Integrin-associated transcriptional characteristics of circulating tumor cells in breast cancer patients.

Background: Integrins enable cell communication with the basal membrane and extracellular matrix, activating signaling pathways and facilitating intracellular changes. Integrins in circulating tumor cells (CTCs) play a significant role in apoptosis evasion and anchor-independent survival. However, the link between CTCs expressing different integrin subunits, their transcriptional profile and, therefore, their functional activity with respect to metastatic potential remains unclear. Methods: Single-cell RNA sequencing of CD45-negative cell fraction of breast cancer patients was performed. All CTCs were divided into nine groups according to their integrin profile. Results: СTCs without the gene expression of integrins or with the expression of non-complementary α and ß subunits that cannot form heterodimers prevailed. Only about 15% of CTCs expressed integrin subunits which can form heterodimers. The transcriptional profile of CTCs appeared to be associated with the spectrum of expressed integrins. The lowest potential activity was observed in CTCs without integrin expression, while the highest frequency of expression of tumor progression-related genes, namely genes of stemness, epithelial-mesenchymal transition (EMT), invasion, proinflammatory chemokines and cytokines as well as laminin subunits, were observed in CTCs co-expressing ITGA6 and ITGB4. Validation on the protein level revealed that the median of integrin ß4+ CTCs was higher in patients with more aggressive molecular subtypes as well as in metastatic breast cancer patients. One can expect that CTCs with ITGA6 and ITGB4 expression will have pronounced metastatic potencies manifesting in expression of EMT and stemness-related genes, as well as potential ability to produce chemokine/proinflammatory cytokines and laminins.

Spatial Heterogeneity of Integrins and Their Ligands in Primary Breast Tumors.

BACKGROUND: The diversity of cell-cell interactions in different regions of a tumor reflects the functional heterogeneity of cancer, which poses challenges in early diagnosis, selection of treatment strategies, and prognosis of breast cancer. Cancer cells interact with each other to form different morphological structures in the tumor and stromal host cells via integrins. The objective of this study was to characterize the morphological and spatial heterogeneity of primary breast tumors in the context of expression profiles of integrins and their ligands. METHODS: We studied spatial transcriptomics using the 10X Visium approach and the Niche Interactions and Communication Heterogeneity in Extracellular Signaling (NICHES) algorithm to map ligand-receptor signaling pathways and visualize the heterogeneity of signaling archetypes in tumor clusters. RESULTS: Cluster analysis of the expression profiles of tumor spots from the samples indicated pronounced inter-tumoral heterogeneity. Integrin-ligand functional clusters were associated with intratumoral heterogeneity, which was manifested by the presence of several morphological loci as observed in histological tumor samples. Inter-tumoral heterogeneity was manifested by a different number of functional clusters, ranging from 2 to 9 for each tumor sample. The main characteristic of these clusters was the significant predominance of non-complementary integrin subunits. Of the 42 functional integrin-ligand pairs in 21 clusters of five samples, 41 pairs occurred only once. The exception was the laminin subunit alpha-5 (LAMA5)-integrin beta 4 (ITGB4) pair, which was detected in two clusters of different samples. CONCLUSIONS: The spatial heterogeneity of integrin-ligand expression clusters in breast cancer contributes significantly to the functional heterogeneity of the tumor, which sets the stage for many scenarios of parenchymatous-stromal relationships, some of which may be effective in the emergence of metastasizing tumor seed cells. The intra- and inter-tumoral spatio-functional heterogeneity of the tumor tissue that we discovered may largely explain why it is difficult to achieve success in most patients with breast cancer using any therapeutic strategy targeting one molecule of the vast array, regardless of the importance of its pathogenetic significance.

Immunohistochemical Analysis of Lung Adenocarcinoma in Russian Mayak Nuclear Workers.

