EpCAM-CD24+ circulating cells associated with poor prognosis in breast cancer patients.

Following the discovery of circulating tumor cells (CTCs) in the peripheral blood of cancer patients, CTCs were initially postulated to hold promise as a valuable prognostic tool through liquid biopsy. However, a decade and a half of accumulated data have revealed significant complexities in the investigation of CTCs. A challenging aspect lies in the reduced expression or complete loss of key epithelial markers during the epithelial-mesenchymal transition (EMT). This likely hampers the identification of a pathogenetically significant subset of CTCs. Nevertheless, there is a growing body of evidence regarding the prognostic value of such molecules as CD24 expressing in the primary breast tumor. Herewith, the exact relevance of CD24 expression on CTCs remains unclear. We used two epithelial markers (EpCAM and cytokeratin 7/8) to assess the count of CTCs in 57 breast cancer patients, both with (M0mts) and without metastasis (M0) during the follow-up period, as well as in M1 breast cancer patients. However, the investigation of these epithelial markers proved ineffective in identifying cell population expressing different combinations of EpCAM and cytokeratin 7/8 with prognostic significance for breast cancer metastases. Surprisingly, we found CD24+ circulating cells (CCs) in peripheral blood of breast cancer patients which have no epithelial markers (EpCAM and cytokeratin 7/8) but was strongly associated with distant metastasis. Namely, the count of CD45-EpCAM-CK7/8-CD24+ N-cadherin-CCs was elevated in both groups of patients, those with existing metastasis and those who developed metastases during the follow-up period. Simultaneously, an elevation in these cell counts beyond the established threshold of 218.3 cells per 1 mL of blood in patients prior to any treatment predicted a 12-fold risk of metastases, along with a threefold decrease in distant metastasis-free survival over a 90-month follow-up period. The origin of CD45-EpCAM-CK7/8-CD24+ N-cadherin-CCs remains unclear. In our opinion their existence can be explained by two most probable hypotheses. These cells could exhibit a terminal EMT phenotype, or it might be immature cells originating from the bone marrow. Nonetheless, if this hypothesis holds true, it's worth noting that the mentioned CCs do not align with any of the recognized stages of monocyte or neutrophil maturation, primarily due to the presence of CD45 expression in the myeloid cells. The results suggest the presence in the peripheral blood of patients with metastasis (both during the follow-up period and prior to inclusion in the study) of a cell population with a currently unspecified origin, possibly arising from both myeloid and tumor sources, as confirmed by the presence of aneuploidy.

Molecular subtype conversion in CTCs as indicator of treatment adequacy associated with metastasis-free survival in breast cancer.

Molecular subtype of breast cancer has a great clinical significance and used as one of the major criteria for therapeutic strategy. Recently, for anticancer therapy, the trend for oncologists is the predominant determination of biomarkers in the existing foci of the disease. In the case of adjuvant therapy prescribed for distant metastases prevention, CTCs could be a suitable object for investigation. CTCs as one of the factors responsible for tumor metastatic potential could be more convenient and informative for evaluation of hormone receptors, Ki-67 and HER2 expression, which are determine molecular subtype in breast cancer patient. In our study, we aimed to investigate the molecular subtype discordance between the primary tumor and CTCs in breast cancer patients. We established conversion of molecular subtype in most of the cases. Namely, conversion was detected in 90% of untreated patients and in 82% of breast cancer patients treated by neoadjuvant chemotherapy. At the same time, molecular subtype conversions in patients treated by neoadjuvant chemotherapy were more diverse. Molecular subtype conversions resulted more often in the unfavorable variants in circulating tumor cells. We stratified all patients according to the adequacy of treatment against converted CTCs molecular subtype. Our study revealed that good response to neoadjuvant chemotherapy observed in case of adequate therapy, namely, when chemotherapy scheme was sufficient against CTCs. It turned out that patients with inadequate therapy were characterized by decreased simulated 5-year metastasis-free survival compared to patients who received appropriate therapy. Thus, detection of molecular subtype conversion in circulating tumor cells could be a perspective tool for optimization of antitumor therapy.

Do tumor exosome integrins alone determine organotropic metastasis?

