Soloxolone methyl induces apoptosis and oxidative/ER stress in breast cancer cells and target cancer stem cell population.

One of the most prevalent malignancies in women and one of the leading causes of cancer-related death is breast cancer. There is a need for new treatment approaches and drugs for breast cancer. Many studies show the high potential of triterpene compounds and their semisynthetic derivatives as anticancer agents due to their ability to induce apoptosis and suppress tumorigenesis. The effects of soloxolone methyl (SM), a semisynthetic derivative of 18-H-glycyrrhetinic acid, on the cytotoxicity and apoptosis of human breast cancer cell line (T-47D) and cancer stem cell (CSCs) population (mammospheres; CD44+/CD24-antigen) derived from breast cancer cells, were examined in this work. The ATP assay was used to determine SM growth-inhibitory effects. Fluorescent staining, caspase-cleaved cytokeratin 18, and flow cytometry analysis were used to determine the mode of the cell death. In addition, cell death was investigated at protein and gene levels by Western Blotting and PCR, respectively. SM resulted in cytotoxicity in a time and dose dependent manner via ROS production and ER stress in T-47D cells in 2 models. The mode of cell death was apoptosis, evidenced by phosphatidylserine exposure, caspase activation, and bax overexpression. In mammospheres as 3D model, SM decreased stem cell properties and induced cell death. Taken together, SM may be a promising agent in the treatment of breast cancer, especially due to its antigrowth activity on CSCs.

Targeting the epithelial-mesenchymal transition (EMT) pathway with combination of Wnt inhibitor and chalcone complexes in lung cancer cells.

Non-small cell lung cancer (NSCLC) is the most common type of the lung cancer. Despite development in treatment options in NSCLC, the overall survival ratios is still poor due to epithelial and mesenchymal transition (EMT) feature and associated metastasis event. Thereby there is a need to develop strategy to increase antitumor response against the NSCLC cells by targeting EMT pathway with combination drugs. Niclosamide and chalcone complexes are both affect cancer cell signaling pathways and therefore inhibit the EMT pathway. In this study, it was aimed to increase antitumor response and suppress EMT pathway in NSCLC cells by combining niclosamide and chalcone complexes. SRB cell viability assay was performed to investigate the anticancer activity of drugs. The drugs were tested on both NSCLC cells (A549 and H1299) and normal lung bronchial cells (BEAS-2B). Then the two drugs were combined and their effects on cancer cells were evaluated. Fluorescence imaging and enzyme-linked immunosorbent assay were performed on treated cells to observe the cell death manner. Wound healing assay, real-time quantitative polymerase chain reaction, and western blot analysis were performed to measure EMT pathway activity. Our results showed that niclosamide and chalcone complexes combination kill cancer cells more than normal lung bronchial cells. Compared to single drug administration, the combination of both drugs killed NSCLC cells more effectively by increasing apoptotic activity. In addition, the combination of niclosamide and chalcone complexes decreased multidrug resistance and EMT activity by lowering their gene expressions and protein levels. These results showed that niclosamide and chalcone complexes combination could be a new drug combination for the treatment of NSCLC.

Sleep Problems in Individuals With Intestinal Stomas and Determining the Quality of Sleep: A Multicenter Study.

PURPOSE: The aim of this study was to determine sleep problems and the sleep quality of individuals with intestinal stomas. DESIGN: Descriptive quantitative design was used. SUBJECTS AND SETTING: The research was carried out from September 2018 to September 2019 at 3 university hospitals with stoma units located in Ankara, Turkey. The study sample comprised 222 individuals with intestinal ostomies who were being monitored in these 3 centers. Approximately half (N = 113, 50.9%) had an ileostomy and 68.5% (N = 152) had a temporary stoma. More than half of the participants (N = 116, 52.3%) reported current sleep problems, 59.5% (N = 132) reported that their stoma affected their sleep, and 64% (N = 142) had problems at night due to their stomas, reporting that they woke up frequently during their main sleep period due to ostomy-related problems. METHODS: Three instruments were used to measure study outcomes: the researcher-designed instrument determining the descriptive characteristics of individuals with intestinal stomas, the Pittsburgh Sleep Quality Index, and Epworth Sleepiness Scale. RESULTS: The mean Pittsburgh Sleep Quality Index score was found to be above 5 (13.42, SD: 3.01) indicating poor sleep quality. The mean Epworth Sleepiness Scale score was 7.54 (SD: 6.37) indicating that participants had daytime sleepiness. The mean Pittsburgh Sleep Quality Index scores of participants with ileostomies ( P = .002) and those with temporary stomas ( P = .009) were found to be significantly higher. CONCLUSIONS: Study findings indicate that individuals with intestinal stomas have poor sleep quality and problems with daytime sleepiness; those with ileostomies and temporary stomas had the worst quality of sleep.