Specimens of lung adenocarcinoma (AdCa) from Russian nuclear workers (n = 54) exposed to alpha particles and gamma rays and from individuals non-exposed to radiation (n = 21) were examined using immunohistochemistry. Estimated significant associations with alpha dose were negative for Ki-67 and collagen IV in AdCa. Associations with gamma-ray dose were negative for tissue inhibitor of matrix metalloproteinase 2 and caspase 3 and positive for matrix metalloproteinase 2 and leukemia inhibiting factor in AdCa. The findings provide some evidence supporting alterations in apoptosis, cell proliferation and extracellular matrix in lung tissues affected by chronic radiation exposure that can contribute to radiogenic cancerogenesis.

Dose rate effect on mortality from ischemic heart disease in the cohort of Russian Mayak Production Association workers.

For improvement of the radiation protection system it is crucial to know the factors that modify the radiation dose-response relationship. One of such key factors is the ionizing radiation dose rate. There are, however, very few studies that examine the impact of the dose rate on radiogenic risks observed in human cohorts exposed to radiation at various dose rates. Here we investigated the impact of the dose rate (in terms of the recorded annual dose) on ischemic heart disease (IHD) mortality among Russian nuclear workers chronically exposed to radiation. We observed significantly increased excess relative risks (ERR) of IHD mortality per unit of external gamma-ray absorbed dose accumulated at higher dose rates (0.005-0.050 Gy/year). The present findings provide evidence for the association between radiation dose rate and ERRs of IHD mortality in occupationally chronically exposed workers per unit total dose. IHD mortality risk estimates considerably increased with increasing duration of uninterrupted radiation exposure at high rates. The present findings are consistent with other studies and can contribute to the scientific basis for recommendations on the radiation protection system.

The Association of Integrins ß3, ß4, and αVß5 on Exosomes, CTCs and Tumor Cells with Localization of Distant Metastasis in Breast Cancer Patients.

Integrins are cell adhesion receptors, which play a role in breast cancer invasion, angiogenesis, and metastasis. Moreover, it has been shown that exosomal integrins provide organotropic metastasis in a mouse model. In our study, we aimed to investigate the expression of integrins ß3, ß4, and αVß5 on exosomes and tumor cells (circulating tumor cells and primary tumor) and their association with the localization of distant metastasis. We confirmed the association of exosomal integrin ß4 with lung metastasis in breast cancer patients. However, we were unable to evaluate the role of integrin ß3 in brain metastasis due to the rarity of this localization. We established no association of exosomal integrin αVß5 with liver metastasis in our cohort of breast cancer patients. The further evaluation of ß3, ß4, and αVß5 integrin expression on CTCs revealed an association of integrin ß4 and αVß5 with liver, but not the lung metastases. Integrin ß4 in the primary tumor was associated with liver metastasis. Furthermore, an in-depth analysis of phenotypic characteristics of ß4+ tumor cells revealed a significantly increased proportion of E-cadherin+ and CD44+CD24- cells in patients with liver metastases compared to patients with lung or no distant metastases.

Heterogeneity of Circulating Epithelial Cells in Breast Cancer at Single-Cell Resolution: Identifying Tumor and Hybrid Cells.

Circulating tumor cells and hybrid cells formed by the fusion of tumor cells with normal cells are leading players in metastasis and have prognostic relevance. This study applies single-cell RNA sequencing to profile CD45-negative and CD45-positive circulating epithelial cells (CECs) in nonmetastatic breast cancer patients. CECs are represented by transcriptionally-distinct populations that include both aneuploid and diploid cells. CD45- CECs are predominantly aneuploid, but one population contained more diploid than aneuploid cells. CD45+ CECs mostly diploid: only two populations have aneuploid cells. Diploid CD45+ CECs annotated as different immune cells, surprisingly harbored many copy number aberrations, and positively correlated to tumor grade. It is noteworthy that cancer-associated signaling pathways areabundant only in one aneuploid CD45- CEC population, which may represent an aggressive subset of circulating tumor cells. Thus, CD45- and CD45+ CECs are highly heterogeneous in breast cancer patients and include aneuploid cells, which are most likely circulating tumor and hybrid cells, respectively, and diploid cells. DNA ploidy analysis can be an effective instrument for identifying tumor and hybrid cells among CECs. Further follow-up study is needed to determine which subsets of circulating tumor and hybrid cells contribute to breast cancer metastasis.

Incidence risks for subtypes of heart diseases in a Russian cohort of Mayak Production Association nuclear workers.

