[Vasculogenic mimicry]. / Vaskulogennaya mimikriya.

Anti-angiogenic drugs are used as an established approach of malignant neoplasms therapy. It has been established that the development of the phenomenon of vasculogenic mimicry - a specific variant of tumor neoangiogenesis, which is formed in highly aggressive solid tumors, is associated with a decrease in the effectiveness of antitumor therapy. This review highlights the mechanisms of development of vasculogenic mimicry in malignant neoplasms, which is one of the alternative options for tumor blood supply. In the formation of vasculogenic mimicry, an important role is assigned to the tumor microenvironment, primarily tumor-associated macrophages and fibroblasts. The signaling pathways that regulate the formation of vasculogenic mimicry channels in tumors have been characterized. The prospects for a targeted impact on molecular targets that initiate and promote vasculogenic mimicry, the impact on which can increase the effectiveness of antitumor therapy, are shown. The review discusses experimental studies of the mechanisms of vasculogenic mimicry formation in malignant neoplasms and the prospects for targeted action on molecules that are components of signaling cascades involved in the development of this model of neoangiogenesis.

[Gene expression profiling in melanoma diagnostics: problems and future application in clinical practice]. / Profilirovanie ekspressii genov v diagnostike melanomy: problemy i perspektivy.

Difficulties in the diagnosis and differential diagnosis of melanoma in the work of a pathologist include not only conflicting structural and morphological features, but also the insufficient effectiveness of biochemical and some molecular markers in immunohistochemical studies. The review presents modern alternative methods for diagnosing malignant tumors based on the assessment of gene expression, the performance, objectivity and reliability of the determination of which may in the future have clinical application as an addition to histopathological methods in the diagnosis and differential diagnosis of various malignant neoplasms, including melanocytic neoplasms, which is changing the paradigm of routine medical practice, introducing diagnostic tests that carry molecular information into it.

[

MicroRNAs epigenetically regulate physiological and pathological processes. Previously, we found that miR-204-5p is expressed at low levels in melanoma cells, and an increase in its level leads to a change in proliferation, migration, and invasion of these cancer cells. Now, using bioinformatics analysis, it has been shown that the target of miR-204-5p is FOXC1 transcription factor, which is implicated in carcinogenesis. Using the luciferase reporter assay, it was found that miR-204-5p suppresses expression of the FOXC1 gene by binding to its 3' non-coding region. Transfection of small interfering RNA (siRNA) targeting FOXC1 into melanoma cells caused a decrease in miR-204-5p levels, which is consistent with the generally accepted concept of feedback regulation of miRNA expression by target genes. According to the results of the MTT test and fluorescence microscopy, the proliferation level of melanoma cells under the influence of siRNA to FOXC1 decreased 72 h after transfection. Changes in the ratio of cells by cell cycle phase were analyzed using flow cytometry. Regulatory relationships between FOXC1 and miR-204-5p, and an inhibitory effect of FOXC1 knockdown on melanoma cell proliferation were revealed. Based on the results, it can be assumed that miR-204-5p regulates proliferation of melanoma cells by affecting FOXC1 expression.

[The significance of C-KIT gene mutations in the diagnosis and prognosis of malignant tumors]. / Znachenie mutatsii gena C-KIT v diagnostike i prognoze techeniya zlokachestvennykh opukholei.

Mutations in the C-KIT gene encoding type III receptor tyrosine kinase that regulates cellular processes, such as differentiation, survival, proliferation, migration, and apoptosis, are found in some neoplasms: gastrointestinal stromal tumor, mastocytosis, melanoma, breast carcinomas, myeloid leukemias, and a number of others. Tumors that exhibit these mutations are sensitive to therapy with tyrosine kinase inhibitors, which makes it necessary to correctly identify the mutation status by C-KIT in order to apply a personalized approach to therapy. This literature review shows that the type and localization of the C-KIT gene mutation are of crucial prognostic value and significance in choosing drugs for antitumor therapy, but traditional diagnostic methods fail to determine accurate mutation characteristics. Routine sequencing techniques focus on identifying the gene mutations associated with specific cellular processes, such as DNA damage and repair. The emergence of next-generation sequencing techniques has solved this problem, making it possible to fully analyze the genome of a malignant neoplasm, with constant screening for new mutations that appear as the tumor develops, affect the prognosis of the disease, and change its sensitivity to the antitumor therapy.

PROONCOGENIC EFFECTS OF INHIBITING THE microRNA miR-106A IN SKIN MELANOMA CELLS IN VITRO.

