesis of ccRCC and the identification of new prognostic markers and targets for therapy.

We aimed to identify and investigate genes that are essential for the development of clear cell renal cell carcinoma (ccRCC) and sought to shed light on the mechanisms of its progression and create prognostic markers for the disease. We used real-time PCR to study the expression of 20 genes that were preliminarily selected based on their differential expression in ccRCC, in 68 paired tumor/normal samples. Upon ccRCC progression, seven genes that showed an initial increase in expression showed decreased expression. The genes whose expression levels did not significantly change during progression were associated mainly with metabolic and inflammatory processes. The first group included CA9, NDUFA4L2, EGLN3, BHLHE41, VWF, IGFBP3, and ANGPTL4, whose expression levels were coordinately decreased during tumor progression. This expression coordination and gene function is related to the needs of tumor development at different stages. Specifically, the high correlation coefficient of EGLN3 and NDUFA4L2 expression may indicate the importance of the coordinated regulation of glycolysis and mitochondrial metabolism. A panel of CA9, EGLN3, BHLHE41, and VWF enabled the prediction of survival for more than 3.5 years in patients with ccRCC, with a probability close to 90%. Therefore, a coordinated change in the expression of a gene group during ccRCC progression was detected, and a new panel of markers for individual survival prognosis was identified.

Systemic analysis of the relationship between the levels of methylation of 21 microRNA genes and the parameters of breast cancer progression was performed on a representative sample of 91 paired specimens of breast cancer and histologically normal tissues and a system of markers for prediction of metastasis was proposed. A significant association of hypermethylation of 11 genes with late (III-IV) clinical stages was found, and for 6 genes (MIR124-1, MIR127, MIR34B/C, MIR9-3, MIR1258, and MIR339) this association was highly significant (p≤0.001, FDR=0.01). For MIR9-3 and MIR339, an association with tumor size was demonstrated (p<0.001, FDR=0.01). No association of the levels of methylation of the analyzed microRNA genes with the degree of differentiation were found. An association with lymph node metastasis was established for 9 microRNA genes; the most significant association was shown for 6 genes MIR125B-1, MIR127, MIR9-3, MIR339, MIR124-3, and MIR1258 (p<0.005, FDR=0.05). Based on these 6 genes, a marker system for predicting breast cancer metastasis was developed by ROC analysis. This system is characterized by 87% sensitivity and 77% specificity (AUC=0.894). The proposed system may have clinical application in the personalized treatment of breast cancer patients.