Determination of thermal comfort among nurses working with personal protective equipment in COVID-19 clinics.

AIMS: This study aimed to determine thermal comfort among nurses working with personal protective equipment in COVID-19 clinic. METHODS: In this study, a descriptive design was carried out between June and September 2020. Sample of the study consisted of 246 nurses (77.6%) who worked in the COVID-19 clinics with personal protective equipment. We used a questionnaire to determine thermal comfort of nurses; a follow-up form to determine the factors affecting thermal comfort; and the ASHRAE Thermal Sensation Scale. Four measurements and follow-ups were made three times. RESULTS: More than half of nurses complained of ambient temperature and ventilation, one-third complained of humidity and nearly half complained of poor air quality. The mean thermal comfort score of nurses working in COVID-19 clinics was 1.19 (SD = 0.75). The thermal comfort of the nurses was negatively affected in all measurements except before wearing personal protective equipment. The highest scores were measured leaving the patient room and before removing personal protective equipment (M = 2.65, SD = 0.58). CONCLUSION: The thermal comfort, work performance and stress levels of the nurses were negatively affected by working with personal protective equipment. This study reveals the necessity of improving the working conditions of nurses, including working hours, environment and personal protective equipment.

Mitochondrial miRNAs (MitomiRs): Their potential roles in breast and other cancers.

Breast cancer is the most common cancer in women worldwide. MicroRNAs (miRNAs) are non-coding RNAs that are involved in the post-transcriptional regulation of gene expression. Although miRNAs mainly act in the cytoplasm, they can be found in the mitochondrial compartment of the cell. These miRNAs called "MitomiR", they can change mitochondrial functions by regulating proteins at the mitochondrial level and cause cancer. In this review, we have aimed to explain miRNA biogenesis, transport pathways to mitochondria, and summarize mitomiRs that have been shown to play an important role in mitochondrial function, especially in the initiation and progression of breast cancer.

Investigation of the efficacy of paclitaxel on some miRNAs profiles in breast cancer stem cells.

Understanding of the functions of microRNAs in breast cancer and breast cancer stem cells have been a hope for the development of new molecular targeted therapies. Here, it is aimed to investigate the differences in the expression levels of let-7a, miR-10b, miR-21, miR-125b, miR-145, miR-155, miR-200c, miR-221, miR-222 and miR-335, which associated with gene and proteins in MCF-7 (parental) and MCF-7s (Mammosphere/stem cell-enriched population/CD44+/CD24-cells) cells treated with paclitaxel. MCF-7s were obtained from parental MCF-7 cells. Cytotoxic activity of paclitaxel was determined by ATP assay. Total RNA isolation and cDNA conversion were performed from the samples. Changes in expression levels of miRNAs were examined by RT-qPCR. Identified target genes and proteins of miRNAs were analyzed with RT-qPCR and western blot analysis, respectively. miR-125b was significantly expressed (2.0946-fold; p = 0.021) in MCF-7s cells compared to control after treatment with paclitaxel. Downregulation of SMO, STAT3, NANOG, OCT4, SOX2, ERBB2 and ERBB3 and upregulation of TP53 genes were significant after 48 h treatment in MCF-7s cells. Protein expressions of SOX2, OCT4, SMAD4, SOX2 and OCT4 also decreased. Paclitaxel induces miR-125b expression in MCF-7s cells. Upregulation of miR-125b may be used as a biomarker for the prediction of response to paclitaxel treatment in breast cancer.

