[Transcription Factor SAP30 Is Involved in the Activation of NETO2 Gene Expression in Clear Cell Renal Cell Carcinoma].

Clear cell renal cell carcinoma (ccRCC) is a common oncourological disease with a high mortality level. The incidence of this type of cancer is constantly increasing, while molecular mechanisms involved in the disease initiation and progression remain far from being fully understood. A problem of the search for novel markers is crucial for improvement of diagnosis and therapy of ccRCC. We have previously found that the disease is characterized by increased expression of the NETO2 gene. In the present study, we showed that isoform 1 (NM_018092.4) makes the main contribution to the upregulation of this gene. Using original CrossHub software, "The Cancer Genome Atlas" (TCGA) project data were analyzed to identify possible mechanisms of NETO2 gene activation in ccRCC. The absence of significant contribution of methylation to the increase of mRNA level of the gene was observed. At the same time, a number of genes encoding transcription factors, which could potentially regulate the expression of NETO2 in ccRCC, were identified. Three such genes (MYCBP, JMY, and SAP30) were selected for the further analysis of their mRNA levels in a set of ccRCC samples with quantitative PCR. We showed a significant increase in mRNA level of one of the examined genes, SAP30, and revealed its positive correlation with NETO2 gene expression. Thus, upregulation of NETO2 gene is first stipulated by the isoform 1 (NM_018092.4), and the probable mechanism of its activation is associated with the increased expression of SAP30 transcription factor.

[Suppression of NR0B2 gene in Clear Cell Renal Cell Carcinoma Is Associated with Hypermethylation of Its Promoter].

Clear cell renal cell carcinoma (ccRCC) is a common urologic malignancy. Understanding of the transcriptional regulation of oncogenes and tumor suppressor genes involved is critical for the development of the treatments for renal tumors. Using ccRCC subdivision of the TCGA dataset, we identified NR0B2 encoding orphan nuclear receptor as a tumor suppressor candidate in renal tissue. In independent cohort of primary renal tumors, quantitative PCR experiments confirmed significant suppression of NR0B2 mRNA in 86% of ccRCC samples studied. In 80% of these cases, we detected the hypermethylation of the NR0B2 pro-moter region. These results suggest that NR0B2 is a tumor suppressor gene in ccRCC, and that the hypermethylation of promoter region is the main mechanism of its downregulation.

[Overexpression of microRNAs miR-9, -98, and -199 Correlates with the Downregulation of HK2 Expression in Colorectal Cancer].

Glycolysis activation is one of the main features of energy metabolism in cancer cells that is associated with the increase in glycolytic enzyme synthesis, primarily, hexokinases (HKs), in many types of tumors. Conversely, in colorectal cancer (CRC) the decrease in the expression of HK2 gene, which encodes one of the key rate-limiting enzyme of glycolysis, was revealed, thus, the study of the mechanisms of its inhibition in CRC is of particular interest. To search for potential microRNAs, inhibiting the expression of HK2 in CRC, we have performed the analysis of data from "The Cancer Genome Atlas" (TCGA) and five microRNA-mRNA target interaction databases (TargetScan, DIANA microT, mirSVR (miRanda), PicTar, and miRTarBase) using original CrossHub software. Seven microRNAs containing binding site on mRNA HK2, which expression is negatively correlated with HK2 expression, were selected for further analysis. The expression levels of these microRNAs and mRNA HK2 were estimated by quantitative PCR on a set of CRC samples. It has been shown, that the expression of three microRNAs (miR-9-5p, -98-5p, and -199-5p) was increased and correlated negatively with mRNA level of HK2 gene. Thus, downregulation of HK2 gene may be caused by its negative regulation through microRNAs miR-9-5p, -98-5p, and -199-5p.

Plasma Level of hsa-miR-619-5p microRNA Is Associated with Prostatic Cancer Dissemination beyond the Capsule.

Profiles of circulating microRNA in the plasma of patients with prostate cancer with pathomorphological stages pT2, pT3, and pT4 are analyzed. The level of circulating microRNA hsa-miR-619-5p is elevated in patients with extracapsular spreading of the tumor, increasing significantly from stage pT2 to stage pT4.

Changes in the Level of Circulating hsa-miR-297 and hsa-miR-19b-3p miRNA Are Associated with Generalization of Prostate Cancer.

We performed diagnostic classification of plasma specimens from patients with non-metastatic and metastatic prostate cancer based on pairs of miRNA that have no individual diagnostic significance. Of 230 miRNA detected in plasma specimens, 3 pairs were diagnostically significant. The miRNA pair hsa-miR-19b-3p and hsa-miR-297 demonstrated highest sensitivity and specificity. Among common target genes of these miRNA, CFL2 gene associated with cell mobility was detected.

