Objective: This study examined the frequency of entosis in solid tumors of various origins (colorectal cancer, breast cancer, and lung cancer) and its association with clinical and pathological characteristics. It also examined survival and copy number alterations (CNAs) in genes associated with stem cells. The aim was to assess the potential prognostic value of entotic events in tumors. Methods: A total of 238 patients were included: 96 with colorectal cancer (CRC), 45 with lung cancer (LC), and 97 with breast cancer (BC). Entotic cell-in-cell (CIC) structures were evaluated on hematoxylin-eosin-stained slides using Mackay's criteria. A CIC frequency >0.1 per 20 high-power fields was considered positive. Clinicopathological parameters, overall survival (CRC), metastasis-free survival (LC and BC), and CNA profiles of stemness-related genes were analyzed. Amplifications of MAP1LC3A and other chromosomal loci were assessed. Results: CRC demonstrated the highest entosis rate, more than two-fold higher compared with BC and LC (p < 0.05). Entosis correlated with high tumor grade (G3) in CRC (p = 0.03). In LC, CIC-positive tumors were more frequent in patients with lymph-node metastases (p = 0.02), whereas in BC, the opposite trend was observed (p = 0.02). It was noted that in patients with stage III-IV LC, the frequency of entosis was significantly higher than in patients with stage I-II cancer (p = 0.03). CIC-positive status was associated with poorer overall survival in CRC (p = 0.03) and reduced metastasis-free survival in LC (p = 0.011). In breast cancer, no statistically significant survival differences were observed. Tumors harboring two or more stemness-gene amplifications showed significantly higher entosis frequency regardless of tumor site. A strong association was identified between entosis and MAP1LC3A amplification. Conclusions: Enosis is not a random morphological phenomenon but a process associated with unfavorable tumor characteristics, high malignancy, reduced survival, and amplification of stem cell-related genes. The results of this study confirm the working hypothesis that entosis may contribute to the emergence of aneuploid clones of tumor cells, including those containing amplifications of stem cell-associated genes. This positions entosis as a potential factor in tumor genetic heterogeneity, which is particularly important in the context of therapeutic selection pressure. The observed association between high entosis frequency and the presence of >= 2 stem cell gene amplifications, as well as its association with poor prognosis in colorectal and lung cancer, highlights its potential value as a prognostic indicator. Furthermore, MAP1LC3A amplification data may serve as a molecular marker of entotic activity and a potential therapeutic target.

The use of postmortem (autopsy) material in fundamental and applied biomedical research significantly facilitates the collection of biomaterial for statistically robust sample cohorts. However, natural adaptive processes to developing cellular stress in the early postmortem period, caused by oxygen and nutrient deprivation, trigger the activation of numerous genes promoting cell survival under stress. Many of these activated pathways are also crucial for tumor cell survival in vivo, as evidenced by various transcriptomic studies. This study aimed to investigate the potential influence of postmortem interval (PMI) duration on gene expression in normal and tumor tissues. Using a model of chemically induced hepatocellular carcinoma in mouse liver, we comparatively analyzed the dynamics of transcript levels for several genes (BRCA1, BRCA2, CHEK1, CHEK2, ATM, CDK12) in paired samples of normal and tumor tissue over a 24-h PMI using RT-qPCR. In normal tissue, gene expression increased significantly, while tumor tissue demonstrated relative transcriptional stability, with no substantial changes in the studied transcript levels. A critical finding was the observed convergence of expression profiles: initial differences between the tissues were completely eliminated by 24 h PMI. This pattern developed despite formally adequate RNA quality (RQN) and the absence of clear signs of progressive autolysis in histology, indicating the insufficiency of standard quality criteria for detecting postmortem changes. These findings collectively underscore the critical importance of minimizing and controlling PMI during the biobanking of oncological samples for reliable transcriptomic research.

