A Randomized Phase II Trial Investigating the Effect of Platelet Function Inhibition on Circulating Tumor Cells in Patients With Metastatic Breast Cancer

BackgroundBlockade of platelet activation and aggregation can inhibit metastasis in preclinical models and is associated with cancer prevention. To test whether disruption of platelet function with clopidogrel and aspirin would decrease the number of circulating tumor cells (CTCs) in patients with metastatic breast cancer, a randomized phase II study was performed.MethodsPatients with metastatic breast cancer who were not currently receiving cytotoxic chemotherapy were eligible. Patients were randomized to receive either clopidogrel and aspirin or to a control group receiving no treatment. Phlebotomy was performed at baseline, at 2 and 4 weeks, and monthly thereafter to obtain specimens to assess CTC, platelet aggregation, and thrombin activity. The primary end point was the proportion of patients with detectable CTCs at 1 month.ResultsForty-eight patients were enrolled and 42 were evaluable at 1 month. Baseline CTC numbers were ≥ 5 in 13% and ≥ 1 in 65% of patients. Despite adequate platelet function inhibition in the treatment group, the proportion of patients with detectable CTCs was similar between the clopidogrel/aspirin and control groups at baseline (P = .21) and 4 weeks (P = .75), showing no treatment effect. Measured endogenous thrombin potential did not correlate with CTC number. No bleeding-related serious adverse events (SAEs) occurred.ConclusionThe baseline CTC numbers were lower than expected, decreasing the ability to detect an impact of platelet inhibition on CTCs. Clopidogrel and aspirin were well tolerated. Future studies evaluating the potential therapeutic role of antiplatelet therapy in breast cancer remain of interest, and they may be informed by these results.     

Single circulating tumor cell detection and overall survival in nonmetastatic breast cancer

BackgroundCirculation of cancer cells in the blood is a mandatory step for metastasis, but circulating tumor cells (CTC) have a low metastatic efficiency in preclinical animal models. In this prospective study, we reported the clinical outcome of nonmetastatic breast cancer patients according to CTC detection.Patients and methodsIn 115 nonmetastatic patients diagnosed with large operable or locally advanced breast cancer, we prospectively detected CTC using the CellSearch system before and after neoadjuvant chemotherapy in a phase II trial (REMAGUS02).ResultsAt baseline, 23% of patients were CTC positive, but only 10% had >1 CTC/7.5 ml of blood. After a median follow-up of 36 months, CTC detection before chemotherapy was an independent prognostic factor for both distant metastasis-free survival [DMFS; P = 0.01, relative risk (RR) = 5.0, 95% confidence interval (CI) 1.4–17] and overall survival (OS; P = 0.007, RR = 9, 95% CI 1.8–45). CTC detection after chemotherapy was of less significance (P = 0.07 and 0.09, respectively). Moreover, CTC detection showed interesting characteristics as an individual predictive test for metastatic relapses (sensibility 55%, specificity 81%, and global accuracy 77%).ConclusionsDetection of ≥1 CTC/7.5 ml before neoadjuvant chemotherapy can accurately predict OS. Our findings may change the clinical management of nonmetastatic breast cancer and indicate that the metastatic efficiency of CTC could be higher than previously reported.     

乳腺癌循环肿瘤细胞研究进展与展望

目的:总结国内外乳腺癌循环肿瘤细胞(CTC)在检测手段及相关临床意义方面的研究进展,以及CTC检测项目在研究肿瘤转移过程中的地位、必要性及未来的发展方向.方法:应用Medline、PubMed及CNKI期刊全文数据库系统,以“乳腺癌、循环肿瘤细胞、转移、上皮间质转化”等为关键词,检索2006-06-2011-06的相关文献,共选择文献48篇加以分析总结.文献纳入标准:1)乳腺癌CTC的起源及其特点;2)乳腺癌CTC的检测方法;3)检测乳腺癌CTC的临床意义;4)乳腺癌CTC的特性.结果:CTC的检测在预测乳腺癌患者无病生存期和总生存期方面有重要价值;同时,它在预测和判断疗效方面优于一些传统方法.除此之外,CTC的分子学和生物学特性,如ER、PR、HER-2表达和上皮间质转化等,以及这些特性与乳腺癌转移的关系已成为新的研究热点.结论:对CTC的深入研究能够使我们进一步了解它,使其成为乳腺癌患者监测转移和预后的常规的检测指标,并有助于了解血行转移的机制,更有望发现乳腺癌治疗的新靶点.     

