[Adrenocortical cancer: morphological variants, immunohistochemical characteristics]. / Adrenokortikal'nyi rak: morfologicheskie varianty, immunogistokhimicheskaya kharakteristika.

Adrenocortical cancer (ACC) is a rare endocrine malignancy of the adrenal cortex, which has an unfavorable prognosis and extremely aggressive clinical behavior in most cases. Nevertheless, cases of a more favorable disease course with late metastasis and slow progression have been described. In 2017, the International Agency for Research on Cancer (IARC) and the World Health Organization (WHO) in the 4th edition of the Classification of Tumors of the Endocrine Organs identified histological variants of ACC, such as classical, oncocytic, myxoid, and sarcomatoid ones, indicating the morphological heterogeneity of this tumor. OBJECTIVE: To provide a detailed description of the morphological variants of ACC with an emphasis on their histological characteristics and the expression of immunohistochemical markers. MATERIALS AND METHODS: A total of 75 cases of ACC were analyzed in the adult population diagnosed as having the morphological variants in accordance with the International Histological Classification of Adrenal Tumors (WHO, 2017). Monoclonal antibodies to SF1, Inhibin A, Melan A, Ki-67, p53, and antimitochondrial antibodies were used for immunohistochemical diagnosis. RESULTS: The classic, oncocytic, and myxoid subtypes of ACC were found in 51 (68%), 15 (20%), and 9 (12%) cases, respectively. The functional activity of the tumors was observed in 43% (n=18) in the classic variant of ACC; moreover, the clinical picture was manifested by the symptoms of hypercorticism (38%) and virilization (5%). There were no significant differences in hormonal activity between different morphological variants. The characteristics of the above histological variants of the tumor was determined with a description of growth patterns that can improve the diagnosis of ACC. The diagnosis of ACC can be confirmed by an immunohistochemical study; the required minimum panel of markers should include SF1, Melan A, and Inhibin A. The Ki-67 proliferative activity index showed significant differences (p=0.0056) when it was determined in the morphological variants of ACC. CONCLUSION: Despite the determination of a minimal immunohistochemical panel to confirm the diagnosis of ACC, it is important to remember that each histological variant may be characterized by the different expression of immunohistochemical markers. The identification of morphological variants of ACC and the use of specific, sensitive, and prognostically significant immunohistochemical markers will allow clinicians and pathologists to more accurately judge the biological properties of this tumor and the clinical course of the disease.

Constitutive Expression of NRAS with Q61R Driver Mutation Activates Processes of Epithelial-Mesenchymal Transition and Leads to Substantial Transcriptome Change of Nthy-ori 3-1 Thyroid Epithelial Cells.

The Q61R mutation of the NRAS gene is one of the most frequent driver mutations of thyroid cancer. Tumors with this mutation are characterized by invasion into blood vessels and formation of distant metastases. To study the role of this mutation in the growth of thyroid cancer, we developed a model system on the basis of thyroid epithelial cell line Nthy-ori 3-1 transduced by a lentiviral vector containing the NRAS gene with the Q61R mutation. It was found that the expression of NRAS(Q61R) in thyroid epithelial cells has a profound influence on groups of genes involved in the formation of intercellular contacts, as well as in processes of epithelial-mesenchymal transition and cell invasion. The alteration in the expression of these genes affects the phenotype of the model cells, which acquire traits of mesenchymal cells and demonstrate increased ability for survival and growth without attachment to the substrate. The key regulators of these processes are transcription factors belonging to families SNAIL, ZEB, and TWIST, and in different types of tumors the contribution of each individual factor can vary greatly. In our model system, phenotype change correlates with an increase in the expression of SNAIL2 and TWIST2 factors, which indicates their possible role in regulating invasive growth of thyroid cancer with the mutation of NRAS(Q61R).

[Histopatological and molecular genetic characteristics of clinically aggressive variants of papillary thyroid carcinoma]. / Gistologicheskaia i molekuliarno-geneticheskaia kharakteristika klinicheski agressivnykh variantov papilliarnogo raka shchitovidnoi zhelezy.

