Ustinia A. Bokova

Background. DNA methylation regulates numerous biological processes, mediating normal development. Alterations in methylation patterns are associated with multiple pathological conditions like hereditary diseases and cancer, making them valuable clinical biomarkers for patient stratification, disease monitoring, early diagnosis, and prediction of response to therapy. Highly targeted, high-throughput methodologies focusing on critical genomic loci enable precise identification of distinct methylation signatures. the aim of the study was to analyze and summarize literature data describing the use of high-throughput DNA methylation analysis technologies, including those based on targeted approaches. Material and Methods . A systematic analysis of literature data was conducted using the PubMed, Web of Science, and Scopus databases, focusing on the characteristics of high-throughput DNA methylation analysis used in cancer and some genetic diseases. A total of 113 sources were analyzed, chronologically covering the period from 2000 to June 2025, 32 of which were used to write the review. Results. The existing technologies for high-throughput methylome analysis, DNA conversion methods, and their advantages and limitations were summarized. In addition, the current targeted enrichment methods, their strengths and weaknesses, and potential applications in scientific and diagnostic practice were discussed. Conclusion. DNA methylation analysis has evolved from a basic research tool into a cornerstone of translational medicine, particularly in oncology. Modern methylome analysis techniques facilitate the discovery of epigenetic markers critical for diagnosing diseases, assessing prognosis, guiding therapy selection, and identifying molecular targets for targeted drugs. Targeted DNA enrichment increases analytical precision and sensitivity while reducing costs. Furthermore, specialized strategies permit targeted analysis even with challenging samples. Combined with the flexibility to focus on specific genomic regions, these advantages make targeted approaches viable not only in academic research but also in routine clinical diagnostics.

Cellular plasticity, a critical feature of embryonic development, is often reactivated in cancer, enabling tumor cells to acquire stem-like properties and facilitate uncontrolled growth. This study compares cellular plasticity in cancer and early human development by integrating single-cell data from three cancers and embryoid bodies. The analysis identifies a shared, highly proliferative progenitor-like state, driven by an E2F-TFDP transcriptional axis and enriched in key cell-cycle regulators UBE2C , TOP2A , BIRC5 , and NUSAP1 . Functionally linked to cell-cycle progression and DNA repair, this state acts as a lineage hub within tumors. Spatial transcriptomics revealed that this state is enriched in lung adenocarcinoma and in lung tissue with basal cell hyperplasia, a potential precursor to squamous cell carcinoma. Validation across eight cancer types demonstrates a prevalence of this program in tumors versus normal cells. Collectively, these findings define a conserved mechanism of tumor plasticity with potential for early detection and targeted therapy. Abstract Figure:

A personalized approach is a priority in the diagnosis and treatment of cancer. Accurate diagnostic tests for assessing the tumor’s molecular characteristics and optimizing the treatment strategy are essential for effective personalized treatment. Multigene NGS panels simultaneously detect a variety of genetic disorders, including single nucleotide substitutions, insertions and deletions, DNA copy number variations, translocations, microsatellite instability, and tumor mutational burden. This is crucial for selecting targeted therapy and immunotherapy, assessing prognosis, and monitoring the progression of cancer. This review discusses the evolution of precision oncology in the 21st century, the possibilities and advantages of NGS in molecular oncology, the technical and analytical characteristics of various multigene NGS panels with both amplification-based and hybridization-based enrichment, and their clinical applications in molecular diagnostics. The review focuses specifically on existing Russian and international NGS technologies. It examines available Russian and international multigene NGS panels, including problems of their application in routine cancer care in Russia.

Aim of the study : to provide a contemporary overview of the nomenclature and terminology of Recklinghausen– Engel syndrome (parathyroid osteodystrophy), its pathogenesis, and characteristic morphological and radiological manifestations. Special attention is paid to the differential diagnosis between brown tumor and skeletal giant cell tumor. The study aims to present the clinical and morphological criteria for distinguishing these conditions, systematized in a comparative table. Material and Methods . A literature search was conducted in the Medline, PubMed, Elibrary, and other databases. Of the 100 identified publications, 30 were included in this systematic review. We report a case of a resected brown tumor of the femur in a 58-year-old man with previously undiagnosed hyperparathyroidism, detailing the morphological diagnostic features and the disease course. Results. Recklinghausen-Engel syndrome (parathyroid osteodystrophy) is a rare but clinically significant metabolic skeletal disorder that develops in the setting of hyperparathyroidism. It is characterized by marked osteoclastic activity, the formation of cystic lesions, subperiosteal resorption, and so-called brown tumors. Despite its benign nature, this syndrome often mimics aggressive giant cell bone tumors, which complicates the interpretation of clinical, radiological, and morphological data and increases the risk of diagnostic errors. Conclusion. This article discusses the diagnostic challenges in the absence of overt clinical symptoms of hyperparathyroidism and emphasizes the key role of the pathomorphologist within a multidisciplinary team. Timely detection of parathyroid hormone hypersecretion and radical removal of the hyperplastic or neoplastic parathyroid gland lead to the regression of bone lesions and a favorable prognosis.

Diagnosis and selection of effective therapy of soft tissue sarcomas (STS) are complicated by low incidence and significant histological variability of these tumors. Development of molecular and genetic testing methods is aimed at improving differential diagnosis of different types of STS and identification of genetic abnormalities which can potentially serve as targets for therapy. Development of effective treatment methods requires adequate preclinical models capable of recreating biological features of the tumors. The article presents molecular and genetic testing methods for STS diagnosis and therapy, advances in in vitro STS models, problems with their use in preclinical studies, as well as possibilities of using primary cell cultures for personalized treatment.