One of the regulators of gene expression at the post-transcriptional level are miRNAs. Due to its multifunctionality, these molecules are considered as potential targets for controlling the biological behavior of tumor cells. To date, several thousand types of microRNAs have been identified and their expression profiles have shown significant during malignant transformation of cells. In this study, we have investigated the effect of miR-106a functional inhibition on the growth, viability and apoptosis of melanoma cells. Comparative analysis of expression profiles in melanona cells and in cells of melanocytic nevi identified by the use of microarray has revealed a significant increase in the miRNA expression level in melanoma cells. Despite this, inhibition of this molecule in melanoma cells has no antitumor effect on cell proliferation, viability, migration activity and apoptosis of melanoma cells, but increases invasive activity and the ability to form colonies. The paper discusses the importance of evaluating changes in miRNA levels in melanoma and other malignancies, relationship îf miR-106a with the pathogenesis of melanoma, as well as the possible role of miR-106a in other pathologies.

MICRORNAS AS ULTRAVIOLET IRRADIATION EFFECTS REGULATORS IN SKIN CELLS.

MicroRNAs belong to small non-coding RNA which regulate gene expression via mRNA degradation or translation inhibition. MicroRNAs are active modulators of gene expression in the skin caused by exogenous factors including ultraviolet irradiation. These effects are realized by targeting transcription factors and signaling systems components. Changes in microRNAs levels started to register in a few hours after exposure to ultraviolet irradiation, wich confirms the presence of an effective fast processes in the scin cells that modulate the functional status of microRNAs. The reported recently ability of microRNAs to be transported by exosomes may be related to systemic effects of ultraviolet irradiation that include the altered immune response and systemic inflammatory reaction. Understanding these processes is important because of the possibility of purposeful influence on the expression and activity of a microRNA that may have implications for diagnosis and therapy of photodermatosis and malignant skin tumors.

[ Bioluminescent assay to detect melanocortin-1 receptor (MC1R) polymorphisms (R160W, R151C, and D294H)].

Several polymorphisms in melanocortin-1 receptor (MC1R) gene are shown to have associations with melanoma risk. In particular, rs1805007, rs1805008, and rs1805009 mutations causing the corresponding R151C, R160W, and D294H changes and associated with the phenotype ("red-hair mutations") are connected with melanoma and non-melanoma skin cancer risks. The work describes the approach to detect these polymorphisms based on primer extension reaction with the following dual bioluminescent assay. Model plasmids with polymorphic MC1R fragments as well as several clinical DNA samples were tested using the developed technique. The results were in good correlation with those obtained by Sanger sequencing.

[Molecular and pathomorphological prognostic markers for melanoma: Current approaches and prospects].

The incidence of melanoma demonstrates a persistent increasing tendency, which justifies the need to study and identify new prognostic markers for the development and course of this disease. The given paper shows current approaches to melanoma staging, including those to applying pathomorphological prognostic criteria, and discusses prospects for using the results of genomic and epigenomic studies of the carcinoma in clinical practice.

Russian study of morphological prognostic factors characterization in BRAF-mutant cutaneous melanoma.

Melanoma is one of the aggressive cancer types causing the majority of deaths in skin cancer patients. Mutational screening of the tumor revealed a number of driver mutations in oncogenes which enabled melanoma classification into a few molecular subtypes. BRAF is a key component of mitogen-activated kinase pathway; its activating mutation leads to accelerated melanoma cells proliferation, invasion and survival. Somatic mutations in BRAF were reported in various malignancies, including thyroid cancer, colorectal cancer and melanoma. Specific features of BRAF-positive tumors could have clinical implications as mutational alterations may have an impact on the biological behavior of the tumor and prognosis of the disease. In the present study, the frequency of BRAF V600E mutation was evaluated in Russian patients with melanocytic lesions, of which 41.25% were primary melanoma and 60% were melanocytic nevi. Melanoma patients with trunk localization were of younger age in the BRAF-positive group as compared with BRAF-negative patients. Immunohistochemical evaluations of Ki-67 expression, as well as matrix metalloproteinase-2, -9, were found to be equal in BRAF-positive and BRAF-negative tumors. MMP-2/MMP-9 immunoreactivity was observed in stromal and/or melanocytic cells both in melanoma and nevi patients. Besides tumor cells, MMP-9 expression was observed in lymphocytes in 27.2% of BRAF-positive and in 19.1% of BRAF-negative patients. Histopathological prognostic markers (Breslow thickness, mitotic index, ulceration, tumor infiltrating lymphocytes pattern) did not show any differences depending on BRAF V600E mutational status. The frequency of BRAF-positive melanomas in Russian cohort is similar to other Caucasian population rates. BRAF V600E mutation harboring tumors are more often observed in younger patients without specific features of morphological prognostic factors.

