Circulating MiR-125b as a marker predicting chemoresistance in breast cancer

Background

Chemotherapy is an important component in the treatment paradigm for breast cancers. However, the resistance of cancer cells to chemotherapeutic agents frequently results in the subsequent recurrence and metastasis. Identification of molecular markers to predict treatment outcome is therefore warranted. The aim of the present study was to evaluate whether expression of circulating microRNAs (miRNAs) can predict clinical outcome in breast cancer patients treated with adjuvant chemotherapy.

Methodology/Principal Findings

Circulating miRNAs in blood serum prior to treatment were determined by quantitative Real-Time PCR in 56 breast cancer patients with invasive ductal carcinoma and pre-operative neoadjuvant chemotherapy. Proliferating cell nuclear antigen (PCNA) immunostaining and TUNEL were performed in surgical samples to determine the effects of chemotherapy on cancer cell proliferation and apoptosis, respectively. Among the miRNAs tested, only miR-125b was significantly associated with therapeutic response, exhibiting higher expression level in non-responsive patients (n = 26, 46%; p = 0.008). In addition, breast cancers with high miR-125b expression had higher percentage of proliferating cells and lower percentage of apoptotic cells in the corresponding surgical specimens obtained after neoadjuvant chemotherapy. Increased resistance to anticancer drug was observed in vitro in breast cancer cells with ectopic miR-125b expression; conversely, reducing miR-125b level sensitized breast cancer cells to chemotherapy. Moreover, we demonstrated that the E2F3 was a direct target of miR-125b in breast cancer cells.

Conclusions/Significance

These data suggest that circulating miR-125b expression is associated with chemotherapeutic resistance of breast cancer. This finding has important implications in the development of targeted therapeutics for overcoming chemotherapeutic resistance in novel anti-cancer strategies.     

The role of plasma IDO activity as a diagnostic marker of patients with colorectal cancer

High levels of indoleamine 2,3-dioxygenase (IDO) are involved in tumour escape mechanisms. The aim of this study is the evaluation of l-kynurenine of plasma as marker of diagnostic and prognostic in patients with colorectal cancer. The study included 78 patients with colorectal cancer, of whom 15 % were in stage I/II, 30 % in stage III, and 55 % in stage IV, and was compared with a control group of 70 healthy subjects. The receiver operating characteristic (ROC) curve analysis showed an area under the curve of 0.917, with a specificity of 100 % and with a sensitivity to detect cancer of the colon of 85.2 %, taking 1.83 μM as a cut-off point. The overall survival analysis also indicated that patients with low levels of l-kynurenine in plasma increased survival rate after 45 months of follow-up (P = 0.032). These results show that the plasma levels of l-kynurenine could be a good biomarker to differentiate individuals with colorectal cancer from healthy individuals.     

CA 15.3 as a tumour marker in breast cancer

CA 15.3 is an antigenic determinant associated with human mammary carcinomas. Two murine monoclonal antibodies have been raised against the determinants, and an immunoradiometric assay (IRMA-Kit, Centocor, USA) has been developed to determine the antigen levels in plasma of cancer patients. Based on the 99% confidence limit of healthy women, plasma values above 30 U/ml are considered abnormal. Plasma samples from 357 women were examined in the present study. Healthy females (n = 84) ranged below the cut-off level between less than 10 and 29 U/ml. Higher values were found in 12.5% of benign breast diseases and in 23.6% of breast cancer patients, including all stages. Depending on the stage of the disease, there were elevated levels in 11% of operable breast cancer patients preoperatively, in 7% of the cases with no evidence of disease after primary treatment and in 63.5% of patients with disseminated mammary carcinoma. In metastasized breast cancer the frequency and the degree of abnormal titers were closely related to the extent of the metastatic disease. Follow-up examinations of 63 patients under cytotoxic therapy showed CA 15.3 changes correlating well with the clinical course in up to 90% of the antigen positive cases. The present data indicate that CA 15.3 may be useful in the surveillance of breast cancer patients. However in our study one third of the patients with metastatic breast cancer did not show any increase in CA 15.3 and must be regarded as antigen negative.     

CA 549 as a marker in breast cancer.

CA 549 is one of several carcinoma associated mucin antigens proposed as a breast cancer tumor marker. In this study, the performance characteristics of the CA 549 assay were validated and the clinical utility of the test was compared with that of other breast cancer markers including CA 15-3, CA M26, CA M29 and carcinoembryonic antigen. The upper limit of normal was established as 15.5 U/ml based on data for 250 control subjects apparently free of disease. Overall, CA 549 had a low negative predictive value (0.51) due to a low sensitivity in the detection of early breast cancer. However, the test had a high positive predictive value (0.93) reflecting a high specificity for the disease. In 56 patients with advanced breast cancer, the sensitivity was 0.71 for CA 549 alone and 0.79-0.84 for CA 549 combined with any of the other markers studied.     