Combination of Histone Deacetylase Inhibitor with Cu(II) 5,5-diethylbarbiturate Complex Induces Apoptosis in Breast Cancer Stem Cells: A Promising Novel Approach.

BACKGROUND: Cancer Stem Cells (CSCs) are a subpopulation within the tumor that play a role in the initiation, progression, recurrence, resistance to drugs and metastasis of cancer. It is well known that epigenetic changes lead to tumor formation in cancer stem cells and show drug resistance. Epigenetic modulators and /or their combination with different agents have been used in cancer therapy. OBJECTIVE: In our study, we scope out the effects of a combination of a histone deacetylases inhibitor, Valproic Acid (VPA), and Cu(II) complex [Cu(barb-κN)(barb-κ2N,O)(phen-κN,N')]·H2O] on cytotoxicity/apoptosis in a stem-cell enriched population (MCF-7s) obtained from parental breast cancer cell line (MCF-7). METHODS: The viability of the cells was measured by the ATP assay. Apoptosis was elucidated via the assessment of caspase-cleaved cytokeratin 18 (M30 ELISA) and a group of flow cytometry analysis (caspase 3/7 activity, phosphatidylserine translocation by annexin V-FITC assay, DNA damage and oxidative stress) and 2',7'- dichlorofluorescein diacetate staining. RESULTS: The VPA combined with Cu(II) complex showed anti-proliferative activity on MCF-7s cells in a doseand time-dependent manner. Treatment with a combination of 2.5 mM VPA and 3.12 µM Cu(II) complex induced oxidative stress in a time-dependent manner, as well as apoptosis evidenced by the increase in caspase 3/7 activity, positive annexin-V-FITC, and increase in M30 levels. CONCLUSION: The results suggest that the combination therapy induces apoptosis following increased oxidative stress, thereby making it a possible promising therapeutic strategy for which further analysis is required.

Factors Affecting the Thermal Comfort of Perioperative Personnel in the OR.

Thermal comfort is an important component of any work setting and can be difficult to achieve in the complex OR environment. This comparative descriptive study sought to identify factors affecting the thermal comfort of perioperative personnel in the OR (N = 68). Researchers used the American Society of Heating, Refrigerating and Air-Conditioning Engineers Thermal Sensation Scale and researcher-developed surveys to collect data. The mean reported thermal comfort level was -0.44 (standard deviation 1.3), which is within acceptable parameters. Factors affecting thermal comfort were gender, professional role, and wearing additional clothing. Additionally, participants who complained about temperature or ventilation were less likely to report thermal comfort levels in the normal range. Health care facility leaders should consider the comfort needs of perioperative personnel when making purchasing decisions about ventilation systems and surgical clothing; however, they also should consider other factors (eg, surgeon comfort, permeability of protective clothing to blood and other body fluids).

Transient Contrast Neurotoxicity After Percutaneous Coronary Intervention Mimicking Subarachnoid Hemorrhage in a Patient With Chronic Kidney Disease.

Transient contrast neurotoxicity is a rare but well-recognized complication of angiography that is due to neurotoxicity of the contrast agent. Patients with renal dysfunction may be inclined to develop contrast medium neurotoxicity due to delayed elimination of the contrast medium in renal metabolism. In this report, we present an unusual case of transient neurotoxicity in a patient with severe chronic kidney disease following percutaneous coronary intervention mimicking clinically and radiologically subarachnoid hemorrhage. The patient's clinical symptoms improved rapidly and fully recovered after hemodialysis and conservative treatment. We believe that performing early hemodialysis is an effective treatment to improve symptoms in end-stage renal disease patients with contrast-induced encephalopathy.

Effects of pre-operative individualized education on anxiety and pain severity in patients following open-heart surgery.