[Inhibition of matrix metalloproteinases 9 and 13 affects the degree of lymphocytic infiltration and the expression levels of microRNA miR-21 and miR-let-7b in melanoma cells in vivo].

OBJECTIVE: To estimate changes in the trend of growth of primary tumor nodules, the degree of lymphocytic infiltration, and the expression levels of oncomicroRNA miR-21 and miR-let-7b when inhibiting matrix metalloproteinases 9 and 13 (MMP-9 and MMP-13) in vivo in C57B16 mice with transplantable melanoma B-16. MATERIAL AND METHODS: Tumor growth was evaluated measuring the volume of primary tumor nodules; the degree of lymphocytic infiltration was microscopically estimated using hematoxylin-eosin-stained tissue specimens, by calculating intratumoral lymphocytes. The expression of oncomicroRNA was quantified by real-time PCR. RESULTS: It was shown that MMP-9 and MMP-13 inhibition had no impact on the growth of primary tumor nodules; selective MMP-9 inhibition failed to affect the degree of lymphocytic infiltration of a primary tumor nodule and to change the expression of oncomicroRNA miR-21 and miR-let-7b; the concomitant inhibition of MMP-9 and MMP-13 altered the immunogenic properties of melanoma, stimulated the lymphocytic infiltration of tumor nodules, and decreased the expression of oncomicroRNA miR-21 and miR-let-7b; the degree of lymphocytic infiltration of primary tumor nodules increased in the dynamics of a tumor process and the expression levels of oncomicroRNA remained unchanged. CONCLUSION: The concomitant inhibition of MMP-9 and MMP-13 affects prognosis and survival in skin melanoma.

[BRAF V600E mutation detection in melanoma patients: clinical and morphological features].

OBJECTIVE: To estimate the frequency of BRAF V600E mutation in melanoma patients and to assess an association of this type of mutation with the clinical and morphological characteristics of the disease. MATERIAL AND METHODS: The mutation of BRAF V600E was analyzed using an allele-specific real-time PCR assay in 71 melanoma patients treated at the Krasnoyarsk Territorial Oncology Dispensary. Before DNA isolation, the glasses containing the material obtained from the patients and stained with hematoxylin and eosin were tested for the content of tumor cells in the sections. Later on, genomic DNA was isolated with a FFPET DNA-Extraction kit (Biolink, Russia) and the real-time PCR assay was carried out. RESULTS: The BRAF V600E mutation was observed in 40.84% of the patients and was not in 59.16%. There were specific features in tumor location and histotype in relation to the status of BRAF. CONCLUSION: To detect whether the V600E mutation is present or absent in the BRAF gene in melanoma patients is an important component in the evaluation of the biological behavior of tumor cells and their performance of biologically significant functions, such as tumor invasion and dissemination and neoangiogenesis, which is important for the further assessment of tumor progression and disease prognosis.

[Modulation of apoptosis in mononuclear cells with different variants of polymorphism C-262T in catalase gene].

Catalase is a clue enzyme metabolizing hydrogen peroxide under conditions of oxidative stress. It is known that the C-262T polymorphism in catalase gene is implicated in higher risk of some diseases onset. In this article we have evaluated the viability of mononuclear leukocytes isolated from patients with different variants of C-262T polymorphism in catalase gene (CC, CT, TT) under conditions of hydrogen peroxide induced oxidative stress. It has been revealed that cells with TT variant of polymorphism have lower viability in the presence of hydrogen peroxide in a concentration of 1000 µM/L in comparison with cells having CC and CT genotypes. Furthermore, unlike CC and CT variants, cells with TT polymorphism showed no activation of mitochondrial apoptotic pathway even with hydrogen peroxide at a concentration of 1 mM. The data obtained may explain the increased risk of diseases related with the activity of the antioxidant system in patients with 262 TT promoter polymorphism in catalase gene.

[Characteristics of CD99 expression in the skin of psoriasis patients].