Metastasis is the most life-threatening event in cancer patients, so the key strategy to treat cancer should be preventing tumor spread. Predicting the site of probable hematogenous metastasis is important for determining the therapeutic algorithm that could prevent the spread of tumor cells. Certain hopes for solving this problem appeared owing to study showing the association between specific integrins on tumor exosomes surface and the site of future metastasis. Numerous experimental data indicate the ability of exosomes to transfer various phlogogenic factors to the target organ, which can lead to the formation of inflammatory foci. Studies of T-lymphocytes homing show that expression of various adhesion molecules including ligands for integrins highly increases on the endothelium during inflammation. Such a mechanism underlies not only in leukocyte transvasation, but, apparently, in the accumulation of bone marrow precursor cells and the formation of a premetastatic niche. This review summarizes the most significant data on the role exosomes to induce inflammation, which leads to the recruiting of bone marrow precursors and the establishment of premetastatic niches.

Assessment of nivolumab exposure and clinical safety of 480 mg every 4 weeks flat-dosing schedule in patients with cancer.

Background: A nivolumab monotherapy flat-dosing regimen of 480 mg every 4 weeks (Q4W) has been approved in several markets, including the United States, Canada, and European Union, as an alternative dosing regimen for several indications. Approvals of this Q4W regimen were based on population pharmacokinetic (PK) analyses, established flat exposure-response relationships, and clinical safety. The objective of this study was to compare the PK exposure of 480 mg Q4W with 3 mg/kg every 2 weeks (Q2W) and 240 mg Q2W using modeling and simulation, and to evaluate clinical safety of the Q4W regimen. Patients and methods: Nivolumab PK exposure for the 480 mg Q4W schedule was simulated for 3817 patients across multiple tumor types and compared with those for the 3 mg/kg Q2W and 240 mg Q2W schedules. The safety profile of the Q4W schedule was assessed by analysis of clinical data from 61 patients who transitioned to nivolumab 480 mg Q4W from 3 mg/kg Q2W during four phase III clinical trials. Results: Compared with 3 mg/kg Q2W, nivolumab 480 mg Q4W produced similar time-averaged concentration, approximately 16% lower trough concentration, and 45% higher peak concentration at steady state. The peak concentration for 480 mg Q4W was significantly lower than that of 10 mg/kg Q2W, a dose previously shown to have an acceptable tolerability and safety profile. Treatment-related adverse events (TRAEs) that started after transitioning from 3 mg/kg Q2W to 480 mg Q4W were reported in 14.8% of patients, with 1.6% of patients reporting grades 3-4 TRAEs. Pooled safety data for these patients are consistent with those for the 3 mg/kg Q2W schedules, and no new safety signals were identified. Conclusions: The time-averaged steady-state exposure and safety profile of nivolumab 480 mg Q4W are consistent with that of 3 mg/kg Q2W across multiple tumor types. Nivolumab 480 mg Q4W represents a new dosing schedule option, and in addition to 240 mg Q2W, provides convenience and flexibility for patient care. Clinical trial numbers: NCT01721772, NCT01668784, NCT01673867, NCT01642004.

[Risk of death from circulatory diseases in a cohort of patients exposed to chronic radiation]. / Risk smerti ot boleznei sistemy krovoobrashcheniia v kogorte rabotnikov, podvergshikhsia khronicheskomu oblucheniiu.

AIM: To assess mortality from circulatory diseases (CD) in a cohort of workers exposed occupationally to chronic radiation in relation to external and internal exposure, by taking into account known non-radiation risk factors (RFs), such as smoking (including smoking index), alcohol consumption, hypertension, and body mass index. SUBJECTS AND METHODS: Mortality from CD (ICD-10: I00 - I99) was studied in a cohort of 22,377 nuclear power plant («Mayak¼ Production Association) workers exposed occupationally to chronic radiation. The study was based on the individual dose estimates of external and internal exposure taken from the new Mayak workers dosimetry system 2008 (MWDS-2008). The quantitative characteristics of smoking (smoking index) were used for the first time to assess the risk for CD in the cohort of workers exposed to chronic radiation. RESULTS: There was a statistically significant linear relationship between CD mortality and external gamma-dose after adjusting for the non-radiation RFs; the excess relative risk per unit dose (ERR/Gy) was 0.05 (95% confidence interval (CI): 0 to 0.11). Introducing an additional adjustment for internal alpha-dose resulted in a twofold increase in ERR/Gy=0.10 (95% CI: 0.02 to 0.21). There was a statistically significant increasing trend in CD mortality with the elevated absorbed dose from internal alpha-radiation in the liver (ERR/Gy=0.27; 95% CI: 0.12 to 0.48). However, ERR/Gy decreased and lost its statistical significance after adjusting for external gamma-dose. CONCLUSION: The results of this study are in good agreement with risk estimates obtained in the Japanese cohort of atomic bomb survivors and in the cohorts of occupationally exposed workers.