OBJECTIVES: Individualized education is important for preparing patients for the operation both physically and psychologically. This study investigated the effects of pre-operative individualized education for open-heart surgery patients on post-operative anxiety and pain severity. METHODS: This quasi-experimental study was conducted in a university hospital between January and October 2014 and involved 109 patients. Data were collected through a form developed by the authors, the State-Trait Anxiety Inventory (STAI), and the visual analog scale (VAS). Patients underwent STAI 1 day before the operation to identify their sources of anxiety and educational needs, and then individualized education was given accordingly. 1 day after the operation, STAI was used to measure patients' state anxiety level, and VAS was used to measure their pain level. The effect of demographic variables on differences in anxiety and pain was investigated. RESULTS: The average age of the participants was 59.62; 69.7% were male, 92.7% were married, 49.5% graduated from primary school, and the majority (71.6%) had coronary heart disease. The most common sources of anxiety reported the participants included lack of knowledge, being away from family, risk of death, and pain. An analysis of the participants' pre-operative mean scores for state anxiety displayed statistically significant differences (P < 0.05) according to age groups and gender. No significant difference was detected between mean pre- and post-operative state anxiety scores. There was a statistically significant relationship between mean pre- and post-operative state anxiety scores and mean pain scores. CONCLUSIONS: The individualized education is given to patients before surgery was found to have potential effects on their post-operative pain levels.

Valproic acid, a histone deacetylase inhibitor, induces apoptosis in breast cancer stem cells.

Cancer stem-like cells (CSCs) are a cell subpopulation that can reinitiate tumors, resist chemotherapy, give rise to metastases and lead to disease relapse because of an acquired resistance to apoptosis. Especially, epigenetic alterations play a crucial role in the regulation of stemness and also have been implicated in the development of drug resistance. Hence, in the present study, we examined the cytotoxic and apoptotic activity of valproic acid (VPA) as an inhibitor of histone deacetylases (HDACs) against breast CSCs (BCSCs). Increased expression of stemness markers were determined by western blotting in mammospheres (MCF-7s, a cancer stem cell-enriched population) propagated from parental MCF-7 cells. Anti-growth activity of VPA was determined via ATP viability assay. The sphere formation assay (SFA) was performed to assess the inhibitory effect of VPA on the self-renewal capacity of MCF-7s cells. Acetylation of histon H3 was detected with ELISA assay. Cell death mode was performed by Hoechst dye 33342 and propidium iodide-based flouresent stainings (for pyknosis and membrane integrity), by M30 and M65 ELISA assays (for apoptosis and primary or secondary necrosis) as well as cytofluorimetric analysis (caspase 3/7 activity and annexin-V-FITC staining for early and late stage apoptosis). VPA exhibited anti-growth effect against both MCF-7 and MCF-7s cells in a dose (0.6-20 mM) and time (24, 48, 72 h) dependent manner. As expected, MCF-7s cells were found more resistant to VPA than MCF-7 cells. It was observed that VPA prevented mammosphere formation at relatively lower doses (2.5 and 5 mM) while the acetylation of histon H3 was increased. At the same doses, VPA increased the M30 levels, annexin-V-FITC positivity and caspase 3/7 activation, implying the induction of apoptosis. The secondary necrosis (late stage of apoptosis) was also evidenced by nuclear pyknosis with propidium iodide staining positivity. Taken together, inhibition of HDACs is cytotoxic to BCSCs by apoptosis. Our results suggested that targeting the epigenetic regulation of histones may be a novel approach and hold significant promise for successful treatment of breast cancer.

Circulating miR-195 as a Therapeutic Biomarker in Turkish Breast Cancer Patients.

BACKGROUND: Dysregulation of miRNA expression may be used as a biomarker for specific tumours because it may contribute to development of cancer. Circulating miRNA profiles have been highlighted for their potential as predictive markers in heterogeneous diseases such as breast cancer. In the literature, there is evidence that miR-195 levels are differentially expressed pre- and post-operative periods in breast cancer patients. At the same time, miRNA expression levels may vary because of ethnic origins. This study aimed to determine expression levels and potential roles of miR-195 in Turkish breast cancer patients. MATERIALS AND METHODS: The expression patterns of miR-195 were initially examined in breast cancer tissues (luminal A and B type) (n=96). Subsequently, blood samples were prospectively collected from preoperative and postoperative Turkish breast cancer patients and disease free controls. Total RNA was isolated, and the expression level of miR-195 was quantified by real-time PCR. RESULTS: We found that miR-195 level was altered in Turkish breast cancer patients, with down-regulation evident in breast cancer tissues compared to normal adjacent specimens. Furthermore, circulating levels of miR- 195 was significantly decreased in post-operative blood samples compared with pre-operative levels (p=0.01 and <0.05). However, miR-195 was significantly increased in pre-operative blood samples of the luminal B type (p= 0.04 and <0.05). CONCLUSIONS: This study represents the first report of a miR-195 expression profile in Turkish breast cancer patients. Our data suggests that miR-195 levels might be a clinically useful biomarker in the earliest stage of Turkish breast cancer patients.