Promoter hypermethylation of the CFTR gene as a novel diagnostic and prognostic marker of breast cancer

Breast cancer is the most commonly diagnosed cancer and the leading cause of cancer‐related deaths among women. New biomarkers with definite diagnostic and prognostic efficacy are urgently needed. Here, we showed that the promoter of the cystic fibrosis transmembrane conductance regulator (CFTR) was hypermethylated in breast cancer. The messenger RNA level of CFTR was downregulated in breast cancer. Notably, all 19 breast cancer patients with hypermethylated CFTR were diagnosed with invasive carcinoma. Moreover, CFTR was upregulated in decitabine (10 μM) treated breast cancer cells. Overexpression of CFTR inhibited cell growth whereas knockdown of CFTR promoted cell invasion. In the tissue array analysis, the CFTR protein level decreased significantly in breast cancer and low CFTR protein level correlated with poor survival with a P‐value of 0.034. Thus, promoter hypermethylation of the CFTR gene might be a novel diagnostic marker of breast cancer.     

Circulating muscle-specific microRNA, miR-206, as a potential diagnostic marker for rhabdomyosarcoma

Presently there is no serum biomarker of rhabdomyosarcoma (RMS). Several studies have shown that profiles of microRNA (miRNA) expression differ among tumor types. Here we evaluated the feasibility of using muscle-specific miRNAs (miR-1, -133a, -133b and -206) as biomarkers of RMS. Expression of muscle-specific miRNAs, especially miR-206, was significantly higher in RMS cell lines than in other tumor cell lines, as well as in RMS tumor specimens. Further, serum levels of muscle-specific miRNAs were significantly higher in patients with RMS tumors than in patients with non-RMS tumors. Normalized serum miR-206 expression level could be used to differentiate between RMS and non-RMS tumors, with sensitivity of 1.0 and specificity of 0.913. These results raise the possibility of using circulating muscle-specific miRNAs, especially miR-206, as landmark biomarkers for RMS.     

Circulating nucleic acids as a tumor marker

Patients suffering from malignant diseases have been shown to have increased amounts of cell free nucleic acids in their circulation. As genetic and epigenetic alterations are increasingly characterized in different types of tumors, such changes can be used to detect tumor-derived nucleic acids in the circulation. To date, nearly all tumor-associated nucleic acids have been detected in the plasma or serum of cancer patients. Moreover, increased levels of circulating viral nucleic acids have also been demonstrated in patients with certain cancers associated with viral infections. The concentration of these tumor-associated nucleic acid species is generally related to the tumor load and the extent of the disease. Serial monitoring of plasma nucleic acids thus provides a good way to follow disease progress and to predict the outcome of such patients. In this review, different approaches of detecting tumor-related nucleic acids in the circulation and their potential as tumor markers in the screening, monitoring and prognostication of malignant diseases are discussed.     

Nuclear NHERF1 expression as a prognostic marker in breast cancer

Our purpose was to investigate whether Na⁺/H⁺ exchanger regulatory factor 1 (NHERF1) expression could be linked to prognosis in invasive breast carcinomas. NHERF1, an ezrin-radixin-moesin (ERM) binding phosphoprotein 50, is involved in the linkage of integral membrane proteins to the cytoskeleton. It is therefore believed to have an important role in cell signaling associated with changes in cell cytoarchitecture. NHERF1 expression is observed in various types of cancer and is related to tumor aggressiveness. To date the most extensive analyses of the influence of NHERF1 in cancer development have been performed on breast cancer. However, the underlying mechanism and its prognostic significance are still undefined. NHERF1 expression was studied by immunohistochemistry (IHC) in a cohort of 222 breast carcinoma patients. Association of cytoplasmic and nuclear NHERF1 expression with survival was analyzed. Disease-free survival (DFS) and overall survival (OS) were determined based on the Kaplan–Meier method. Cytoplasmic NHERF1 expression was associated with negative progesterone receptor (PgR) (P=0.017) and positive HER2 expression (P=0.023). NHERF1 also showed a nuclear localization and this correlated with small tumor size (P=0.026) and positive estrogen receptor (ER) expression (P=0.010). Multivariate analysis identified large tumor size (P=0.011) and nuclear NHERF1 expression (P=0.049) to be independent prognostic variables for DFS. Moreover, the nuclear NHERF1(−)/ER(−) immunophenotype (27%) was statistically associated with large tumor size (P=0.0276), high histological grade (P=0.0411), PgR-negative tumors (P<0.0001) and high proliferative activity (P=0.0027). These patients had worse DFS compared with patients with nuclear NHERF1(+)/ER(+) tumors (75.4% versus 92.6% P=0.010). These results show that the loss of nuclear NHERF1 expression is associated with reduced survival, and the link between nuclear NHERF1 and ER expression may serve as a prognostic marker for the routine clinical management of breast cancer patients.     