Circulating small extracellular vesicles (sEVs) are emerging as potential biomarkers for glioblastoma progression. This study aimed to compare the levels of matrix metalloproteinases (MMP2 and MMP9), terminal complement complex (C5b-9), and VEGF-A in circulating sEVs in glioblastoma patients (GBMPs) with and without tumor recurrence. Using differential ultracentrifugation, sEVs were isolated from blood samples of GBMPs with no tumor recurrence for over one year (n = 6) and after first relapse (n = 14). The vesicles were characterized and quantified using flow cytometry. In both groups, C5b-9 was predominantly detected on tumor-specific circulating sEVs (glial fibrillary acidic protein (GFAP)-positive sEVs) with high VEGF-A expression, while C5b-9 was significantly less frequent on sEVs with low VEGF-A expression (p < 0.05). GFAP+VEGF+dimMMP2-C5b-9+ vesicles were rarely detected in GBMPs without relapse, suggesting their potential utility as biomarkers for a favorable relapse-free prognosis. In recurrent GBMPs, a positive correlation was observed between GFAP+VEGF+bright MMP2+C5b-9+ sEVs and MGMT gene promoter methylation levels (r = 0.543, p < 0.05). Additionally, a trend toward a negative correlation was found between GFAP+VEGF+bright MMP2+C5b-9- sEVs and mutant p53 expression in primary tumor tissue (r = −0.44, p = 0.114). These findings suggest that sEV profiles may serve as valuable prognostic markers for glioblastoma recurrence and treatment responses.

Background/Objectives: In gastric cancer, only a subset of patients benefit clinically from neoadjuvant chemoimmunotherapy, underscoring the need for robust biomarkers that can predict treatment responses and guide personalized immunotherapy. This study aimed to characterize the immune microenvironment of gastric tumors and identify predictive markers associated with therapeutic efficacy. Methods: We prospectively enrolled 16 patients with histologically confirmed, PD-L1–positive (CPS ≥ 1) gastric adenocarcinoma (T2–4N0–1M0). All patients received eight cycles of FLOT chemotherapy combined with pembrolizumab. Treatment response was assessed by Mandard tumor regression grading. Spatial transcriptomic profiling (10x Genomics Visium) and multiplex immunofluorescence were used to evaluate tumor-infiltrating immune cell subsets and PD-1 expression at baseline and after treatment. Results: Transcriptomic analysis differentiated the immune landscapes of responders from non-responders. Responders exhibited elevated expression of IL1B, CXCL5, HMGB1, and IFNGR2, indicative of an inflamed tumor microenvironment and type I/II interferon signaling. In contrast, non-responders demonstrated upregulation of immunosuppressive genes such as LGALS3, IDO1, and CD55, along with enrichment in oxidative phosphorylation and antigen presentation pathways. Multiplex immunofluorescence confirmed a higher density of FoxP3+ regulatory T cells in non-responders (median 5.36% vs. 2.41%; p = 0.0032). Notably, PD-1+ CD8+ T cell and PD-1+ FoxP3+ Treg frequencies were significantly elevated in non-responders, suggesting that PD-1 expression within cytotoxic and regulatory compartments may contribute to immune evasion. No substantial differences were observed in PD-L1 CPS or PD-1+ B cells and PD-1+ macrophages. Conclusions: Our findings identify PD-1+ CD8+ T cells and PD-1+ FoxP3+ Tregs as potential biomarkers of resistance to neoadjuvant chemoimmunotherapy in gastric cancer. Transcriptional programs centered on IL1B/CXCL5 and LGALS3/IDO1 define distinct immune phenotypes that may guide future combination strategies targeting both effector and suppressive arms of the tumor immune response.

BACKGROUND:Squamous cell carcinoma accounts for up to 95% of malignant laryngeal tumors, whereas leiomyosarcoma is exceptionally rare, with fewer than 50 cases reported worldwide. The synchronous occurrence of two histogenetically distinct malignancies in the larynx is extremely uncommon and represents a diagnostic and therapeutic challenge. CASE PRESENTATION:We describe a 66-year-old Caucasian male with synchronous laryngeal tumors: a well-differentiated squamous cell carcinoma of the right vocal fold and a high-grade leiomyosarcoma of the subglottic region. Diagnosis was established using endoscopy, histopathology, and immunohistochemistry. The patient underwent total laryngectomy with permanent tracheostomy followed by adjuvant radiotherapy. At follow-up, no recurrence or progression was detected. CONCLUSION:This case illustrates the rare coexistence of epithelial and mesenchymal malignancies in the larynx and emphasizes the importance of multimodal diagnostic evaluation to guide optimal surgical management.