BRCA mutations cause reduction in miR-200c expression in triple negative breast cancer.

Triple negative breast cancer (TNBC) is the most aggressive and poorly understood subclass of breast cancer (BC). Over the recent years, miRNA expression studies have been providing certain detailed overview that aberrant expression of miRNAs is associated with TNBC. Although TNBC tumors are strongly connected with loss of function of BRCA genes, there is no knowledge about the effect of BRCA mutation status on miRNA expressions in TNBC cases. The aims of this study were to evaluate the expression profile of miRNAs that plays role in TNBC progression and the role of BRCA mutations in their regulation. The expression level of BC associated 13 miRNAs was analyzed in 7 BRCA mutations positive, 6 BRCA mutations negative TNBC cases and 20 non-tumoral tissues using RT-PCR. According to RT2 Profiler PCR Array Data Analysis, let-7a expression was 4.67 fold reduced in TNBCs as compared to normal tissues (P=0.031). In addition, miR-200c expression was 5.75 fold reduced in BRCA mutation positive TNBC tumors (P=0.005). Analysis revealed a negative correlation between miR-200c and VEGFA expressions (r=-468). Thus, miR-200c may be involved in invasion and metastasis in TNBC cases with BRCA mutation. In this study we provide the knowledge on the first report of association between microRNA-200c and BRCA mutations in TNBC. Further studies and evaluations are required, but this miRNA may provide novel therapeutic molecular targets for TNBC treatment and new directions for the development of anticancer drugs.

Evaluation of genetic variations in miRNA-binding sites of BRCA1 and BRCA2 genes as risk factors for the development of early-onset and/or familial breast cancer.

Although genetic markers identifying women at an increased risk of developing breast cancer exist, the majority of inherited risk factors remain elusive. Mutations in the BRCA1/BRCA2 gene confer a substantial increase in breast cancer risk, yet routine clinical genetic screening is limited to the coding regions and intron- exon boundaries, precluding the identification of mutations in noncoding and untranslated regions. Because 3' untranslated region (3'UTR) polymorphisms disrupting microRNA (miRNA) binding can be functional and can act as genetic markers of cancer risk, we aimed to determine genetic variation in the 3'UTR of BRCA1/BRCA2 in familial and early-onset breast cancer patients with and without mutations in the coding regions of BRCA1/ BRCA2 and to identify specific 3'UTR variants that may be risk factors for cancer development. The 3'UTRs of the BRCA1 and BRCA2 genes were screened by heteroduplex analysis and DNA sequencing in 100 patients from 46 BRCA1/2 families, 54 non-BRCA1/2 families, and 47 geographically matched controls. Two polymorphisms were identified. SNPs c.*1287C>T (rs12516) (BRCA1) and c.*105A>C (rs15869) (BRCA2) were identified in 27% and 24% of patients, respectively. These 2 variants were also identified in controls with no family history of cancer (23.4% and 23.4%, respectively). In comparison to variations in the 3'UTR region of the BRCA1/2 genes and the BRCA1/2 mutational status in patients, there was a statistically significant relationship between the BRCA1 gene polymorphism c.*1287C>T (rs12516) and BRCA1 mutations (p=0.035) by Fisher's Exact Test. SNP c.*1287C>T (rs12516) of the BRCA1 gene may have potential use as a genetic marker of an increased risk of developing breast cancer and likely represents a non-coding sequence variation in BRCA1 that impacts BRCA1 function and leads to increased early-onset and/or familial breast cancer risk in the Turkish population.

BRCA1/2 germline mutations and their clinical importance in Turkish breast cancer patients.

