Melanoma is an aggressive cancer characterized by the acidification of the extracellular environment. Here, we showed for the first time that extracellular media acidification increases proliferation, migration, and invasion of patient-derived metastatic melanoma cells and up-regulates cell-surface expression of acid-sensitive channels containing the ASIC1a, α-ENaC, and γ-ENaC subunits. No influence of media acidification on these processes was found in normal keratinocytes. To control metastatic melanoma progression associated with the ASIC1a up-regulation, we proposed the ASIC1a inhibitor, -mambalgin-2 from Dendpoaspis polylepis venom. Recombinant analog of mambalgin-2 cancelled acidification-induced proliferation, migration, and invasion of metastatic melanoma cells, promoted apoptosis, and down-regulated cell-surface expression of prooncogenic factors CD44 and Frizzled 4 and phosphorylation of transcription factor SNAI. Confocal microscopy and affinity purification revealed that mambalgin-2 interacts with heterotrimeric ASIC1a/α-ENaC/γ-ENaC channels on the surface of metastatic melanoma cells. Using the mutant variant of mambalgin-2 with reduced activity toward ASIC1a, we confirmed that the principal molecular target of mambalgin-2 in melanoma cells is the ASIC1a subunit. Bioinformatic analysis confirmed up-regulation of the ASIC1 expression as a marker of poor survival prognosis for patients with metastatic melanoma. Thus, targeting ASIC1a by drugs such as mambalgin-2 could be a promising strategy for metastatic melanoma treatment.

Metastatic melanoma is a highly malignant tumor. Melanoma cells release extracellular vesicles (EVs), which contribute to the growth, metastasis, and malignancy of neighboring cells by transfer of tumor-promoting miRNAs, mRNA, and proteins. Melanoma microenvironment acidification promotes tumor progression and determines EVs’ properties. We studied the influence of EVs derived from metastatic melanoma cells cultivated at acidic (6.5) and normal (7.4) pH on the morphology and homeostasis of normal keratinocytes. Acidification of metastatic melanoma environment made EVs more prooncogenic with increased expression of prooncogenic mi221 RNA, stemless factor CD133, and pro-migration factor SNAI1, as well as with downregulated antitumor mir7 RNA. Incubation with EVs stimulated growth and migration both of metastatic melanoma cells and keratinocytes and changed the morphology of keratinocytes to stem-like phenotype, which was confirmed by increased expression of the stemness factors KLF and CD133. Activation of the AKT/mTOR and ERK signaling pathways and increased expression of epidermal growth factor receptor EGFR and SNAI1 were detected in keratinocytes upon incubation with EVs. Moreover, EVs reduced the production of different cytokines (IL6, IL10, and IL12) and adhesion factors (sICAM-1, sICAM-3, sPecam-1, and sCD40L) usually secreted by keratinocytes to control melanoma progression. Bioinformatic analysis revealed the correlation between decreased expression of these secreted factors and worse survival prognosis for patients with metastatic melanoma. Altogether, our data mean that metastatic melanoma EVs are important players in the transformation of normal keratinocytes.

We have previously shown that extracellular vesicles secreted by metastatic melanoma cells stimulate the growth, migration, and stemness of normal keratinocytes. This study showed for the first time that extracellular vesicles secreted by the metastatic melanoma cell lines mel H, mel Kor, and mel P contain, both at the mRNA and protein levels, the 7-type nicotinic acetylcholine receptor (7-nAChR), which is involved in the regulation of the oncogenic signaling pathways in epithelial cells. Incubation with the vesicles secreted by mel H cells and containing the highest amount of mRNA coding 7-nAChR increased the surface expression of 7-nAChR in normal Het-1A keratinocytes and stimulated their growth. Meanwhile, both of these effects disappeared in the presence of -bungarotoxin, an 7-nAChR inhibitor. A bioinformatic analysis revealed a correlation between the increased expression of the CHRNA7 gene coding 7-nAChR in patients with metastatic melanoma and a poor survival prognosis. Therefore, extracellular vesicles derived from metastatic melanoma cells can transfer mRNA coding 7-nAChR, thus enhancing the surface expression of this receptor and stimulating the growth of normal keratinocytes. Targeting of 7-nAChR may become a new strategy for controlling the malignant transformation of keratinocytes.