Circulating exosomal CPNE3 as a diagnostic and prognostic biomarker for colorectal cancer

Exosomal proteins are emerging as relevant diagnostic and prognostic biomarkers for cancer. This study was aimed at illustrating the clinical significance of exosomal Copine III (CPNE3) purified from the plasma of colorectal cancer (CRC) patients. The CPNE3 expression levels in CRC tissues were analyzed by real-time PCR, western blot, and immunohistochemistry. Plasma exosomes were isolated to examine the CPNE3 level using ELISA. Pearson’s correlation analysis was performed to investigate the CPNE3 levels between CRC tissues and matched plasma samples. Receiver operating characteristic curve analysis was developed to measure the diagnostic performance of exosomal CPNE3. The Kaplan–Meier method and Cox's proportional hazards model were utilized to determine statistical differences in survival times. CPNE3 showed increased expressions in the CRC tissues. A moderately significant correlation was found between CPNE3 expression in CRC tissues by immunohistochemistry and matched serum exosomal CPNE3 expression by ELISA (r = 0.645,(r = 0.645, p < 0.001). < 0.001). Exosomal CPNE3 yielded a sensitivity of 67.5% and a specificity of 84.4% in CRC at the cutoff value of 0.143 pg per 1ug1 ug exosome. Combined data from carcinoembryonic antigen and exosomal CPNE3 achieved 84.8% sensitivity and 81.2% specificity as a diagnostic tool. CRC patients with lower exosomal CPNE3 levels had substantially better disease-free survival (hazard ratio [HR], 2.9; 95% confidence interval [CI]: 1.3–6.4; p = 0.009) = 0.009) and overall survival (HR, 3.4; 95% CI: 1.2–9.9; p = 0.026) = 0.026) compared with those with higher exosomal CPNE3 levels. Exosomal CPNE3 show potential implications in CRC diagnosis and prognosis.     

Comprehensive analysis of novel three-long noncoding RNA signatures as a diagnostic and prognostic biomarkers of human triple-negative breast cancer

Currently, traditional predictors of prognosis (tumor size, nodal status, progesterone receptor [PR], estrogen receptor [ER], or human epidermal growth factor receptor-2 [HER2]) are insufficient for precise survival prediction for triple-negative breast cancer (TNBC). Long noncoding RNAs (lncRNAs) have been observed to exert critical functions in cancer, including in TNBC. Nevertheless, systematically tracking expression-based lncRNA biomarkers based on the sequence data for the prediction of prognosis in TNBC has not yet been investigated. To ascertain whether biomarkers exist that can distinguish TNBC from adjacent normal tissue or nTNBC, we implemented a comprehensive analysis of lncRNA expression profiles and clinical data of 1097 BC samples from The Cancer Genome Atlas database. A total of 1510 differentially expressed lncRNAs in normal and TNBC samples were extracted. Similarly, 672 differentially expressed lncRNAs between nTNBC and TNBC samples were detected. The receiver operating characteristic curve analysis indicated that three upregulated lncRNAs (AC091043.1, AP000924.1, and FOXCUT) may be of strong diagnostic value for predicting the existence of TNBC in the training and validation sets (area under the curve (AUC > 0.85). Kaplan-Meier analysis demonstrated that the other three lncRNAs (AC010343.3, AL354793.1, and FGF10-AS1) were associated with the prognosis of TNBC patients (P < 0.05). We used the three overall survival (OS)-related lncRNAs to establish a three-lncRNA signature. Multivariate Cox regression analysis suggested that the three-lncRNA signature was a prognostic factor independent of other clinical variables ( P < 0.01) for predicting OS in TNBC patients that could be utilized to classify patients into high- or low-risk subgroups. Our results might provide efficient signatures for clinical diagnosis and prognostic evaluation of TNBC.     