Heart diseases are one of the main causes of death. The incidence risks were assessed for various types of heart diseases (HDs) in a cohort of Russian nuclear workers of the Mayak Production Association (PA) who had been chronically occupationally exposed to external gamma and/ or internal alpha radiation. The study cohort included all workers (22,377 individuals) who had been hired at the Mayak PA during 1948-1982 and followed up until 31 December 2018. The mean gamma-absorbed dose to the liver (standard deviation) was 0.43 (0.63) Gy, and the mean alpha-absorbed dose to the liver was 0.25 (1.19) Gy. Excess relative risk (ERR) per unit liver-absorbed dose (Gy) was calculated based on maximum likelihood. At the end of the follow-up, 559 chronic rheumatic heart disease (CRHD), 7722 ischemic heart disease (IHD) [including 2185 acute myocardial infarction (AMI) and 3976 angina pectoris (AP)], 4939 heart failure (HF), and 3689 cardiac arrhythmia and conduction disorder (CACD) cases were verified in the study cohort. Linear model fits of the gamma dose response for HDs were best once adjustments for non-radiation factors (sex, attained age, calendar period, smoking status and alcohol consumption) and alpha dose were included. ERR/Gy in males and females was 0.17 (95% confidence intervals: 0.10, 0.26) and 0.23 (0.09, 0.38) for IHD; 0.18 (0.09, 0.29) and 0.26 (0.08, 0.49) for AP; - 0.01 (n/a, 0.1) and - 0.01 (n/a, 0.27) for AMI; 0.27 (0.16, 0.40) and 0.27 (0.10, 0.49) for HF; 0.32 (0.19, 0.46) and 0.05 (- 0.09, 0.22) for CACD; 0.73 (- 0.02, 2.40) and - 0.12 (- 0.50, 0.69) for CRHD, respectively. Sensitivity analyses demonstrated the persistence of a significant dose-response regardless of exclusion/inclusion of adjustments for known potential non-radiation confounders (smoking, alcohol consumption, body mass index, hypertension, diabetes mellitus), and it was only the magnitude of the risk estimate that varied. The risks of HD incidence were not modified with sex (except for the CACD risk). This study provides evidence for a significant association of certain types of HDs with cumulative dose of occupational chronic external exposure to gamma radiation.

The Novel Association of Early Apoptotic Circulating Tumor Cells with Treatment Outcomes in Breast Cancer Patients.

Stemness and epithelial-mesenchymal plasticity are widely studied in the circulating tumor cells of breast cancer patients because the roles of both processes in tumor progression are well established. An important property that should be taken into account is the ability of CTCs to disseminate, particularly the viability and apoptotic states of circulating tumor cells (CTCs). Recent data demonstrate that apoptosis reversal promotes the formation of stem-like tumor cells with pronounced potential for dissemination. Our study focused on the association between different apoptotic states of CTCs with short- and long-term treatment outcomes. We evaluated the association of viable CTCs, CTCs with early features of apoptosis, and end-stage apoptosis/necrosis CTCs with clinicopathological parameters of breast cancer patients. We found that the proportion of circulating tumor cells with features of early apoptosis is a perspective prognosticator of metastasis-free survival, which also correlates with the neoadjuvant chemotherapy response in breast cancer patients. Moreover, we establish that apoptotic CTCs are associated with the poor response to neoadjuvant chemotherapy, and metastasis-free survival expressed at least two stemness markers, CD44 and CD133.

Incidence risks for cerebrovascular diseases and types of stroke in a cohort of Mayak PA workers.