The study of factors contributing to the adjuvant hormone therapy resistance in estrogen-positive breast cancer (BC) is an important aspect of scientific research. Tamoxifen resistance can be determined by the regulation of the estrogen receptor signaling pathway via membrane tyrosine kinases. Among the various membrane kinases, the activation of the transforming growth factor β1 (TGF-β1) and the triggering of alternative PI3K/Akt signaling can be critical in the hormonal resistance. The aim of the study was to analyze the protein expression of TGF-β1, TGF-βR1, TGF-βR2, pAkt1, cyclin D1 as well as the cell populations of TGF-β1/TGF-βR2, TGF-βR1/TGF-βR2, pAkt!/TGF-βR2 and cyclin D1/TGF-βR2 in tumor tissue depending on the response to tamoxifen treatment. This study included 65 BC patients (T1-4N0-3M0) who received breast-conserving surgery or radical mastectomy, radiation and/or chemotherapy (if indications) and adjuvant hormonal therapy with tamoxifen (5 years, 20 mg/day). Patients without any recurrence or metastasis the adjuvant tamoxifen therapy were classified as a tamoxifen-sensitive subgroup ((TS), N = 55 (84.6%)); patients with distant metastasis or recurrence were classified as resistant to tamoxifen ((TR), N = 10 (15.4%)). Proteins expression as well as cell population expression was assessed using a CytoFLEX flow cytometer (Beckman Coulter, USA). Progression-free survival (PFS) was estimated according to the Kaplan-Meier method, and survival differences between groups were determined using the log-rank test. P-value of less than 0.05 was considered statistically significant. Tumors with high of TGF-βR1 and TGF-PR2 protein expression were more prevalent among TS patients (p = 0.036 and p = 0.006; respectively. See Fig, 1). Increased risk of breast cancer progression was associated with a high pAkt1 expression in tumor cells (p = 0.024, See Fig, 1). Patients with the high pAkt1+/TGF-βR2+ and cyclin D1+/TGFβR2- expression in tumor tissue were less sensitive to tamoxifen treatment (p = 0.006 and р = 0.000; respectively). However, a high percentage of pAkt!-/TGF-βR2+ and cyclin D1+/TGFβR2+ cells was detected in the tumor tissue of patients with the response to treatment (p = 0.001 and p = 0.003, respectively). High TGF-βR2 and pAkt!-/TGF-βR2+ expression was associated with longer progression-free survival (log-rank p = 0.006 and log-rank p = 0.008). Moreover, we demonstrated a significant relationship of both high pAkt1 and pAkt1+/TGF-βR2+, pAkt!+/TGF-βR2- expression with poor survival of breast cancer patients (log-rank p = 0.002 and log-rank p = 0.001, respectively). Our data indicate the possible use of the identified parameters as markers associated with the efficacy of adjuvant tamoxifen therapy. The article contains 1 Figure, 10 References. The Authors declare no conflict of interest.

Background: Proteasomes degrade intracellular proteins. Different proteasome forms were identified. Proteasome inhibitors are used in cancer therapy, and novel drugs directed to specific proteasome forms are developed. Breast cancer (BC) therapy depends on the subtype of the tumor, determined by the expression level of Ki67, HER-2, estrogen and progesterone receptors. Relationships between the presence of specific proteasome forms and proteins that determine the BC subtype remain unclear. Here, using gene expression data in 19,145 tumor samples from 144 datasets and tissues from 159 patients with different subtypes of BC, we investigated the association between the activity and expression of proteasomes and levels of BC subtype markers. Methods: Bioinformatic analysis of proteasome subunit (PSMB1-10) gene expression in BC was performed. Proteasome heterogeneity in BC cell lines was investigated by qPCR. By Western blotting, proteasome composition was assessed in cells and patient tissue lysates. Proteasome activities were studied using fluorogenic substrates. BC molecular subtypes were determined by immunohistochemistry. Results: BC subtypes demonstrate differing proteasome subunit expression pattern and strong PSMB8-10 co-correlation in tumors. A significant increase in chymotrypsin- and caspase-like proteasome activities in BC compared to adjacent tissues was revealed. The subunit composition of proteasomes in tumor tissues of BC subtypes varied. Regression analysis demonstrated a positive correlation between proteasome activities and the expression of Ki67, estrogen receptors and progesterone receptors. Conclusion: BC subtypes demonstrate differences within the proteasome pool. Correlations between the proteasome activity, hormone receptors and Ki67 indicate possible mutual influence. Obtained results facilitate development of novel drug combinations for BC therapy.