BRCA1/BRCA2 genes were screened in 117 patients with breast cancer by sequencing. Fourteen percent of patients tested positive for BRCA1/BRCA2 mutations. Four frame shift mutations, four pathogenic missense mutations, and 25 different sequence variations were detected. BRCA mutation positivity was significantly associated with Ki67 (p = .001). BRCA protein expressions were decreased in the patients harboring important mutations and polymorphisms (BRCA1;P508 stop, V1740G, Q1182R, Q1756P and BRCA2;V2466A) related with disease. Our findings contribute significantly to the types of germline BRCA1/BRCA2 mutations and their biological effects in Turkish women. These data could help guide the management of BRCA1/BRCA2 mutation-carrying patients when considering breast-conserving therapy.

Expression status of let-7a and miR-335 among breast tumors in patients with and without germ-line BRCA mutations.

The genetic factors of cancer predisposition remain elusive in the majority of familial and/or early-onset cases of breast cancer (BC). This type of BC is promoted by germ-line mutations that inactivate BRCA1 or BRCA2. On the other hand, recent studies have indicated that alterations in the levels of miRNA expression are linked to this disease. Although BRCA1 and BRCA2 gene mutations have been reported to commonly lead to alterations in genes that encode cancer-related proteins, little is known regarding the putative impact of these mutations on noncoding miRNAs. In the present study, we aimed to determine whether miRNA dysregulation is involved in the pathogenesis of BRCA-mutated BC. An expression analysis of 14 human miRNAs previously shown to be related to BC diagnosis, prognosis, and drug resistance was conducted using tissues from 60 familial and/or early-onset patients whose peripheral blood samples had been screened for BRCA1 and BRCA2 mutations through sequence analysis. Let-7a and miR-335 expression levels were significantly downregulated in the tumors of patients with a BRCA mutation compared with those of patients without a BRCA mutation (P = 0.04 and P = 0.02, respectively). Our results defined the associations between the expression status of let-7a and miR-335 and BRCA mutations. The expression analysis of these miRNAs might be used as biomarkers of the BRCA mutation status of early-onset and/or familial BC.

Molecular markers for patients with thymic malignancies: not feasible at present?

BACKGROUND: Thymomas and thymic carcinomas are rare malignancies and devising clinically effective molecular targeted therapies is a major clinical challenge. The aim of the study was to analyze BLC2 and vascular endothelial growth factor receptor (VEGFR) expression and KRAS and EGFR mutational status and to correlate them with the clinical characteristics of patients with thymomas and thymic carcinomas. MATERIALS AND METHODS: A total of 62 patients (mean age: 50.4 ± 13.2 years) with thymomas and thymic carcinomas were enrolled. The expression of BLC2 and VEGFR in tumor cells and normal tissues was evaluated by RT-PCR. The mutational status of the KRAS and EGFR genes was investigated by PCR with sequence specific primers. RESULTS: The BLC2 and VEGFR expression levels did not differ significantly between tumor and normal tissues. Moreover, there were no clearly pathogenic mutations in KRAS or EGFR genes in any tumor. None of the molecular markers were significantly related to clinical outcomes. CONCLUSIONS: Changes in levels of expression of BLC2 and VEGFR do not appear to be involved in thymic tumorigenesis. Moreover, our data suggest that KRAS and EGFR mutations do not play a major role in the pathogenesis of thymomas and thymic carcinomas.

Microsatellite instability status affects gene expression profiles in early onset colorectal cancer patients.

BACKGROUND: The association between microsatellite instability (MSI) status and gene expression profiles in the early onset sporadic colorectal cancer (CRC) has not been clearly established. The aim of this study was to identify the altered gene expression patterns depending on the MSI status of early onset CRC and determine specific biomarkers that could provide novel therapeutic molecular targets in the Turkish population. MATERIALS AND METHODS: MSI markers (BAT25, BAT26, D2S123, D5S346, and D17S250) were investigated in tumors from 36 early onset sporadic CRC patients in whom gene expression profiles were analyzed previously. The relationship between the gene expression profiles depending on MSI status was evaluated. RESULTS: A total of 15 tumors (16.66%) were identified as having MSI and 21 tumors (58.33%) were identified as having microsatellite stability (MSS). CK20 and MAP3K8 upregulation, observed in MSS tumors, was significantly associated with lymph node metastasis, recurrence, and/or distant metastasis and a short median survival (P < 0.05). REG1A upregulation is also correlated with recurrence and/or distant metastasis and a short median survival in patients with MSI tumors (P < 0.05). CONCLUSIONS: High expression levels of CK20 and MAP3K8 in MSS tumors and REG1A in MSI tumors correlated with a poor prognosis in CRC patients. Further studies and validations are required; these genes may provide novel therapeutic molecular targets for the development of anticancer drugs related to MSI status for early onset CRC treatment.