Incidence risks for cerebrovascular diseases (CeVD) and some types of stroke in a cohort of 22,377 Russian Mayak nuclear workers chronically exposed to ionising radiation and followed up until the end of 2018 are reported. Among total 9469 cases of CeVD, 2078 cases were strokes that included 262 hemorrhagic strokes (HS) and 1611 ischemic strokes (IS). Data evaluation was performed with categorical and dose-response analyses estimating the relative risk (RR) and excess relative risk (ERR) per unit cumulative liver absorbed dose of external gamma-ray or internal alpha-particle exposure based on a linear model utilizing the AMFIT module of the EPICURE software. CeVD incidence was found to be significantly associated with cumulative radiation dose: ERR/Gy was 0.37 (95% confidence interval (CI) 0.27, 0.47) in males and 0.47 (95% CI 0.31, 0.66) in females for external exposure, and 0.31 (95% CI 0.11, 0.59) in males and 0.32 (95% CI 0.11, 0.61) in females for internal exposure. When the model for the analysis of external radiation effect did not include an adjustment for alpha radiation dose (and vice versa), the radiogenic risk estimate increased notably both for males and for females. In contrast, exclusion from or inclusion in the model of additional adjustments for non-radiation factors did not notably change the risk estimates. ERR/Gy of external gamma dose for CeVD incidence significantly decreased with increasing attained age (males and females) and duration of employment (females). No significant associations of either stroke or its types with cumulative gamma-ray dose of external exposure or alpha-particle dose of internal exposure were found.

The Incidence Risk for Primary Glaucoma and Its Subtypes following Chronic Exposure to Ionizing Radiation in the Russian Cohort of Mayak Nuclear Workers.

Secondary glaucoma is a typical normal tissue complication following radiation therapy involving ocular radiation exposure at high fractionated dose (several tens of Gy). In contrast, recent studies in acutely exposed Japanese atomic bomb survivors showed a significantly increased risk for normal-tension glaucoma (NTG, a subtype of primary open-angle glaucoma) at much lower dose, but such information is not available in any other cohorts. We therefore set out to evaluate the incidence of risk for primary glaucoma and its subtypes in a Russian cohort of Mayak Production Association nuclear workers who received chronic radiation exposure over many years. Of these, we found a significantly increased relative risk (RR) of NTG incidence (RR = 1.88 95% confidence intervals (CI): 1.01, 3.51; p = 0.047) in workers exposed to gamma rays at cumulative brain absorbed dose above >1 Gy. We observed the linear relationship between NTG incidence and brain absorbed gamma dose with an excess relative risk per unit brain absorbed dose of 0.53 (95% CI: 0.01, 1.68; p < 0.05), but not for any other subtypes nor for total primary glaucoma. Such elevated risk of radiogenic NTG incidence, if confirmed in other cohorts, has significant implications for normal tissue complications in radiotherapy patients receiving ocular radiation exposure, and for ocular radiation protection in radiation workers.

Mortality from various diseases of the circulatory system in the Russian Mayak nuclear worker cohort: 1948-2018.

This paper reports on the findings from the study of mortality from diseases of the circulatory system (DCS) in Russian nuclear workers of the Mayak Production Association (22 377 individuals, 25.4% female) who were hired at the facility between 1948 and 1982 and followed up until the end of 2018. Using the AMFIT module of the EPICURE software, relative risks (RRs) and excess RRs per unit absorbed dose (ERR/Gy) for the entire Mayak cohort, the subcohort of workers who were residents of the dormitory town of Ozyorsk and the subcohort of migrants from Ozyorsk were calculated based on maximum likelihood. The mean cumulative liver absorbed gamma-ray dose from external exposure was 0.45 (0.65) Gy (mean (standard deviation)) for men and 0.37 (0.56) Gy for women. The mean cumulative liver absorbed alpha dose from internal exposure to incorporated plutonium was 0.18 (0.65) Gy for men and 0.40 (1.92) Gy for women. By the end of the follow-up, 6019 deaths with DCS as the main cause of death were registered among Mayak Production Association workers (including 3828 deaths in the subcohort of residents and 2191 deaths in the subcohort of migrants) over 890 132 (622 199/267 933) person-years of follow-up. The linear model that took into account non-radiation factors (sex, attained age, calendar period, smoking status and alcohol drinking status) and alpha radiation dose (via adjusting) did not demonstrate significant associations of mortality from DCS, ischaemic heart disease (IHD) and cerebrovascular disease with gamma-ray exposure dose in the entire cohort, the resident subcohort or the migrant subcohort (either in men or women). For the subcohort of residents, a significant association with gamma dose was observed for mortality from ischaemic stroke in men with ERR/Gy = 0.43 (95% CI 0.08; 0.99); there were no significant associations with liver absorbed gamma dose for any other considered outcomes. As for internal exposure, for men no significant associations of mortality from any DCS with liver absorbed alpha dose were observed, but for women positive associations were found for mortality from DCS (the entire cohort and the resident subcohort) and IHD (the entire cohort). No significant associations of mortality from various types of DCS with neutron dose were observed either in men or women, although neutron absorbed doses were recorded in only 18% of the workers.