Introduction. Luminal breast cancer (BC) occupies the largest proportion in the structure of the entire molecular landscape of this disease. During the ongoing treatment, most often hormone therapy, in part of patients, the disease progression develops, which makes necessary the search for new molecular predictors. Aim of the research. To study the clinical and morphological features of the disease depending on BMI-1 and ROR1 expression in the primary tumor in luminal breast cancer patients during aromatase inhibitors therapy. Materials and methods. The study included 80 patients with T1-2N0-1M0 operable primary breast cancer at the age of 62.1 ± 8.1 years. The primary tumor tissue was studied using immunohistochemistry. Antibodies to ROR1 and BMI-1 were used. The presence and degree of immunostaining, and the percentage of positively stained tumor cells were assessed. The expression parameters of the studied markers were assessed in relation to various clinical and pathological data of the disease. Results. 70 patients had the luminal A subtype, 10 patients had the luminal B/HER2-negative subtype. Positive BMI-1 expression was observed in 64% of cases, ROR1 expression was less common and amounted to 24%. As tumor grade increases, the number of cases with positive ROR1 expression increases (p < 0.05). The level of proliferative activity (Ki67) correlated with positive ROR1 expression (ρ = 0.312, p = 0.03) and BMI-1 (ρ = 0.310, p < 0.031). In presence of metastases to regional lymph nodes, the expression of both markers was significantly higher. The occurrence of distant metastases is associated with high levels of BMI-1 expression by primary tumor cells (p < 0.05). Conclusion. The study clearly demonstrates the correlation of ROR1 and BMI-1 proteins with the clinical and morphological parameters of the primary tumor and the course of the disease in luminal subtypes of breast cancer.

Colorectal cancer is the most common malignancy worldwide. A number of pathological and molecular genetic criteria are currently used as predictors of the disease. They include assessment of MMR deficiency or MSI/MSS status, which among others, determine the immunogenicity of the tumor. In this regard, the evaluation of PD-L1, CTLA-4, and LAG-3 immune checkpoint molecules in different tumor compartments according to MMR status deserves special attention. Multiplex immunohistochemistry was used to evaluate the expression of immune checkpoint molecules in the tumor core and at the invasive margin. Data analysis showed the predominance of PD-L1 (p = 0.011), CTLA-4 (p = 0.004), and LAG-3 (p = 0.013) expression at the invasive margin of dMMR carcinomas compared to pMMR samples. Quantitative analysis of TILs population in the tumor core and at the invasive margin allowed establishment of the predominance of CD3+ and CD8+ lymphocytes at the invasive margin of dMMR carcinomas. Study of the CD163+ macrophages population in the same tumor compartments revealed the predominance of the studied TAMs in the core and at the invasive margin of dMMR carcinomas and the predominance of CD163+ macrophages with PD-L1-phenotype in the tumor stroma. This study revealed a significant predominance of PD-L1, CTLA-4, LAG-3, and CD 3+ ,CD8+ lymphocytes in dMMR colorectal carcinomas. Further research on the immune landscape in different tumor compartments will likely have high prognostic value for CRC patients, as it might expand the criteria for prescribing immunotherapy.

Aim. To examine the pattern of morphological changes, RNA quality number, and gene expression in mouse tissues sampled at autopsy under controlled experimental conditions.Materials and methods. Balb/c mice were euthanized and subsequently subjected to necropsy at 0, 3, 12, 24, 48, and 72 hours of the postmortem period. During the first three hours following euthanasia, the mice were maintained at room temperature, after which they were transferred to a refrigerator (4 º С). Total RNA was extracted from tissue samples taken from the kidney, liver, and brain; the integrity of the RNA samples was assessed by capillary electrophoresis, and the RNA quality number (RQN) was calculated. The expression levels of Actb, Epas1, and Rps18 housekeeping genes were evaluated by real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) with original primers and probes using the TaqMan assay. The histologic examination was performed according to standard techniques.Results. Degradation of RNA extracted from mouse kidney tissues appeared to be greater than that of RNA taken from the liver. In the meantime, a negative linear correlation was observed between RQN and the duration of the postmortem interval for liver and kidney samples. In contrast, no significant changes in the RQN score were observed for brain RNA samples at any of the time points. The expression of the Epas1 and Rps18 genes was significantly decreased in mouse kidney and liver tissues. However, the level of Epas1 and Rps18 gene expression in the brain remained stable at all time points and did not exhibit a significant decrease at 72 hours after euthanasia. No obvious morphological changes were detected by the histologic examination, which does not exclude the presence of ultrastructural pathological changes.Conclusion. RQN in autopsy tissues serves as a crucial predictor of sample quality for molecular biology studies, including gene expression analysis.