Relevance. Non-small cell lung cancer (NSCLC) is the leading cause of cancer-­related mortality worldwide due to the high recurrence and metastasis rates. It is generally accepted that metastases and recurrences are formed by tumor cells with a highly invasive, stem and chemoresistant phenotype. Tumor hybrid cells (THCs) formed by the fusion of tumor cells with a wide range of normal cells: macrophages, fibroblasts, mesenchymal stem cells, etc. are considered to be potential metastasis and recurrence-­initiating cells. However, the phenotypic diversity of THCs, and their association with disease progression remain poorly understood. The aim of the study was to characterize the population composition of THCs in NSCLC and its association with clinicopathological characteristics, metastasis and recurrence. Materials and Methods. A total of 50 patients with NSCLC were included. Fresh frozen samples of tumor tissue obtained during resection and morphologically verified were used to analyze types and number of THCs. THCs were analyzed by flow cytometry in primary tumors using markers for tumor cells, cancer stem cells, leukocytes, macrophages and fibroblasts. Results and Discussion. THCs were detected in all NSCLC patients. Most THCs demonstrated leukocyte, macrophage and stem characteristics. The number and frequency of THCs depended on neoadjuvant chemotherapy. THCs with leukocyte and stem cell markers (pan-­CK+CD45+CD44+CD73+) were associated with locoregional recurrence, whereas THCs with macrophage and stem cell markers (EpCAM+CD45+CD44+CD73+CD163+) — with distant metastases. Conclusion. This study is the first to comprehensively describe the population composition of THCs in NSCLC, their association with clinicopathological characteristics, neoadjuvant chemotherapy and disease prognosis. Detection of prognostically relevant THCs could be an effective approach for predicting the risk of metastasis and recurrence of NSCLC and the basis for the development of therapy focused on the prevention of cancer progression.

Soft tissue sarcomas are rare tumors (about 1 % of all malignant neoplasms) and include more than 70 histological subtypes, the pathogenetic features of which remain unclear. This is largely due to both quantity and volume of clinical material and high heterogeneity of the disease. Given the rarity and heterogeneity of each individual subtype of soft tissue sarcoma, there is an urgent need to develop universal model systems to understand the molecular changes that determine tumor biology. Such systems include CDX models (cell line-derived xenograft), created from cell lines, PDX (patient-derived xenograft), obtained from primary tumor/metastasis cells, both a whole fragment of surgical material and from a cell suspension; humanized animals containing various human immune cells, and GEM (genetically engineered mouse) models, which are created through transfection of genetic changes characteristic of different subtypes of soft tissue sarcomas. To create these systems, not only widely available mouse models are used, but also other animals, such as fish (Danio rerio) , rats, pigs, and dogs. Another important goal of using animal models is to screen the effectiveness of modern drugs. To date, treatment of various subtypes of soft tissue sarcomas is based on standard protocols of chemotherapy (doxorubicin, epirubicin, dacarbazine, ifosfamide) and surgical resection. In the case of inoperable forms or late stages of soft tissue sarcomas, animal models are a potential tool in predicting the effectiveness of therapy and personalized selection of treatment regimens. In this regard, studies of the mechanisms of targeted action on specific molecules and the use of humanized animals for the development of new approaches to immunotherapy are of particular relevance. The current review discusses animal model systems of the three most common types of soft tissue sarcomas: liposarcomas, undifferentiated pleomorphic and synovial sarcomas, as well as the use of these models to find new therapeutic solutions. Conclusion. Currently, PDX and GEM models are widely used to identify molecules and signaling pathways involved in the development of sarcomas, identify tumor-initiating cells, and assess the chemoresistance of known drugs and new drugs at the level of the entire tumor ecosystem. However, the key problems of animal models of soft tissue sarcomas remain changes in their composition and phenotype compared to the original tumor, poor survival of surgical material, and lack of cellular immunity in immunocompetent models, high cost, and the length of time it takes to create and maintain the model. A solution to one of the problems may be the use of humanized animals with PDX, which implies the presence in the model of immune, stromal and tumor components that are as close as possible to the human body.

Cancers are one of the leading causes of mortality in the world. Cellular and physiological mechanisms of cancer development remain not well defined. In vivo models are an attractive approach for understanding of cancer origin and progression. This review presents current state of experimental in vivo systems including syngeneic models, patient-derived xenografts (PDX), cell line-derived xenografts (CDX) and various animals – humanized and genetically engineered models (GEM). These models provide opportunities for developing patients’ avatars, lifetime visualization of tumor migration and invasion at the organism level, and the evaluation of new therapeutic methods aimed at primary tumors, metastases, and cancer prevention. We also discuss the problems of choosing the optimal model and potential solutions for their overcoming.

Background. Cancer of unknown primary (CUP) is a metastatic lesion with diffcult identifcation of the primary tumor site using standard diagnostic approaches. Although the incidence of CUP is not high, this type of cancer often shows a high aggressiveness and therapy resistance and results in poor patient survival. The mechanisms of CUP origin are not clear, and further studies are needed.This study aims to analyze the mutational landscape of CUP and identify specifc genetic alterations.Material and Methods. Whole exome sequencing was used to analyze the mutational landscape of CUP. Results. CUP had single nucleotide variants (SNVs) in the EPHA8 (ephrin receptor) gene. CUP also harbored copy number variations (CNAs) in the ID2, FOXD4, ZMYND11, ZNF596, KIDINS220, LRRN1, GEMIN4, CEP72, TPPP, and MXRA5 genes. According to functional enrichment analysis, these genes are involved in the regulation of transcription, biogenesis of microRNA, cellular cytoskeleton, adhesion, extracellular matrix remodeling, proliferation, apoptosis, and epithelial-mesenchymal transition.Conclusion. Cancer of unknown primary harbors mutations in the genes that regulate different biological processes particularly cell motility.