Chronic bronchitis incidence in the extended cohort of Mayak workers first employed during 1948-1982.

OBJECTIVES: This paper describes findings from the study of chronic bronchitis (CB) incidence after occupational exposure to ionising radiation among workers employed at Russian Mayak Production Association (PA) during 1948 and 1982 and followed up until 2008 based on 'Mayak Worker Dosimetry System 2008'. METHODS: Analyses were based on 2135 verified cases among 21 417 workers. Rate ratios (RR) were estimated by categorical analysis for non-radiation and radiation factors. Excess rate ratios per Gy (ERR/Gy) of external or internal exposures with adjustments via stratification on other factors were calculated. RESULTS: The interesting finding in relation to non-radiation factors was a sharp increase in the RR for CB incidence before 1960, which could be caused by a number of factors. Analyses restricted to the follow-up after 1960 revealed statistically significant associations of the CB incidence and external γ-ray radiation, ERR/Gy=0.14 (95% CI 0.02 to 0.28) having adjusted for sex, attained age, calendar period, plant, smoking status and lung α-particle dose, and internal α-particle radiation, ERR/Gy=1.14 (95% CI 0.41 to 2.18) having adjusted for sex, attained age, calendar period, plant, smoking status and lung γ-ray dose and ERR/Gy=1.19 (95% CI 0.32 to 2.53) having additionally adjusted for pre-employment occupational hazards and smoking index instead of smoking status. CONCLUSIONS: Analyses of CB incidence in the study cohort identified positive significant association with occupational exposure to radiation: however, there are no similar studies of CB incidence in relation to radiation in other cohorts to date with which a meaningful comparison of the results could be made.

Smoking-related DNA adducts as potential diagnostic markers of lung cancer: new perspectives.

In recent years, the new direction such as identification of informative circulating markers reflecting molecular genetic changes in the DNA of tumor cells was actively developed. Smoking-related DNA adducts are very promising research area, since they indicate high pathogenetic importance in the lung carcinogenesis and can be identified in biological samples with high accuracy and reliability using highly sensitive mass spectrometry methods (TOF/TOF, TOF/MS, MS/MS). The appearance of DNA adducts in blood or tissues is the result of the interaction of carcinogenic factors, such as tobacco constituents, and the body reaction which is determined by individual characteristics of metabolic and repair systems. So, DNA adducts may be considered as a cumulative mirror of heterogeneous response of different individuals to smoking carcinogens, which finally could determine the risk for lung cancer. This review is devoted to analysis of the role of DNA adducts in lung carcinogenesis in order to demonstrate their usefulness as cancer associated markers. Currently, there are some serious limitations impeding the widespread use of DNA adducts as cancer biomarkers, due to failure of standardization of mass spectrometry analysis in order to correctly measure the adduct level in each individual. However, it is known that all DNA adducts are immunogenic, their accumulation over some threshold concentration leads to the appearance of long-living autoantibodies. Thus, detection of an informative pattern of autoantibodies against DNA adducts using innovative multiplex ELISA immunoassay may be a promising approach to find lung cancer at an early stage in high-risk groups (smokers, manufacturing workers, urban dwellers).

Cerebrovascular diseases incidence and mortality in an extended Mayak Worker Cohort 1948-1982.