The article demonstrates a detailed analysis of the results of the rounds of quality control of immunohistochemical studies conducted by the Central Committee of the Immunohistochemical Quality Control Center of the Russian Medical Academy of Continuous Professional Education of the Ministry of Health of Russia in 2023. Typical shortcomings and errors in the immunohistochemical examination of various tumors have been identified and ways to eliminate them are given. Particular attention is paid to defining a panel of standard breast cancer markers and eliminating the shortcomings of immunohistochemical examination of markers of accompanying diagnosis.

Cardiac myxoma in its morphology is a typical benign tumor, meanwhile, the fact of its localization in the heart chamber, directly in the constant blood flow, largely determines the clinical behavior of this neoplasm, which is often manifested by the development of characteristics that formally determine the aggressive and even malignant nature of the course. Accordingly, the malignancy of cardiac myxoma is determined more by its clinical behavior (recurrence, multifocality of the lesion, the presence of mechanisms of spread similar to metastasis) rather than by its histological picture. In the structure of primary benign tumors of the heart, myxoma occupies a dominant position and its incidence is up to 85%. According to some authors, the tumor develops from multipotent mesenchymal stem cells of the endocardium, mainly in the area of the fossa ovale, while according to others the histogenesis of the tumor remains unclear. The obligate morphology element is the myxoma cell. The presence of so-called "ring" structures is special, regular and highly specific, and Gamna - Gandy bodies, foci of calcification and superficial thrombosis are considered characteristic secondary destructive morphological signs. The review describes the morphology features, specific clinical manifestations, immunohistochemical parameters of cardiac myxoma, and presents information available in the literature on the mechanisms of tumor spread (metastasis).

КЛИНИЧЕСКАЯ И ЭКСПЕРИМЕНТАЛЬНАЯ ХИРУРГИЯ■ СЕРДЕЧНАЯ И СОСУДИСТАЯ ХИРУРГИЯ Резюме Актуальность.Приблизительно у 20-30% пациентов с синдромом Нунан встречается асимметричная гипертрофическая кардиомиопатия (ГКМП) с обструкцией выходного отдела левого желудочка (ВОЛЖ) и митральной недостаточностью.Данных о роли трансаортальной расширенной септальной миоэктомии у детей с данным синдромом относительно мало.Цель -оценить результаты трансаортальной расширенной септальной миоэктомии у детей с обструктивной ГКМП и синдромом Нунан.Материал и методы.Проведен ретроспективный анализ результатов после трансаортальной расширенной септальной миоэктомии, выполненной в кардиохирургическом отделении № 2 НИИ кардиологии Томского НИМЦ с 2010 по 2023 г.Пациенты были разделены на 2 группы.В исследование были включены 40 пациентов.В I группу вошли 14 пациентов с ГКМП и синдромом Нунан, II группу составили 26 несиндромальных пациентов с ГКМП.Результаты.Медиана возраста составила 5,2 [2,3; 10,5] и 7,5 [3,4; 13,2] года для I и II групп (p=0,281).При этом возраст первичной постановки диагноза в I группе был 1 [1; 3], а во II группе -12 [6; 51] мес (р<0,001).Частота интраоперационных осложнений в I группе -7,1% (n=1), во II группе -23,1% (n=6).У всех пациентов I группы диагностировалась аномалия подклапанного аппарата митрального клапана, в 14,3% случаев для устранения обструкции потребовалось его протезирование механическим двустворчатым протезом.Летальности в раннем и среднесрочном послеоперационном периоде наблюдения не было.Медиана времени наблюдения за пациентами обеих групп составила 3,0 [2,9; 3,2] года.Значимые различия, по данным среднесрочных наблюдений, у пациентов I группы по сравнению со II группой наблюдались в остаточном градиенте на ВОЛЖ и z-индексе задней стенки левого желудочка (p=0,001; p=0,011).Заключение.Трансаортальная расширенная септальная миоэктомия позволяет эффективно устранить обструкцию ВОЛЖ у детей с синдромом Нунан и сопутствующей обструктивной ГКМП с удовлетворительными ранними и среднесрочными послеоперационными результатами.Финансирование.Исследование не имело спонсорской поддержки.