Introduction. The increasing clinical use of cellular technologies suggests the possibility of long-term storage of the cellular product while maintaining its viability and basic properties. The procedures of cell’s cryopreservation used in laboratory as well in clinical practice differ a lot. Each method includes two tasks to solve: what is the optimal freezing medium to use and what cryopreservation procedure to prefer. In this paper, we present the method utilized in our center for bone marrow cell cryopreservation. The freezing was carried out in nitrogen vapor after adding the medium containing 95 % dextran [average mw 50 000–70 000] and 5 % dimethylsulfoxid.Purpose. To show that the proposed method of cryopreservation of dendritic cells is highly effective, simple, reproducible and most convenient for clinical use.Materials and methods. Viability, expression of surface antigens and stimulating activity towards allogeneic T lymphocytes of cryopreserved mature dendritic cells cultured from peripheral blood monocytes were evaluated.Results. The first cryopreservation resulted in the death of a small amount of cells. The second freezing procedure increased the proportion of dead cells. Meanwhile, the difference in the expression of the surface antigens in fresh, cryopreserved and re-cryopreserved dendritic cells was not statistically significant. The level of stimulating activity of fresh and cryopreserved dendritic cells did not significantly differ. Conversely the proliferation of allogeneic T lymphocytes was decreased after stimulation with re-cryopreserved dendritic cells.Conclusion. The presented method of cryopreservation allows to preserve the viability and basic functions of dendritic cells. After thawing dendritic vaccine could be administered to patients after being diluted in an isotonic saline without washing, which makes this method the most convenient for clinical use.

Меланома - одна из наиболее агрессивных злокачественных опухолей человека. Заболеваемость меланомой и смертность от неё неуклонно растут. Ультрафиолетовое излучение – основной фактор риска развития меланомы, однако, по-видимому, это – не единственный экологический фактор, вовлеченный в меланомогенез. Возможность вирусной природы меланомы обсуждается давно. Среди кандидатов на роль этиологических агентов меланомы фигурируют папилломавирусы. Интерес к ним велик ввиду того, что в эту группу вирусов входят сильные канцерогены, вызывающие рак шейки матки и некоторые другие формы рака. Цель обзора - анализ данных литературы об ассоциации меланомы с вирусами папиллом человека. Несколько исследовательcких групп обнаружили папилломавирусы в образцах меланомы кожи и слизистых оболочек; при этом иногда выявлялись и вирусы типов высокого онкогенного риска. Проверка экспрессии генома вируса при обнаружении в образце меланомы вирусной ДНК в подавляющем большинстве работ не проводилась. Объективной трудностью при проведении подобных исследований является проблема отрицательного контроля: папилломавирусы регулярно обнаруживаются на нормальной коже и слизистых оболочках человека, что делает реальной угрозу контаминации образца опухоли вирусной ДНК invivo. Преодолению этого затруднения мог бы помочь комплексный подход: выявление вируса на трех уровнях, - ДНК, мРНК вирусных онкогенов и соответствующих онкобелков. Имеются ограниченные данные об ухудшении прогноза заболевания при положительном по папилломавирусам статусе меланомы, но систематическое наблюдение за сколь-нибудь значительными группами больных, различающимися по данному статусу опухоли, не проводилось. Вопрос об ассоциированности меланомы с вирусами папиллом человека остается открытым.

Imatinib mesilate is a well-known antitumor target inhibitor of protein tyrosine kinase, which is effective in different cancer types expressing Bcr / Abl and, in particular, in hemoblastosis. A higher interest to imatinib during the COVID-19 epidemic is explained by the fact that cancer patients are one of the COVID-19 risk groups. Moreover, imatinib target mechanism of action, which is effective in cancer, can have a high potential against the most severe COVID-19 complication such as the disease associated pulmonary fibrosis. COVID-19 associated interstitial pulmonary fibrosis develops as an autoimmune process caused by systemic inflammation with atypical (idiopathic) pneumonia resulting from acute respiratory distress syndrome with the tyrosine kinase mechanism of signaling pathway activation and cellular response. Experi-mental and clinical results showing antifibrotic and dose-related antithrombotic imatinib effect demonstrate perspective use of this antitumor agent to correct COVID-19 associated pneumonia causing a high death rate of patients with COVID-19.The review presents literature data of 2001–2020 discussing pathologic genetic and clinical characteristics of the fibrosis which exacerbates COVID-19 pneumonia in adults. The sequence of the disease processes demonstrates that disease progression with the decreasing oxygen saturation in the peripheral blood intensifies local thrombosis in the lungs. As a result, hypoxia is developing, which is difficult to control and can cause lethal outcome in severe cases. Yet, the conventional antifibrotic and thrombolytic agents can only partially control the process of pneumofibrosis including that of cancer patients. The approximate antifibrotic dose of imatinib 400 mg / day is therapeutic for oncopatho-logy. The antitumor drug registered in many countries and well described side effects and contraindications needs no long-term registration studies for a new indication, therefore, it may be easily prepared for clinical testing.