Papillary carcinoma is the most commonly diagnosed form of well-differentiated thyroid cancer that is generally characterized by a favorable prognosis. However, a number of relatively rare variants of this tumor, such as papillary carcinoma of high cells, papillary carcinoma of columnar cells, a diffuse sclerosing variant and recently described cancer of shoe nail cell type, are characterized by a less favorable clinical course, a high frequency of distant metastasis, and relatively low overall and relapse-free survival rates. In this connection, it is important to recognize these options at the stage of a primary morphological study. This review of the literature considers the morphological, clinical and molecular genetic features of the above variants of papillary thyroid carcinoma.

[The Novel Short Isoform of Securin Stimulates the Expression of Cyclin D3 and Angiogenesis Factors VEGFA and FGF2, but Does Not Affect the Expression of MYC Transcription Factor].

Pituitary tumor-transforming gene-1 (PTTG1) encodes securin, a multifunctional protein involved in development of various types of cancer. Securin participates in the regulation of sister chromatids separation and the expression of multiple genes involved in the control of the cell cycle, metabolism, and angiogenesis. In several human cell lines, we have found a novel short isoform of securin mRNA, which does not contain exons 3 and 4. After the translation of this new mRNA, a shortened protein is produced that, like the full-size form, is able to activate the transcription of cyclin D3 gene (CCND3), which controls the G1/S transition and angiogenesis factors VEGFA (vascular endothelial growth factor), and FGF2 (fibroblast growth factor 2) in HEK293 cells. However, unlike the full-size protein, the short isoform of PTTG1 does not affect the MYC gene expression because it lacks the DNA-binding domain, which is needed for its interactions with the MYC promoter. Furthermore, the short form of securin does not influence the expression of MYC transcriptional targets, such as TP53 and IL-8. Thus, we found a novel isoform of securin which is able to activate a more restricted repertoire of genes compared to the full-size protein.

Isolation of Large Amounts of Highly Pure Mitochondria for "Omics" Studies.

Ultracentrifugation on a density gradient remains the only reliable way to obtain highly pure mitochondria preparations. However, it is not readily available for any laboratory and has a serious disadvantage of providing low mitochondria yield, which can be critical when working with limited starting material. Here we describe a combined method for isolation of mitochondria for proteomic studies that includes cell disruption by sonication, differential centrifugation, and magnetic separation. Our method provides remarkable enrichment of mitochondrial proteins as compared to differential centrifugation, magnetic separation, or their combination, and it enables the strongest depletion of cytoplasmic components, as assessed by two-dimensional electrophoresis, mass spectrometry, and Western blot. It also doubles the yield of mitochondria. However, our method should not be used for functional studies as most of the isolated organelles demonstrate disturbed structure in electron microphotographs.

Inflammatory Immune Infiltration in Human Tumors: Role in Pathogenesis and Prognostic and Diagnostic Value.

The cellular microenvironment directly and indirectly influences tumor development and possesses prognostic and in some cases diagnostic value. Over the years, understanding of structural organization of the immune/inflammatory moiety of neoplasms as well as in-depth phenotypic and transcriptomic profiling of its cellular components together provide more and more insights in both basic and translational medical science. In this review, we will discuss the specific roles of various stromal cells and their impact on neoplastic progression as well as address the use of quantitative and phenotypic analysis of immune/inflammatory infiltrate for diagnostics and predicting the clinical course of human malignancies.

[Preparation of recombinant serpins B3 and B4 and investigation of their specific interactions with antibodies using hydrogel-based microarrays].