MicroRNA hsa-miR-4674 in Hemolysis-Free Blood Plasma Is Associated with Distant Metastases of Prostatic Cancer.

We analyzed microRNA profile in hemolysis-free blood plasma of patients with prostatic cancer. The metastatic form of prostatic cancer was found to be associated with increased levels of hsa-miR-22-3p, hsa-miR-663a, and hsa-miR-4674 in comparison with non-metastatic form. Common candidate target genes of these microRNA include JUNB, KMT2A, and XPO6.

Plasma Levels of hsa-miR-619-5p and hsa-miR-1184 Differ in Prostatic Benign Hyperplasia and Cancer.

Peripheral blood plasma profiles of circulating microRNA expression were analyzed in patients with prostatic cancer and benign hyperplasia. In prostatic cancer, significant increase in hsa-miR-619-5p and hsa-miR-1184 microRNA expression and significant decrease in hsalet-7b-5p and hsa-let-7c-5p microRNA expression were observed. The role of the relationship between the microRNA expression and the activities and functions of host genes with introns encoding these microRNA is discussed.

Analysis of Plasma microRNA Associated with Hemolysis.

We analyzed the effect of hemolysis on microRNA profi le of blood plasma. It was found that hemolysis of ~0.05% erythrocytes in a sample signifi cantly affected the concentration of 9 microRNA: hsa-miR-486-5p, hsa-miR-16-5p, hsa-miR-451a, hsa-miR-106a-5p, hsa-miR-17-5p, hsa-miR-93-5p, hsa-miR-20a-5p, hsa-miR-107, and hsa-miR-20b-5p. The effect of hemolysis on plasma content of miR-17 family microRNA was demonstrated.

Profile of microRNA in Blood Plasma of Healthy Humans.

Analysis of the plasma microRNA profile can be used for the diagnosis of various pathological and physiological conditions. Complete microRNA microprofiling is an extremely important task. Here we used microarray analysis allowing measurement of the expression of 2500 microRNA (MirBase, version 20). About 10% known microRNA were found in the plasma. Most of the detected microRNA (69 microRNA; ~30%) were encoded by mirtrons.

[Molecular genetic mechanisms of drug resistance in prostate cancer].

The major problem in prostate cancer treatment is the development of drug resistance and especially important, cross-resistance. The mechanisms of drug resistance, which are divided into ligand-dependent (requiring the presence of androgens in the cell) and independent (not requiring the presence of androgens) are reviewed. The mechanisms are mainly represented with mutations of the androgen receptor and expression of aberrant constitutively active splice variants, as well as up-regulation of genes involved in androgens synthesis.

Suppression of ITGB4 Gene Expression in PC-3 Cells with Short Interfering RNA Induces Changes in the Expression of ß-Integrins Associated with RGD-Receptors.

We studied the effect of transfection of PC-3 prostate cancer cells with a plasmid encoding shRNA complimentary to a fragment of integrin ß4 (ITGB4). The results attest to considerable changes in the transcriptome of transfected cells. For instance, compensatory changes in the expression of integrin family genes were found.

[Evaluation of Gene Expression of Hexokinases in Colorectal Cancer with the Use of Bioinformatics Methods].

One of the hallmarks of cancer is the change of energy metabolism, mainly activation of glycolysis that occurs even at early stages of tumorigenesis. The glycolysis activation can be caused by overexpression of hexokinases, primarily HK1 and HK2. Colorectal cancer, which takes the third place in the cancer morbidity and mortality rates worldwide, is believed to be accompanied with overexpression of HK2, which is .considered a marker of poor prognosis. With the use of the developed CrossHub tool, we performed the analysis of the Cancer Genome Atlas RNA-Sequencing data, which, on the contrary, revealed the prevalence of the down-regulation of HK2 gene and only slight expression alterations in HK1 gene. The Cancer Genome Atlas is the largest resource in the field of molecular oncology that accumulated genomic, transcriptomic and methylomic data for thousands of sample of more than 20 cancers. The transcriptome analysis data for colorectal cancer (283 tumor samples and 41 matched normal samples) were in accord with the results of further qPCR expression level evaluation. Up-regulation of HK1 and HK2 genes was observed only in a part of samples: 12% for HK1 and 30% for HK2. At the same time, the HK2 mRNA level decrease was shown in 50% of cases. Correlation analysis revealed the consistency in HK1 and HK2 expression alterations (Spearman's rank correlation coefficient r(s) = 0.43, p < 0.01), that could be explained by common deregulation mechanisms of these genes in colorectal tumors. The HK3 expression level was significantly increased in 60% of samples. Most likely, just hexokinase 3 contributes significantly to the activation of glycolysis in colorectal cancer.