Mesenchymal circulating tumor cells (CTCs) and OCT4 mRNA expression in CTCs for prognosis prediction in patients with non-small-cell lung cancer

Purpose

Circulating tumor cells (CTCs) with epithelial-to-mesenchymal transition (EMT) phenotypes might be related to tumor progression while OCT4 expression is involved in tumor metastasis and poor prognosis. But the possible clinical significance of EMT phenotypes of CTCs from non-small-cell lung cancer (NSCLC) patients has still to be demonstrated. Furthermore, none has been investigated the expression of OCT4 in CTCs. We therefore identified the EMT phenotype-based subsets of CTCs and determined the OCT4 expression status of CTCs in NSCLC patients, to explore their possible clinical relevance.

Methods

37 NSCLC patients and ten healthy volunteers were enrolled, respectively. The Canpatrol? CTC enrichment technique was used to isolate and identify the EMT phenotype-based subsets of CTCs. OCT4 expression in each CTC was also determined. Results were correlated with patients’ clinico-pathological features.

Results

CTCs were detected in 33 of 37 (89.2%) NSCLC patients, and no CTCs were identified in ten healthy volunteers. Three CTCs phenotypes, including epithelial, biophenotypic, and mesenchymal CTCs were identified based on the expression of EMT markers. Mesenchymal CTCs were more commonly found in patients with distant metastasis. Patients with distant metastasis tended to have a higher median CTCs number. OCT4-positive was observed in 21 of 28 (75.0%) patients. High expression of OCT4 tended to occur in advanced patients as well as in distant metastatic patients.

Conclusions

The findings suggest that identification of CTCs by EMT markers as well as evaluation of OCT4 expression status by assessment of OCT4 expression in CTCs could serve as potential adjuncts for evaluating metastasis and prognosis in NSCLC patients.

     

MicroRNA‐708‐3p mediates metastasis and chemoresistance through inhibition of epithelial‐to‐mesenchymal transition in breast cancer

Metastasis and chemoresistance remain major challenges in the clinical treatment of breast cancer. Recent studies show that dysregulated microRNAs (miRNAs) play an important role in metastasis and chemoresistance development in breast cancer. Herein, we identified downregulated expression of miR‐708‐3p in breast cancers. In particular, miR‐708‐3p expression was significantly decreased in specimens from breast cancer patients with metastasis compared to that in specimens from patients with no metastasis. Consistent with clinical data, our in vitro data show that miR‐708‐3p was more significantly decreased in invasive breast cancer cell lines. In addition, our data show that inhibition of miR‐708‐3p significantly stimulated breast cancer cell metastasis and induced chemoresistance both in vitro and in vivo. In contrast, overexpression of miR‐708‐3p dramatically inhibited breast cancer cell metastasis and enhanced the sensitivity of breast cancer cells to chemotherapy both in vitro and in vivo. Furthermore, we identified that miR‐708‐3p inhibits breast cancer cell epithelial‐to‐mesenchymal transition (EMT) by directly targeting EMT activators, including ZEB1, CDH2 and vimentin. Taken together, our findings suggest that miR‐708‐3p acts as a cancer suppressor miRNA and carries out its anticancer function by inhibiting EMT in breast cancer. In addition, our findings suggest that restoration of miR‐708‐3p may be a novel strategy for inhibiting breast cancer metastasis and overcoming the chemoresistance of breast cancer cells.     