PFKFB3 overexpression in monocytes of patients with colon but not rectal cancer programs pro-tumor macrophages and is indicative for higher risk of tumor relapse.

Introduction: Circulating monocytes are main source for tumor-associated macrophages (TAMs) that control tumor growth, angiogenesis, metastasis and therapy resistance. We raised the questions how monocyte programming is affected by growing tumors localized in colon and rectal sections, and how treatment onsets affect monocyte programming in the circulation. Methods: Patients with rectal cancer and colon cancer were enrolled in the study. Peripheral blood monocytes were characterized by phenotypic analysis using flow cytometry, by transcriptomic analysis using RNA sequencing and by gene expression analysis using real-time RT-PCR. Phenotypic analysis was performed with IF/confocal microscopy. Spatial transcriptomic analysis was applied using GeoMX DSP-NGS. Results: In patients with rectal cancer, increased amount of CCR2+ monocytes was indicative for the absence of both lymphatic and hematogenous metastasis. In contrast, in patients with colon cancer CD163+ monocytes were indicative for LN metastasis. NGS analysis identified tumor-specific transcriptional programming of monocytes in all CRC patients compared to healthy individuals. The key transcriptional difference between monocytes of patients with colon and rectal cancer was increased expression of PFKFB3, activator of glycolysis that is currently considered as therapy target for major solid cancers. PFKFB3-expressing monocyte-derived macrophages massively infiltrated tumor in colon. Nanostring technology identified correlation of PFKFB3 with amount and tumor-promoting properties of TAMs in colon but not in rectal cancer. PFKFB3 was indicative for tumor relapse specifically in colon cancer. Discussion: Our findings provide essential argument towards CRC definition to cover two clinically distinct cancers - colon cancer and rectal cancer, that differentially interact with innate immunity.

Heterogeneous Manifestations of Epithelial-Mesenchymal Plasticity of Circulating Tumor Cells in Breast Cancer Patients.

To date, there is indisputable evidence of significant CTC heterogeneity in carcinomas, in particular breast cancer. The heterogeneity of CTCs is manifested in the key characteristics of tumor cells related to metastatic progression - stemness and epithelial-mesenchymal (EMT) plasticity. It is still not clear what markers can characterize the phenomenon of EMT plasticity in the range from epithelial to mesenchymal phenotypes. In this article we examine the manifestations of EMT plasticity in the CTCs in breast cancer. The prospective study included 39 patients with invasive carcinoma of no special type. CTC phenotypes were determined by flow cytometry before any type of treatment. EMT features of CTC were assessed using antibodies against CD45, CD326 (EpCam), CD325 (N-cadherin), CK7, Snail, and Vimentin. Circulating tumor cells in breast cancer are characterized by pronounced heterogeneity of EMT manifestations. The results of the study indicate that the majority of heterogeneous CTC phenotypes (22 out of 24 detectable) exhibit epithelial-mesenchymal plasticity. The variability of EMT manifestations does not prevent intravasation. Co-expression of EpCAM and CK7, regardless of the variant of co-expression of Snail, N-cadherin, and Vimentin, are associated with a low number of CTCs. Intrapersonal heterogeneity is manifested by the detection of several CTC phenotypes in each patient. Interpersonal heterogeneity is manifested by various combinations of CTC phenotypes in patients (from 1 to 17 phenotypes).

Risk of skin cancer by histological type in a cohort of workers chronically exposed to ionizing radiation.