CK19, CK20, EGFR and HER2 status of circulating tumor cells in patients with breast cancer.

AIMS AND BACKGROUND: The major cause of death in breast cancer patients is metastasis. Various biomarkers have been used for the early detection of circulating tumor cells in the peripheral blood of breast cancer patients. The aims of the current study were to analyze circulating tumor cells in the blood of breast cancer patients by investigating EGFR, CK19, CK20 and HER2 expression profiles and to evaluate their prognostic importance. METHODS: CK19, CK20 and EGFR gene expression profiles were evaluated in the blood samples of 84 female patients with primary invasive ductal breast cancer and 20 healthy female volunteers using SYBR green-based real-time qPCR assays. HER2 expression analyses were conducted in 46 patients who had an HER2-positive primary tumor and in 30 healthy women to determine the cutoff level of positivity. RESULTS: The positive rates of CK20, EGFR, CK19 and HER2 mRNA expression in the peripheral blood were 28.57% (24/84), 20.23% (17/84), 5.95% (5/84) and 2.17% (1/46), respectively. The high positive ratio of CK20 mRNA expression in the peripheral blood of breast cancer was identified for the first time in the current study. Significant differences were identified in CK20 expression status and several clinical parameters related with aggressiveness of tumors using a binary logistic regression analysis. Higher CK20-positive levels were observed in patients who had lymph node metastasis and advanced-grade primary tumors, which were estrogen receptor-negative. We have demonstrated that CK20 may be a novel biomarker that is useful to identify circulating tumor cells and predict breast cancer progression. CONCLUSIONS: The results suggest that the investigation of CK20 mRNA with other biomarkers in the peripheral blood of breast cancer patients may be useful to monitor the presence of disseminated tumor cells in the blood circulation and to predict the prognosis of breast cancer.

The promoter hypermethylation status of GATA6, MGMT, and FHIT in glioblastoma.

Glioblastoma (GBM) is an aggressive and lethal cancer, accounting for the majority of primary brain tumors in adults. GBMs are characterized by large and small alterations in genes that control cell growth, apoptosis, angiogenesis, and invasion. Epigenetic alterations also affect the expression of cancer genes, either alone or in combination with genetic mechanisms. The current evidence suggests that hypermethylation of promoter CpG islands is a common epigenetic event in a variety of human cancers. A subset of GBMs is also characterized by a locus-specific and genome-wide decrease in DNA methylation. Epigenetic alterations are important in the molecular pathology of GBM. However, there are very limited data about these epigenetic alterations in GBM. Alterations in promoter methylations are important to understand because histone deacetylases are targets for drugs that are in clinical trial for GBMs. The aim of the current study was to investigate whether the promoter hypermethylation of putative tumor suppressor genes was involved in GBM. We examined the methylation status at the promoter regions of GATA6, MGMT, and FHIT using the methylation-specific polymerase chain reaction in 61 primary GBMs. Our results reveal that there is no promoter hypermethylation of FHIT in the examined GBM tissue specimens. In contrast, the promoter hypermethylation of GATA6 and MGMT was detected in 42.8 and 11.11% of GBMs, respectively. The frequency of MGMT promoter hypermethylation was low in the group of patients we evaluated. In conclusion, our study demonstrates that promoter hypermethylation of MGMT is a common event in GBMs, whereas GATA6 is epigenetically affected in GBMs. Furthermore, inactivation of FHIT by epigenetic mechanisms in GBM may not be associated with brain tumorigenesis.