Incidence and mortality from cerebrovascular disease (CVD) [International Classification of Diseases 9th revision (ICD-9) codes: 430-438] was studied in a cohort of 22,377 workers first employed at the Mayak Production Association (Mayak PA) in 1948-1982 and followed up to the end of 2008. The cohort size was increased by 19% and follow-up extended by 3 years over the previous analysis. Radiation doses were estimated using an updated dosimetry system: Mayak Worker Dosimetry System 2008 (MWDS-2008). For the first time, in an analysis of this cohort, quantitative smoking data were used. Workers of the study cohort were exposed occupationally to prolonged external gamma rays and internal alpha particles. The mean (±standard deviation) total dose from external gamma rays was 0.54 ± 0.76 Gy (95% percentile 2.21 Gy) for males and 0.44 ± 0.65 Gy (95% percentile 1.87 Gy) for females. The mean plutonium body burden in the 31% of workers monitored for internal exposure was 1.32 ± 4.87 kBq (95% percentile 4.71 kBq) for males and 2.21 ± 13.24 kBq (95% percentile 4.56 kBq) for females. The mean total absorbed alpha-particles dose to the liver from incorporated plutonium was 0.23 ± 0.77 Gy (95% percentile 0.89 Gy) in males and 0.44 ± 2.11 Gy (95% percentile 1.25 Gy) in females. After adjusting for nonradiation factors (gender, age, calendar period, employment period, facility, smoking, alcohol consumption), there were significantly increasing trends in CVD incidence associated with total absorbed dose from external gamma rays and total absorbed dose to the liver from internal alpha-particle radiation exposure. Excess relative risks per Gy (ERR/Gy) were 0.46 (95% CI 0.37, 0.57) and 0.28 (95% CI 0.16, 0.42), respectively, based on a linear dose-response model. Adjustments for additional factors (hypertension, body mass index, duration of employment, smoking index and total absorbed dose to the liver from internal exposure during the analysis of external exposure and vice versa) had little effect on the results. The categorical analyses showed that CVD incidence was significantly higher among workers with total absorbed external gamma-ray doses greater than 0.1 Gy compared to those exposed to lower doses and that CVD incidence was also significantly higher among workers with total absorbed internal alpha-particle doses to the liver from incorporated plutonium greater than 0.01 Gy compared to those exposed to lower doses. The results of the categorical analyses of CVD incidence were in good agreement with a linear dose response for external gamma-ray doses but for internal alpha-particle doses the picture was less clear. For the first time an excess risk of CVD mortality was seen in workers whose livers were exposed to internal alpha-particle doses greater than 0.1 Gy compared to those workers who were exposed to doses of less than 0.01 Gy. A significant increasing trend for CVD mortality with internal alpha-particle dose was revealed in the subcohort of workers exposed at doses <1.0 Gy after having adjusted for nonradiation factors, ERR/Gy = 0.84 (95% CI, 0.09, 1.92). These updated results provide good evidence for a linear trend in risk of CVD incidence with external gamma-ray dose. The trend for CVD incidence with internal alpha-particle dose is less clear due to the impact of issues concerning the use of dose estimates based on below the limit of detection bioassay measurements.

Chronic bronchitis in the cohort of Mayak workers first employed 1948-1958.

Incidence of chronic bronchitis has been studied in a cohort of 12,210 workers first employed at one of the main plants of the Mayak nuclear facility during 1948-1958 and followed up to 31 December 2005. Information on external gamma doses is available for virtually all of these workers; in contrast, plutonium body burden was measured only for 30% of workers. During the follow-up period in the study cohort 1,175 incident cases of chronic bronchitis were verified. The analyses of nonradiation factors revealed that the underlying risk of chronic bronchitis incidence increased with increasing attained age and was higher among smokers compared with never-smokers as would be expected. The most interesting finding in relationship to nonradiation factors was a sharp increase in the baseline chronic bronchitis risk before 1960. The cause of this is not clear but a number of factors may play a role. Based on the follow-up data after 1960, the analysis showed a statistically significant linear dose response relationship with cumulative external gamma-ray dose (ERR/Gy = 0.14, 95% CI 0.01, 0.32). Based on the same subset but with an additional restriction to members with cumulative internal lung dose below 1 Gy, a statistically significant linear dose response relationship with internal alpha-radiation lung dose from incorporated plutonium was found (ERR/Gy = 2.70, 95% CI 1.20, 4.87). In both cases, adjustment was made for nonradiation factors, including smoking and either internal or external dose as appropriate. At present there are no similar incidence studies with which to compare results. However, the most recent data from the atomic bomb survivor cohort (the Life Span Study) showed statistically significant excess mortality risk for respiratory diseases of 22% per Gy and this value is within the confidence bounds of the point estimate of the risk from this study in relation to external dose.