The incidence risk of non-melanoma skin cancer (NMSC), in particular basal cell carcinoma (BCC) and squamous cell carcinoma (SCC), was investigated in a cohort of workers of the Russian nuclear facility, the Mayak Production Association (PA), who had been occupationally exposed to low dose-rate ionizing radiation over prolonged periods. The study cohort included all workers who had been hired at the enterprise in 1948-1982 and followed up to 31.12.2018 (22,377 individuals, 25% of females). The mean cumulative skin absorbed dose of external gamma-ray exposure was 0.50 ± 0.73 Gy (the range of 0.00-8.84 Gy); the mean cumulative skin absorbed dose of neutron exposure was 0.002 ± 0.004 Gy (the range of 0.0000002-0.153 Gy). Relative risk and excess relative risk per unit skin absorbed dose of external exposure (RR and ERR/Gy) were estimated using AMFIT module of EPICURE software. Over the entire follow-up period 295 (84.8%) BCC, 48 (13.8%) SCC and 5 (1.4%) skin appendage cell carcinomas (SACC) were registered among NMSC in members of the study cohort. A significant linear association of the BCC incidence with the cumulative skin absorbed dose of external gamma-ray exposure was observed: ERR/Gy = 0.57 (95% CI 0.24, 1.06). Inclusion of an adjustment for neutron dose in the model resulted in a modest reduction of the BCC risk estimate [ERR/Gy = 0.55 (95% CI 0.23, 1.03)]. No significant association was revealed for SCC incidence with cumulative skin absorbed dose of external gamma-ray exposure [ERR/Gy = 0.14 (95% CI - 0.23, 0.91)]; inclusion of the neutron dose adjustment in the model did not modify the estimated SCC risk. No modification of the BCC and SCC incidence risks by sex, age at hire, attained age and facility type was observed.

Database of Families of Workers Chronically Exposed to Radiation: Data and Biospecimen Resources.

ABSTRACT: Animal experiment findings suggest that high doses of ionizing radiation exposure (>1.0 Gy) may cause genetic and epigenetic effects in offspring. However, epidemiological studies of offspring of radiation-exposed parents did not find increased risks of any health effects. Findings of cellular/experimental investigations and studies of human health effects are contradicting, and further investigations are needed to help resolve ambiguities using updated and/or improved data. This paper provides a detailed description of a database of families of workers of the first Russian nuclear facility, Mayak Production Association, located in the Southern Urals in the Chelyabinsk region close to Ozyorsk city, which started its operation in 1948 and today consists of reactors, radiochemical and plutonium production plants, and auxiliary facilities. The Mayak worker cohort includes 22,377 individuals (25% females) who were hired at one of the main Mayak PA facilities between 1948 and 1982 and were externally or internally exposed to ionizing radiation over prolonged periods. Advantages of the cohort include its large size, extensive follow-up period (70 y), individually measured doses from external and internal exposure and the wide range of these doses, heterogeneity by gender/age/ethnicity/initial health status, complete data on vital status and causes of death, available medical information on morbidity and reproduction, available data on non-radiation factors, and stored biological specimens donated by more than one-third of the cohort members. Based on medical and dosimetry database "Clinics" containing raw data on workers of the study cohort, the Mayak workers' family and offspring database was created. To date, it comprises 12,195 family couples (a husband and a wife) and 16,585 offspring. Biological specimens are available for more than 1,000 family triads (a husband, a wife, and their child). Stages of assembling the database and its descriptive characteristics are presented in this paper. Examples of potential applications of the database for investigations of non-targeted and transgenerational radiation effects in offspring of exposed parents are discussed.

Effect of Early-Stage Human Breast Carcinoma on Monocyte Programming.