Significance of circulating tumor cells in the portal vein regarding metastases and vascular invasion in hepatocellular carcinoma patients

BackgroundVascular invasion is an important risk factor of poor prognosis in hepatocellular carcinoma (HCC) patients. The detection of circulating tumor cells (CTCs) in the blood is direct evidence of tumor presence. There are few reports on CTCs and metastasis and vascular invasion of HCC. The purpose of this study was to analyze the significance of CTCs in the portal vein regarding metastases and vascular invasion in HCC patients.MethodsA total of 104 HCC patients diagnosed and treated in Zhengzhou University People’s Hospital were enrolled. Surgery was performed in 60 individuals. Portal vein blood samples were collected before treatment for CTCs detection. We used the isolation by size of epithelial tumor cells (ISET) and fluorescence in situ hybridization (FISH) to enrich and classify CTCs from blood samples. The patients were divided into metastasis and nonmetastasis groups according to the metastasis status before treatment. Differences in clinical indicators such as alpha-fetoprotein (AFP) levels, tumor size, CTCs count, and macrovascular tumor thrombus between the two groups were analyzed as well as the associations of CTCs count with the above indicators. For individuals with postoperative pathology, the relationship between CTCs counts and microvascular invasion (MVI) was analyzed.ResultsThe amounts of portal vein CTCs were higher in patients with metastases compared with the nonmetastases group (20 vs. 7; z=3.795; P<0.001). Multivariate logistic regression analysis showed that the CTC count was a risk factor for HCC metastasis [odds ratio (OR) =1.044; 95% CI: 1.011–1.079]. The sensitivity and specificity of CTC count in predicting HCC metastasis were 82.93% and 52.38%, respectively. CTC count was significantly correlated with tumor size (r_s=0.308; P=0.001), vascular invasion (z=4.211; P<0.001), and MVI (z=12.763; P=0.002). A threshold CTC count of seven showed the most significant power for predicting metastasis.ConclusionsVascular invasion positivity was closely related to HCC metastasis. Portal vein CTC count before treatment was correlated with vascular invasion and could be considered one of the factors affecting HCC metastasis. However, the ability of CTC count was limited in predicting HCC metastasis due to insufficient specificity.     

Trastuzumab versus observation for HER2 nonamplified early breast cancer with circulating tumor cells (EORTC 90091-10093, BIG 1-12, Treat CTC): a randomized phase II trial

BackgroundTrastuzumab improves the outcome of women with HER2 positive breast cancer. We aimed to assess whether trastuzumab decreases the detection rate of circulating tumor cells (CTCs) in women with high risk, HER2 nonamplified, early breast cancer.Patients and methodsThe EORTC 90091-10093 BIG 1–12 Treat CTC is a phase II trial, conducted in 70 hospitals and 6 CTC laboratories across 5 European countries. Patients with centrally confirmed HER2 nonamplified breast cancer and ≥1 centrally confirmed CTC per 15 ml of blood by CellSearch® following surgery and (neo)adjuvant chemotherapy were randomized (1 : 1) to 6 cycles of trastuzumab intravenously versus 18 weeks of observation. Randomization was stratified for center, locally confirmed estrogen receptor status and adjuvant versus neoadjuvant chemotherapy. The primary end point was rate of detection of ≥1 CTC per 15 ml of blood at week 18. Secondary end points were invasive disease-free survival (iDFS) and cardiac safety.ResultsBetween 30 April 2013 and 17 October 2016, 1317 patients were screened; 95 (7.2%) had detectable CTC(s), and 63 (4.8%) were randomized to trastuzumab (n = 31) or observation (n = 32). Fifty-eight patients were assessable for the primary end point, 29 in each arm. In 9 of the 58 patients, CTC(s) were still detected at week 18 : 5 in the trastuzumab and 4 in the observation arm (one-sided Fisher’s exact test, P = 0.765). An Independent Data Monitoring Committee recommended stopping further accrual for futility for the primary end point. Median follow-up at database lock was 13 months (IQR 4–16.5). The 1-year iDFS was 93.8% (95% CI 77.3–98.4) in the observation versus 84.8% (95% CI 63.4–94.2) in the trastuzumab arm. No grade 2–4 cardiac events were observed in the trastuzumab arm.ConclusionTrastuzumab does not decrease the detection rate of CTCs in HER2 nonamplified, nonmetastatic breast cancer.     