The pattern of CD99 expression in the skin was studied in 22 patients with disseminated psoriasis. The patients' skin biopsy specimens were immunohistochemically stained by the standard procedure using monoclonal antibodies to CD99. The progression of psoriasis was characterized by the increased expression of the protein in the dermis (lymphocytes and vascular endothelium) and epidermis (keratinocytes) with the antigen being located not only in the cytoplasm, but also in the membrane. In remission, the content of CD99-positive cells in the dermis and epidermis was significantly reduced with the prevalence of cytoplasmic localization in the epidermal keratinocytes. It is assumed that the expression of CD99 plays an important role in the pathogenesis of psoriasis, in the processes of emigration of leukocytes, and in their tropism toward to the epidermis.

Analysis of the application of MMP-9 inhibitor in skin melanoma: experimental study.

Experiment on C57Bl/6 mice with modeled skin melanoma showed that selective inhibition of matrix metalloproteinase-9 increased lifetime and reduced the number of PCNA(+) tumor cells and intensity of neoangiogenesis. Inhibition of matrix metalloproteinase-9 prevented tumor necrosis. The results suggest that matrix metalloproteinase-9 is involved not only in the regulation of extracellular matrix degradation, but also in the processes of cell proliferation and neoangiogenesis in skin melanoma. Therefore, this enzyme can be considered as a potential therapeutic target.

[TSPO ligand PK11195 and MAPK inhibitor UO126 modulate tspo expression level].

Translocator protein TSPO and MAPK signaling partway are important regulators of numerous cell functions including carcinogenesis, cell proliferation and apoptosis. We have studied MAPK inhibitor UO126 and TSPO specific ligand PK11195 effects on TSPO expression level in melanoma cells. Using immunocytochemical staining, we have detected increased expression of TSPO after incubation with PK11195 and UO126 in all concentrations. These results were confirmed by real time PCR: the increase in mRNA TSPO expression level was detected after incubation with PK11195 in concentration 100 nmol/L, and with UO126 in concentration 10 micromol/L. In the case of a combination of treatments with PK11195 and UO126, we also observed an increase in the level of TSPO expression. So, we conclus de that TSPO ligand PK11195 and MAPK signaling partway inhibitor UO126 modulate TSPO expression. These data are crucial for indentifying of regulatory processes for TSPO expression. Pathogenetic interconnection between MAPK signaling partway and TSPO is important for novel therapeutic approaches in melanoma treatment.

Interaction between single nucleotide polymorphism in catalase gene and catalase activity under the conditions of oxidative stress.

Catalase is an antioxidant enzyme the activity of which is crucial for the protection against damage caused by reactive oxygen species. The -262C>T polymorphism in the promoter region of catalase gene was found to be associated with altered catalase levels. In this study, peripheral blood mononuclear cells catalase activity was measured after H(2)O(2)-induced oxidative stress. C/T and T/T genotypes were associated with the decrease of catalase levels in contrast to C/C donors who had elevated catalase activity in the presence of 0.4 and 0.7 mM H(2)O(2). Genotype-dependent response of catalase activity to oxidative stress might be related to the predisposition of catalase mutant allele carriers to disorders mediated by oxidative stress.

Changes in the levels of N-cadherin and PCNA in skin melanoma cells are mediated through matrix metalloproteinase 9.

Immunohistochemical studies revealed increased level of matrix metalloproteinase 9 in skin melanoma cells. Inhibition of matrix metalloproteinase 9 with interfering RNA changed the level of PCNA and reduced N-cadherin content in melanoma cells. This attests to the involvement of matrix metalloproteinase 9 in the realization of invasion and metastatic growth and in the regulation of tumor growth and progress.

[Apoptosis modulation by PMA occurs independently of TsPO expression in squamous cell carcinoma cells].

Phorbol esters are known to modulate protein kinase C activity - enzyme involved in cell proliferation, differentiation and apoptosis regulation in skin cells. Besides it phorbol-12-myristate 13-acetate was shown possible to modulate promoter activity of TsPO - protein that is involved in steroidogenesis and cell proliferation regulation. Caspase-3 and TsPO expression was measured in squamous cell carcinoma cells after incubation with phorbol-12-myristate 13-acetate and ultraviolet radiation. Following alterations of TsPO and caspase-3 levels are explained by different mechanisms of regulation.

Peculiarities of cell cycle kinetics in normal keratinocytes and melanocytes after exposure to UV-B radiation.

We studied changes in cell cycle and apoptosis in normal keratinocytes and melanocytes after exposure to UV-B radiation. UV-B radiation modulated proliferation rate in the studied cell and produced an apoptosis-modulating effect in epidermal melanocytes.