The objective of this work was to obtain preparations of recombinant squamous-cell carcinoma antigens (serpins B3 and B4) and to investigate their interactions with different monoclonal antibodies using hydrogel-based microarrays (biochips). Two genetic constructs encoding full-length serpin B3 and serpin B4 molecules were created to produce recombinant SPB3 and SPB4 proteins carrying a N-terminal His6-tag. Monoclonal antibodies against serpin B3 (H3, C5, H5, H81, and G9) were also obtained. An experimental gel-based biological microchip was designed to contain gel elements that carry immobilized antibodies against SPB3, immobilized commercial monoclonal SCC107 and SCC140 antibodies against squamous-cell carcinoma antigen (SCCA), and gel elements with immobilized SPB3 or SPB4. Judging by the specificity of recombinant SPB3 and SPB4, which bind to monoclonal antibodies against SCCA and, according to the manufacturer's data, can recognize conformational epitopes of both SPB3 and SPB4, it was concluded that the obtained recombinant serpins had the correct tertiary structure. A biochip-based direct immunoassay showed that SPB4 could bind effectively only to SCC107 and SCC140 antibodies, while SPB3 interacted specifically not only with these antibodies, but also with H3 and C5 monoclonal antibodies. Using biochip-based sandwich immunoassay, a pair of monoclonal antibodies SCC107/C5 that interacted specifically with serpin B3 but did not interact with serpin B4 was identified. Thus, it has been demonstrated that serpin B3 can be selectively determined in the presence of highly homologous serpin B4 using a biochip-based assay.

[Innate immunity against viruses].

Viruses are obligate parasites which are able to infect cells of all living organisms. Multiple antiviral defense mechanisms have appeared early in evolution of the immune system. Higher vertebrates have the most complex antiviral immunity which is based on both innate and adoptive immune responses. However, majority of living organisms, including plants and invertebrates, rely exclusively on innate immune mechanisms for protection against viral infections. There are some striking similarities in several components of the innate immune recognition between mammals, plants and insects, rendering these signaling cascades as highly conserved in the evolution of the immune system. This review summarizes recent advances in the field of innate immune recognition of viruses, with particular interest on pattern-recognition receptors.

Autoantibodies to tumor-associated antigens as cancer biomarkers.

Malignant tumors induce humoral immune response in cancer patients, although the incidence of such autoantibody responses against individual tumor-associated antigens (TAA) is rather low. To increase predictive value of TAA-recognizing autoantibodies as potential cancer biomarkers, TAAs should be combined into protein arrays. Here we review recent advances in the application of such arrays and summarize data concerning most promising antigens. We also review the methods of cloning TAA-recognizing autoantibodies, generation of human hybridomas and screening of recombinant human immunoglobulin libraries.

Cancer-associated antigens and antigen arrays in serological diagnostics of malignant tumors.

The appearance of antibodies to cancer-associated antigens in biological fluids (particularly, in blood sera) of cancer patients is now a well-established fact, and their detection by immunochemical methods is a promising approach to diagnostics of malignant neoplasms. In this review, we consider some immunobiological aspects of the most extensively studied cancer-associated B-cell antigens, various applications of autoantibodies as cancer biomarkers, and prospects for the use of antigen arrays for improving diagnostic sensitivity.

[Contemporary molecular-biological methods of cancer diagnosis and monitoring based on the humoral immune response to tumor-associated antigens].

Humoral immune response to tumor-associated antigens in cancer patients can be used as a basis for disease diagnosis and monitoring. Moreover, identification of molecular targets of such response may be used to develop antigen-specific anticancer vaccines. Here, we review the main approaches to identification and study of tumor-associated antigens recognized by serum antibodies. We also focus on the challenges that must be met before serological antigens can be used in clinical cancer diagnostics.

[Onconeural antibodies as a tool for diagnosis of malignant tumors and paraneoplastic neurological disorders].

Paraneoplastic neurological syndromes (PNS) are autoimmune neurodegenerative diseases that develop as a result of the cross-reactivity of the tumor-specific immune effectors with neurons of central and peripheral nervous systems. So-called onconeural antibodies, which are detected in sera of PNS patients, are not only crucial diagnostic markers of PNS and associated tumors, but also have a considerable potential in the serological diagnosis of cancer as a whole. In this review we discuss the role of onconeural antibodies in serological diagnosis of PNS and associated tumors as well as their potential in diagnosis and prognostication of a broad spectrum of malignant tumors.