循环肿瘤细胞在转移性乳腺癌中的表达及其与多种血清肿瘤标志物的关系

目的 探讨转移性乳腺癌的循环肿瘤细胞(CTC)和血清肿瘤标志物(CEA、CA153和CA125)的表达水平以及两者之间的相关性。方法 采用CellSearch自动检测系统检测93例转移性乳腺癌患者开始新治疗前的循环肿瘤细胞(CTC),同时采用电化学发光法检测这些患者血清肿瘤标志物(CEA、CA153和CA125)的表达水平。结果 CTC阳性率为60%(56/93),与患者激素受体的状态、Her-2的状态、既往治疗的线数以及内脏转移无关。CEA的阳性率为56%(52/93),ER或PR阳性的患者CEA的阳性率更高(χ²=4.550,P=0.045),与Her-2的状态、既往治疗的线数以及内脏转移无关;CA153的阳性率为47%(44/93),与患者激素受体的状态、Her-2的状态、既往治疗的线数以及内脏转移无关;CA125的阳性率为41%(38/93);既往治疗大于等于二线的患者CA125的阳性率更高(χ²=4.501,P=0.038),与患者激素受体的状态、Her-2的状态以及是否有内脏转移无关。CTC和CEA之间呈正相关(r=0.296,P=0.004);CTC和CA153之间呈正相关(r=0.286,P=0.005);CTCs和CA125两者之间无相关性(r=0.184,P=0.077)。结论 转移性乳腺癌患者的循环肿瘤细胞(CTC)检出率高,而且与血清肿瘤标志物(CEA和CA153)有明显相关性,提示联合检测转移性乳腺癌的循环肿瘤细胞(CTC)和血清肿瘤标志物(CEA和CA153)可能会对其治疗决策有所帮助,同时CTC有可能成为转移性乳腺癌新的治疗靶点。     

Circulating tumor cells in locally advanced pancreatic adenocarcinoma: the ancillary CirCe 07 study to the LAP 07 trial

BackgroundPancreatic carcinoma is one of the leading causes of cancer-related mortality. At the time of diagnosis, 30% of patients present with a locally advanced pancreatic carcinoma (LAPC). As circulating tumor cells (CTCs) count may be a surrogate of the cancer metastatic abilities, CTC detection rates and prognostic value were studied in a prospective cohort of LAPC patients.Patients and methodsAn LAP07 international multicenter randomized study assesses in patients whose LAPC is controlled after 4 months of chemotherapy whether chemoradiotherapy could increase survival versus continuation of chemotherapy. A subgroup of patients included in the LAP07 trial was screened for CTCs (CellSearch®) before the start of the chemotherapy and after 2 months of treatment. Patient characteristics and survival were obtained prospectively and were correlated with CTC detection.ResultsSeventy-nine patients were included. One or more CTCs/7.5 ml were detected in 5% of patients before treatment and in 9% of patients after 2 months of treatment (overall detection rate: 11% of patients). CTC positivity was associated with poor tumor differentiation (P = 0.04), and with shorter overall survival (OS) in multivariable analysis (RR = 2.5, P = 0.01), together with anemia.ConclusionsThe evaluation of micrometastatic disease using CTC detection appears as a promising prognostic tool in LAPC patients.     

Circulating tumour cells in locally advanced breast cancer

Material and methods

A prospective study was conducted to determine the value of changes in circulating tumour cell (CTC) levels prior to and after the first cycle of neoadjuvant treatment in early prediction of pathologic response in locally advanced breast cancer (LABC). Two blood samples were obtained from 72 eligible LABC patients to isolate and enumerate CTCs before neoadjuvant chemotherapy started on day 1, and on day 21, immediately before second cycle administration.

Results

Sixty patients (83.3%) had <1 CTC in the first sample and response rates in this cohort were pathologic complete response (PCR) in 2 patients (5%), partial response (PR) in 35 (87.5%), stable disease (SD) in 2 (5%) and progressive disease (PD) in 1 (2.5%). Twelve patients (16.7%) had >2 CTCs in the first sample; these patients were more likely to have triple negative tumours. All 12 had fewer CTCs in the second sample. Response rates in this second cohort of 12 patients were PCR in 4 (34%), PR in 6 (50%), SD in 1 (8%) and PD in 1 (8%). PCR rate was markedly better in this second cohort (p<0.0042; OR 14.5, 95% CI 2.3–92).

Discussion

This study suggests that the presence of CTCs prior to neoadjuvant therapy might be a predictor of response to this therapy.     