Search for potential gastric cancer markers using miRNA databases and gene expression analysis.

AIM: The aim of this study was to identify genes that are differentially expressed in gastric tumors and to analyze the association of their expression level with tumor clinicopathologic features. METHODS: In the present research, we used bioinformatic-driven search to identify miRNA that are down-regulated in gastric tumors and to find their potential targets. Then, the expression levels of some of the target mRNAs were investigated using reverse transcription polymerase chain reaction (RT-PCR) analysis. RESULTS: As a result of the bioinformatics analysis, fifteen genes were found to be potentially differentially expressed between the tumors and normal gastric tissue. Five of them were chosen for the further analysis (WNT4, FGF12, EFEMP1, CTGF, and HSPG2) due to their important role in cell proliferation and differentiation. Expression levels of these genes were evaluated in our collection of frozen tissue samples of gastric tumor and paired normal stomach epithelia. Increased FGF12 expression was observed in diffuse type of gastric cancer while WNT4 mRNA was found to be down-regulated in intestinal type of gastric cancer. Besides, CTGF gene overexpression was revealed in diffuse type of stomach cancer in comparison with that in intestinal type. Up-regulation of CTGF was also associated with lymph node metastasis. CONCLUSIONS: The findings show its expedient to perform further investigations in order to clarify diagnostic and prognostic value of CTGF, FGF12, and WNT4's in stomach cancer as well as the role of these genes in carcinogenesis.

Modeling of respiratory system dysfunction among nuclear workers: a preliminary study.

Numerous studies have reported on cancers among Mayak Production Association (PA) nuclear workers. Other studies have reported on serious deterministic effects of large radiation doses for the same population. This study relates to deterministic effects (respiratory system dysfunction) in Mayak workers after relatively small chronic radiation doses (alpha plus gamma). Because cigarette smoke is a confounding factor, we also account for smoking effects. Here we present a new empirical mathematical model that was introduced for simultaneous assessment of radiation and cigarette-smoking-related damage to the respiratory system. The model incorporates absolute thresholds for smoking- and radiation-induced respiratory system dysfunction. As the alpha radiation dose to the lung increased from 0 to 4.36 Gy, respiratory function indices studied decreased, although remaining in the normal range. The data were consistent with the view that alpha radiation doses to the lung above a relatively small threshold (0.15 to 0.39 Gy) cause some respiratory system dysfunction. Respiratory function indices were not found to be influenced by total-body gamma radiation doses in the range 0-3.8 Gy when delivered at low rates over years. However, significant decreases in airway conductance were found to be associated with cigarette smoking. Whether the indicated cigarette smoking and alpha radiation associated dysfunction is debilitating is unclear.

Chromosomal aberrations in lymphocytes of peripheral blood among Mayak facility workers who inhaled insoluble forms of 239PU.

A cytogenetic study was performed on 79 plutonium (Pu) workers chronically exposed to alpha radiation from inhaled, low-transportable (insoluble) compounds of airborne 239Pu and to external gamma rays. Body burden estimates for 239Pu ranged from 0 to 15.5 kBq. Chromosomal aberrations (CAs) (stable and unstable) among peripheral blood lymphocytes and cumulative alpha radiation doses were evaluated approximately 25 y after first contact with 239Pu. For the cytogenetic analyses, a standard two-day peripheral blood lymphocyte culture technique was applied. While alpha radiation doses continually increase up to the time of cytogenetic measurements, significant gamma ray exposures essentially ceased long before the time of measurement, so that alpha and gamma doses were not correlated. For the exposed workers, the mean 239Pu body burden (estimate), evaluated at the time of the cytogenetic measurement, was 1.23 +/- 0.26 kBq and the corresponding mean absorbed external gamma ray dose (estimate) to the total body was 0.076 +/- 0.009 Gy. Single and multivariate regression analyses were performed on the CA data. Stable, unstable and total aberrations increased as the 239Pu body burden increased over the range 0-4.5 kBq. However, above this range little additional increase was observed. CAs were weakly correlated with time since the first intake of 239Pu. No relationship between chromatid aberrations and 239Pu incorporation was found. Unstable (but not stable) aberrations were correlated with gamma radiation dose. No significant relationship of CA and smoking was found.