Circulating nucleic acids as a new diagnostic tool

The discovery of circulating nucleic acids in the 1940s opened up new possibilities for the non-invasive detection, monitoring and screening of various human disorders. Several tumour markers that enable early cancer detection or tumour behaviour prediction have been detected in the plasma of cancer patients. Maternal plasma analysis can be used to detect certain fetal abnormalities, with the quantification of cell-free nucleic acids used to screen for several pregnancy-associated disorders. Some other applications are in transplant monitoring and graft rejection assessment, and in certain medical emergencies such as trauma and burn severity stratification. Many studies have yielded promising results in this field, but the techniques have yet to be applied in routine clinical practice. Large-scale studies using similar technologies and a broad spectrum of patients are still needed to verify the results of the various studies.     

Evaluation of human LOX-12 as a serum marker for breast cancer

The high concentration of prostaglandins has been associated with chronic inflammatory diseases and several types of human cancers. This is due to the over expression of inflammatory enzymes like Cyclooxygenase (COX), Lipoxygenase (LOX) etc. The aim of this study was to quantify the LOX-12 with clinicopathological parameter of breast cancer patients and its response after chemotherapy to establish serum LOX-12 as a prognostic marker. This case-controlled study was performed on 86 biopsy proven breast cancer patients. Blood and tissue samples were collected from the patients. Serum LOX-12 of the study group was quantified by Surface Plasmon Resonance (SPR) and ELISA techniques by antibody–antigen interaction strategy. A significant increase in LOX-12 levels was observed in breast cancer patients (Mean ± SD = 40.54 ± 13.61 ng/ml) as compared to healthy controls (Mean ± SD = 13.42 ± 2.4 ng/ml) (p < 0.0001). Serum LOX-12 levels were significantly higher (p < 0.002) in patients with lymph node involvement. More than 75% patients had shown significant (p < 0.0001) reduction of LOX-12 levels after chemotherapy. This was also confirmed by ELISA. This study for the first time had co-related the quantity of serum LOX-12 with breast cancer and also with the effect of chemotherapy.     

Serum pleiotrophin as a diagnostic and prognostic marker for small cell lung cancer

Pleiotrophin (PTN) is involved in tumour progression, angiogenesis and metastasis. The purpose of this study was to investigate the expression level of PTN in the serum of patients with small cell lung cancer (SCLC) and to explore the clinical significance of PTN. Serum samples from 128 patients with SCLC, 120 healthy volunteers (HV) and 60 patients with benign lung disease (BLD) were collected. The levels of serum PTN were determined with ELISA and its correlation with the clinical data was examined. The serum PTN levels in SCLC patients were significantly higher than that in BLD patients (P < 0.05) or HV (P < 0.05). With a cutoff value of 258.18 ng/mL, the sensitivity and specificity of PTN to SCLC patients and BLD patients, SCLC patients and HV were 79.2% and 91.7%, 86.7% and 95.8% respectively. An area under the curve for all stages of SCLC resulting from PTN, which was significantly better than the other tumour markers tested including progastrin‐releasing peptide and neuron‐specific enolase. High serum PTN levels appear to correlate with poor survival in patients with SCLC. These results suggest that PTN levels in the serum could be a new effective biomarker for the diagnosis and prognosis of SCLC.     

Cyclin E as molecular marker in the management of breast cancer: a review

Over the past decade numerous molecular markers have been identified that may play a role in breast carcinogenesis and prognosis. The most commonly used markers in clinical practice are the estrogen receptor, progesterone receptor and HER-2/neu. Recent studies found cyclin E to be a promising prognostic indicator in breast cancer and examined its potential as a target for therapy. Further studies demonstrated that cyclin E levels were periodic during the cell cycle, with levels of protein peaking in the G1 phase. This peak in cyclin E levels also correlated with maximum enzymatic function of the cyclin E-cdk2 complex, suggesting a critical role of cyclin E in regulating G1 to S-phase transition. Studies examining the relevance of cyclin E alterations in breast cancer have shown gene amplifications in some breast cancer cell lines, data that provide strong support for the role of cyclin E in breast carcinogenesis. It is believed that the most significant cyclin E alteration is post-translational cleavage of full-length cyclin E into low molecular weight forms that are hyperactive compared to the 50-kDa, full-length protein and correlate with increasing stage and grade of breast cancer. The role of cyclin E in the prognosis and therapy of breast cancer is reviewed according to recent publications.     

Small non-coding RNA profiling in breast cancer: plasma U6 snRNA,miR-451a and miR-548b-5p as novel diagnostic and prognostic biomarkers

Molecular Biology Reports - Breast cancer is a leading cause of cancer-related death in women. Most cases are invasive ductal carcinomas of no special type (NST breast carcinomas). In this...