A novel platform for detection of CK+ and CK- CTCs

Metastasis is a complex, multistep process that begins with the epithelial-mesenchymal transition (EMT). Circulating tumor cells (CTC) are believed to have undergone EMT and thus lack or express low levels of epithelial markers commonly used for enrichment and/or detection of such cells. However, most current CTC detection methods target only EpCAM and/or cytokeratin (CK) to enrich epithelial CTCs, resulting in failure to recognize other, perhaps more important, CTC phenotypes that lack expression of these markers. Here, we describe a population of complex aneuploid CTCs that do not express CK or CD45 antigen in patients with breast, ovarian, or colorectal cancer. These cells were not observed in healthy subjects. We show that the primary epithelial tumors were characterized by similar complex aneuploidy, indicating conversion to an EMT phenotype in the captured cells. Collectively, our study provides a new method for highly efficient capture of previously unrecognized populations of CTCs. Significance: Current assays for CTC capture likely miss populations of cells that have undergone EMT. Capture and study of CTCs that have undergone EMT would allow a better understanding of the mechanisms driving metastasis.     

HER2 status of circulating tumor cells in patients with metastatic breast cancer: a prospective, multicenter trial

There is a growing body of evidence that HER2 status can change during disease recurrence or progression in breast cancer patients. In this context, re-evaluation of HER2 status by assessment of HER2 expression on circulating tumor cells (CTCs) is a strategy with potential clinical application. The aim of this trial was to determine the HER2 status of CTCs in metastatic breast cancer patients comparing two CTC assays. A total of 254 patients with metastatic breast cancer from nine German university breast cancer centers were enrolled in this prospective study. HER2 status of CTCs was assessed using both the FDA-approved CellSearch^® assay and AdnaTest BreastCancer?. Using the CellSearch assay, 122 of 245 (50%) patients had ≥5 CTCs, and HER2-positive CTCs were observed in 50 (41%) of these patients. Ninety of 229 (39%) patients were CTC positive using AdnaTest BreastCancer, and HER2 positivity rate was 47% (42 of 90). The rate of breast cancer patients with HER2-negative primary tumors but HER2-positive CTCs was 32% (25 of 78) and 49% (28 of 57) using the CellSearch assay and AdnaTest BreastCancer, respectively. Considering only those patients who had CTCs on both tests (n = 62), concordant results regarding HER2 positivity were obtained in 50% of the patients (31/62) (P = 0.96, κ = ?0.006). HER2-positive CTCs can be detected in a relevant number of patients with HER2 negative primary tumors. Therefore, it will be mandatory to correlate the assay-dependent HER2 status of CTCs to the clinical response on HER2-targeted therapies.     

Circulating Tumor Cells are Associated with Bone Metastasis of Lung Cancer

Lung cancer (LC) is the leading cause of cancer mortality worldwide, predominantly due to the difficulty of early diagnosis and its high metastatic potential. Recently, increasing evidence suggests that circulating tumour cells (CTCs) are responsible for cancer metastatic relapse, and CTCs have attracted interest in cancer metastasis detection and quantification. In present study, we collected blood samples from 67 patients with bone metastasis, and 30 patients without such metastasis, and searched for CTCs. Then the association of CTC numbers with bone metastasis and other clinico-pothological variants was analyzed. Results demonstrated that when 5 or 1 was taken as a threshhold for the CTC number, there were significantly higher positivity of CTCs in the bonemetastasis group than in the non-metastasis group. While the increase in CTC number was not significantly associated with any other clinicopathological factor, including age, gender, pathological type, intrapulmonary metastasis and lymph node metastasis, the CTC number in patients with positivity of the last above mentioned variants was obviously higher than in patients with negativity of the two variants. Taken together, the CTC number appears to be significantly associated with the bone metastasis from lung cancer.     

High independent prognostic and predictive value of circulating tumor cells compared with serum tumor markers in a large prospective trial in first-line chemotherapy for metastatic breast cancer patients