Most intracranial arachnoid cysts are thought to be non-tumorous, congenital, intra-arachnoid cerebrospinal fluid collections that account for about 1% of all intracranial space-occupying lesions. In children, the prevalence of this pathology is 2.6%; in adults, it reaches 1.4%. The disease is more often registered in men. Most often arachnoid cysts are supratentorial. Their most common locations are in the middle cranial fossa and the retrocerebellar cistern. Less often they can be detected on the convexity of the brain hemispheres; however, cases of arachnoid cysts at more unusual sites have also been described, including in newborns. The pathology is often characterized by an asymptomatic course, while certain symptoms may have an acute onset, which is due to compression of brain structures caused by the large cyst size.This article describes a clinical case of a large intracranial arachnoid cyst in a 28-year-old man. It was not verified in the antemortem diagnosis, but was revealed according to the autopsy findings (macroscopic features of the cyst, histologic presentation with specific morphological changes, and findings of computed tomography of the cerebral hemispheres).

Molecular genetic analysis of tumor tissues is the most important step towards understanding the mechanisms of cancer development; it is also necessary for the choice of targeted therapy. The Hi-C (high-throughput chromatin conformation capture) technology can be used to detect various types of genomic variants, including balanced chromosomal rearrangements, such as inversions and translocations. We propose a modification of the Hi-C method for the analysis of chromatin contacts in formalin-fixed paraffin-embedded (FFPE) sections of tumor tissues. The developed protocol allows to generate high-quality Hi-C data and detect all types of chromosomal rearrangements. We have analyzed various databases to compile a comprehensive list of translocations that hold clinical importance for the targeted therapy selection. The practical value of molecular genetic testing is its ability to influence the treatment strategies and to provide prognostic insights. Detecting specific chromosomal rearrangements can guide the choice of the targeted therapies, which is a critical aspect of personalized medicine in oncology.

OBJECTIVE:Study of the features of expression of immune checkpoint proteins PD-L1, CTLA4 and LAG3 in the microenvironment of colon adenocarcinoma depending on MMR status.MATERIAL AND METHODS:The study group consisted of 32 patients with a morphologically confirmed diagnosis of colon cancer; all of them underwent surgical treatment in the form of hemicolonectomy or resection. The work assessed samples of tumor tissue obtained as a result of surgery, the study was carried out in 3 stages: morphological examination of histological slides of colon tumors at the light-optical level, immunohistochemistry examination of tumor samples to determine the dMMR/pMMR status of carcinoma using a panel of antibodies to proteins of the unpaired nucleotide repair system MLH1, MSH2, MSH6 and PMS2, multiplex analysis of PD-L1, CTLA4, LAG3, CD3+, CD8+, CD163+ markers using the Vectra 3.0.3 tissue scanning system (Perkin Elmer, USA).RESULTS:Significant differences in the expression of PD-L1, CTLA4, LAG3 in the area of the invasive tumor margin were revealed between the dMMR and pMMR groups of colon adenocarcinomas in patients comparable in clinical and morphological characteristics and treatment. In the group of tumors with dMMR status, an increase in the expression of all studied markers was noted. The number of CD3+ TILs was also significantly higher in the invasive margin of tumors with dMMR status. Similarly, in this group of colon carcinomas, a large number of CD163+ macrophages were noted both in the center and in the invasive margin zone. No statistically significant differences were found in the expression of immune checkpoints and the composition of TILs in the central zone of tumors with different MMR status.CONCLUSION:A study using multiplex immunohistochemical analysis showed that MMR-deficient colon adenocarcinomas are characterized by more pronounced immune infiltration and increased expression of immune checkpoints in microenvironmental cells, mainly in the area of invasive tumor growth. The data obtained may be important for understanding the mechanisms of immune-mediated control of tumor growth and the choice of immunotherapy tactics depending on MMR status.