Circulating monocytes are a major source of tumor-associated macrophages (TAMs). TAMs in human breast cancer (BC) support primary tumor growth and metastasis. Neoadjuvant chemotherapy (NAC) is a commonly used treatment for BC patients. The absence of the response to NAC has major negative consequences for the patient: increase of tumor mass, delayed surgery, and unnecessary toxicity. We aimed to identify the effect of BC on the subpopulation content and transcriptome of circulating monocytes. We examined how monocyte phenotypes correlate with the response to NAC. The percentage of CD14-, CD16-, CD163-, and HLA-DR-expressing monocytes was quantified by flow cytometry for patients with T1-4N0-3M0 before NAC. The clinical efficacy of NAC was assessed by RECIST criteria of RECIST 1.1 and by the pathological complete response (pCR). The percentage of CD14+ and СD16+ monocytes did not differ between healthy women and BC patients and did not differ between NAC responders and non-responders. The percentage of CD163-expressing CD14lowCD16+ and CD14+CD16+ monocytes was increased in BC patients compared to healthy women (99.08% vs. 60.00%, p = 0.039, and 98.08% vs. 86.96%, p = 0.046, respectively). Quantitative immunohistology and confocal microscopy demonstrated that increased levels of CD163+ monocytes are recruited in the tumor after NAC. The percentage of CD14lowCD16+ in the total monocyte population positively correlated with the response to NAC assessed by pCR: 8.3% patients with pCR versus 2.5% without pCR (p = 0.018). Search for the specific monocyte surface markers correlating with NAC response evaluated by RECIST 1.1 revealed that patients with no response to NAC had a significantly lower amount of CD14lowCD16+HLA-DR+ cells compared to the patients with clinical response to NAC (55.12% vs. 84.62%, p = 0.005). NGS identified significant changes in the whole transcriptome of monocytes of BC patients. Regulators of inflammation and monocyte migration were upregulated, and genes responsible for the chromatin remodeling were suppressed in monocyte BC patients. In summary, our study demonstrated that presence of BC before distant metastasis is detectable, significantly effects on both monocyte phenotype and transcriptome. The most striking surface markers were CD163 for the presence of BC, and HLA-DR (CD14lowCD16+HLA-DR+) for the response to NAC.

Heterogeneity of Stemlike Circulating Tumor Cells in Invasive Breast Cancer.

The presence of stem and epithelial-mesenchymal-transition (EMT) features in circulating tumor cells (CTCs) determines their invasiveness, adaptability to the microenvironment, and resistance to proapoptotic signals and chemotherapy. It also allows them to fulfil the role of metastatic "seeds". We evaluated the heterogeneity of stem CTCs by their CD44, ALDH1, and CD133 expression depending on N-cadherin expression in breast-cancer patients. A total of 38 female patients were selected for this study. CTC phenotypes were determined by flow cytometry before any type of treatment. Multiplex immunofluorescence was used for the evaluation of tumor-cell heterogeneity in primary lesions. In patients who had CD44-CD24- CTCs, a subset of cells with the expression of other stem-cell markers (CD133 and ALDH1) were detected. Expression of CD133 and/or ALDH1 may be associated with expression of N-cadherin: all populations of N-cadherin+ CTCs demonstrate stem features; in the absence of N-cadherin expression, true nonstem (CD44-CD24-CD133-ALDH1-) cells are found. The heterogeneity of stem marker expression in CTCs was observed regardless of N-cadherin expression. In our study, stromal cell-derived factor-1 (SDF-1) receptor expression in CTCs did not depend on stemlike traits, but was instead associated with N-cadherin expression. Subpopulations of tumor cells, detected both in tumors and blood, were identified. Breast cancer was characterized by pronounced interpersonal and intrapersonal heterogeneity of CTCs by the presence and combination of various stem features and N-cadherin expression. To complete the characterization of stemlike features of CTCs, we suggest the simultaneous use of the three stem markers.

Risk of stomach cancer incidence in a cohort of Mayak PA workers occupationally exposed to ionizing radiation.