BackgroundCirculating tumor cells (CTCs) are a prognostic marker in metastatic breast cancer, but comparisons with serum tumor markers (CA 15-3, carcinoembryonic antigen and lactate dehydrogenase) variations are needed.Patients and methodsCTCs were counted with CellSearch® at baseline, before cycle 2 (C2) and cycle 3 or 4 (C3/4) in 267 metastatic breast cancer patients on first-line chemotherapy with/without targeted therapy.ResultsBaseline CTC detection rate was 65% with ≥1 CTC/7.5 ml threshold and 44% with ≥5 CTC/7.5 ml and was independent of subtypes (luminal, triple negative, human epithelial growth factor receptor 2 (HER2)+). CTCs were associated with tumor markers, bone/liver involvement, tumor burden and performance status. CTC detection ≥1 CTC/7.5 ml was a strong prognostic factor for progression-free survival (PFS), P < 0.0001. Threshold of CTC ≥5 was statistically significant for PFS and overall survival (OS), P = 0.03 on multivariate analysis. Among patients with ≥5 CTC/7.5 ml at baseline, 50% had <5 CTC/7.5 ml at C2. Changes were correlated with both PFS and OS (P < 0.0001). All patients receiving anti-HER2 therapy had <5 CTC/7.5 ml after three cycles of treatment.ConclusionThis is the largest prospective series validating the prognostic value of CTC independently from serum tumor marker. Elevated CTCs before C2 are an early predictive marker of poor PFS and OS, which could be used to monitor treatment benefit. CTC decrease under treatment seems stronger with targeted therapy.     

Denosumab treatment is associated with the absence of circulating tumor cells in patients with breast cancer

Background

The presence of circulating tumor cells (CTCs) in patients with breast cancer correlates to a bad prognosis. Yet, CTCs are detectable in only a minority of patients with progressive breast cancer, and factors that influence the abundance of CTCs remain elusive.

Methods

We conducted CTC isolation and enumeration in a selected group of 73 consecutive patients characterized by progressive invasive breast cancer, high tumor load and treatment discontinuation at the time of CTC isolation. CTCs were quantified with the Parsortix microfluidic device. Clinicopathological variables, blood counts at the time of CTC isolation and detailed treatment history prior to blood sampling were evaluated for each patient.

Results

Among 73 patients, we detected at least one CTC per 7.5 ml of blood in 34 (46%). Of these, 22 (65%) had single CTCs only, whereas 12 (35%) featured both single CTCs and CTC clusters. Treatment with the monoclonal antibody denosumab correlated with the absence of CTCs, both when considering all patients and when considering only those with bone metastasis. We also found that low red blood cell count was associated with the presence of CTCs, whereas high CA 15-3 tumor marker, high mean corpuscular volume, high white blood cell count and high mean platelet volume associated specifically with CTC clusters.

Conclusions

In addition to blood count correlatives to single and clustered CTCs, we found that denosumab treatment associates with most patients lacking CTCs from their peripheral circulation. Prospective studies will be needed to validate the involvement of denosumab in the prevention of CTC generation.

     

Molecular characterization and heterogeneity of circulating tumor cells in breast cancer

Introduction

This study analyzes peripheral blood samples from breast cancer (BC) patients. CTCs from peripheral blood were enriched by size-based separation and were then cultivated in vitro. The primary aim of this study was to demonstrate the antigen independent CTC separation method with high CTC recovery. Subsequently, CTCs enriched several times during the treatment were characterized molecularly.

Methods

Patients with different stages of BC (N = 167) were included into the study. All patients were candidates for surgery, surgical diagnostics, or were undergoing chemotherapy. In parallel, 20 patients were monitored regularly and in addition to CTC presence, also CTC character was examined by qPCR, with special focus on HER2 and ESR status.

Results

CTC positivity in the cohort was 76%. There was no significant difference between the tested groups, but the highest CTC occurrence was identified in the group undergoing surgery and similarly in the group before the start of neoadjuvant treatment. On the other hand, the lowest CTC frequencies were observed in the menopausal patient group (56%), ESR+ patient group (60%), and DCIS group (44.4%). It is worth noting that after completion of neoadjuvant therapy (NACT) CTCs were present in 77.7% of cases. On the other hand, patients under hormonal treatment were CTC positive only in 52% of cases.

Discussions

Interestingly, HER2 and ESR status of CTCs differs from the status of primary tumor. In 50% of patients HER2 status on CTCs changed not only from HER2+ to HER2?, but also from HER2– to HER2+ (33%). ESR status in CTCs changed only in one direction from ESR+ to ESR?.

Conclusions

Data obtained from the present study suggest that BC is a heterogeneous disease but CTCs may be detected independently of the disease characteristics in 76% of patients at any time point during the course of the disease. This relatively high CTC occurrence in BC should be considered when planning the long-term patient monitoring.