Stomach cancer is a widespread health condition associated with environmental and genetic factors. Contribution of ionizing radiation to stomach cancer etiology is not sufficiently studied. This study was aimed to assess an association of the stomach cancer incidence risk with doses from occupational radiation exposure in a cohort of workers hired at main Mayak production association facilities in 1948-1982 taking into account non-radiation factors including digestive disorders. The study cohort comprised 22,377 individuals and by 31.12.2013 343 stomach cancer diagnoses had been reported among the cohort members. Occupational stomach absorbed doses were provided by the Mayak Worker Dosimetry System- 2008 (MWDS-2008) for external gamma ray exposure and by the Mayak Worker Dosimetry System- 2013 (MWDS-2013) for internal exposure to plutonium. Excess relative risks (ERR) per Gy for stomach cancer were estimated using the Poisson's regression. Analyses were run using the AMFIT module of the EPICURE software. The stomach cancer incidence risk in the study cohort was found to be significantly associated with the stomach absorbed dose of gamma rays: ERR/Gy = 0.19 (95% CI: 0.01, 0.44) with a 0 year lag, and ERR/Gy = 0.20 (95% CI: 0.01, 0.45) with a 5 year lag. To estimate the baseline risk, sex, attained age, smoking status and alcohol consumption, chronic diseases (peptic ulcer, gastritis and duodenitis) were taken into account. No modifications of the radiogenic risk by non-radiation factors were found in the study worker cohort. No association of the stomach cancer incidence risk with internal exposure to incorporated plutonium was observed.

Morphological features of pulmonary fibrosis in workers occupationally exposed to alpha radiation.

Purpose: The article reports on a comparative analysis of biological specimens of lung tissues collected from workers with pulmonary fibrosis induced by internal exposure to plutonium alpha-particles (plutonium-induced pulmonary fibrosis [PuPF]) and with etiologically different pulmonary fibrosis (non-PuPF) that developed as an outcome of a chronic obstructive pulmonary disease (COPD).Materials and methods: To perform histological examinations, lung tissues were sampled during autopsy. Six samples of various lung regions (the apical region, the lingula of the left lung and the inferior lobe) were collected from each donor. The resected tissue samples were fixed in 10% neutral-buffered formalin during 24 h and embedded into paraffin blocks (FFPE). FFPE blocks with lung tissue specimens collected from 56 workers with PuPF, 34 workers with non-PuPF and 35 workers without any lung disease were used in the study. To perform microscopic examination, lung tissue specimens were hematoxylin and eosin stained. To examine the connective-tissue scaffold of lung stroma and identify foci of pulmonary fibrosis, the cut sections of paraffin blocks were stained by Van Gizon's method (to assess the total volume of fibrosis-affected tissues), Gomori's technique (to define the reticular scaffold of lung stroma) and Weigert's technique (to examine elastic fibers). Morphological patterns of all biological specimens were studied using immunohistochemistry. To fit the empirical data, the Weibull's model was used.Results and conclusions: The study found qualitative and quantitative morphological features specific for PuPF compared to non-PuPF. The study demonstrated that hyper-production of collagen type V plays a key role in PuPF. The collagen type V content in fibrotic foci in lung tissue specimens from workers with PuPF was found to be increased.

The Russian Radiobiological Human Tissue Repository: characteristics of biological specimens donated by nuclear workers with lung cancer.

Purpose: Characteristics of biological specimens donated by nuclear workers with lung cancer.Materials and methods: Biological specimens were identified at the Radiobiological Human Tissue Repository (RHTR). It was established at the Southern Urals Biophysics Institute in Russia and has been developed and supported within the bilateral US-Russian Agreement on International Cooperation for Minimization of the Effects of Prolonged Radiation Exposure. Biological specimens were collected from workers of the Russian nuclear production facility Mayak PA who were exposed to gamma radiation and/or alpha particles. To determine a histologic type of lung cancer, immunohistochemistry was used.Results and conclusions: Today biological specimens donated by 343 registrants with lung cancer are available at the RHTR. Among them, 255 donors (74%) are Mayak PA workers hired at the main facilities (reactors, plutonium production, and radiochemical plants) in 1948-1982. These workers donated about 6024 specimens of lung tissues (tumor and tumor-free) stored mostly as formalin-fixed paraffin-embedded tissue blocks (31%) and histology slides (64%); in addition, they donated 1800 specimens of blood/blood components, buccal epithelium cells, and sputum. Among histologic types identified for these lung cancer cases, adenocarcinoma and small cell carcinoma were prevalent. Information about individual doses from external and internal radiation exposure, data on quantitative smoking characteristics and diseases are available for all workers with lung cancer. Complete information on radiation exposure, health status and non-radiation factors annotated to RHTR registrants and the high quality of the available biological specimens are a unique resource for studying biological mechanisms of radiation-induced lung cancer.