     

Downregulated miR-367-3p,miR-548aq-5p,and miR-4710 in Human Whole Blood: Potential Biomarkers for Breast Cancer With Axillary Lymph Node Metastasis

BackgroundIncreasing studies have shown that microRNAs (miRNAs) have great diagnostic value in cancer. Axillary lymph node metastasis (ALNM) is closely related to the prognosis of breast cancer. However, it remains unknown whether miRNAs in whole blood could be promising biomarkers in breast cancer ALNM.MethodsAn miRNA microarray was used to screen potential differentially expressed miRNA candidates in whole blood of three breast cancer patients with ALNM and three without ALNM. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect candidate differentially expressed miRNAs in the whole blood of 109 breast cancer patients. Furthermore, bioinformatics analysis was carried to predict the potential targets and enriched pathway of miRNAs.ResultsQRT-PCR validated the fact that miR-367-3p, miR-548aq-5p and miR-4710 are downregulated in breast cancer with ALNM compared to it without ALNM. Receiver operating characteristic (ROC) curve analysis revealed that miR-367-3p, miR-548aq-5p and miR-4710 have good diagnostic values. Notably, the three-miRNA signature showed better predictive value, with an area under ROC curve (AUC) of 0.7414. Bioinformatics analysis revealed that the miRNAs could participate in a complex network and thus be involved in cancer-related pathways.ConclusionsOur findings support the potential of miR-367-3p, miR-548aq-5p and miR-4710 and the three-miRNA signature as biomarkers for breast cancer with ALNM.     

Mesenchymal marker and LGR5 expression levels in circulating tumor cells correlate with colorectal cancer prognosis

Purpose

The presence of circulating tumor cells (CTCs) has been found to correlate with colorectal cancer (CRC) prognosis, whereas epithelial-mesenchymal transition (EMT) in CTCs has been found to be associated with CRC metastasis. LGR5 is a known target of Wnt signaling and plays an important role in CRC development. The aim of this study was to assess the clinical relevance of EMT and LGR5 expression in CTCs from CRC patients.

Methods

Sixty-six CRC patients were included in this study. The detection and expression of EMT phenotypes in CTCs from these patients were assessed using CanPatrol? CTC enrichment and mRNA in situ hybridization (ISH), respectively. LGR5 expression in the CTCs was assessed using mRNA ISH.

Results

CTCs were detected in 86.4% (57/66) of the CRC patients included. Both the numbers of total CTCs and of CTCs displaying a mesenchymal phenotype (M+ CTCs) were found to significantly correlate with advanced disease stages and the occurrence of metastasis (p?<?0.05). An adjusted multivariate analysis also indicated that the number of M+ CTCs significantly correlated with the occurrence of metastasis (p?=?0.031). Additionally, we found that a high LGR5 expression level significantly correlated with the occurrence of metastasis (p?<?0.05). We also found that the presence of ≥ 6 CTCs or?≥?3 M+ CTCs per 5 ml blood significantly correlated with disease progression (p?<?0.05). Patients with ≥ 6 CTCs or?≥?3 M+ CTCs per 5 ml blood were found to exhibit poorer progression-free survival (PFS) and overall survival (OS) rates (p?<?0.05 in all cases). Using Cox regression analyses, we found that only total CTC numbers remained as independent prognostic factors for a worse PFS (p?=?0.043).

Conclusions

From our data we conclude that CTC numbers and EMT phenotypes may serve as prognostic markers for disease progression and metastasis in CRC patients. In addition, we conclude that LGR5 expression in CTCs may serve as a marker for CRC metastasis.

     

A Pilot Study Assessing the Incidence and Clinical Significance of Circulating Tumor Cells in Esophagogastric Cancers

BackgroundCirculating tumor cells (CTCs) have been found to be of clinical utility in predicting response to treatment and prognosis in several malignancies. Less is known of the prevalence and clinical relevance of CTCs in esophagogastric adenocarcinoma, with the available data arising from heterogeneous patient populations using varied detection methods.Patients and MethodsA pilot study was undertaken to assess the prevalence of CTCs in patients with advanced esophageal or gastric adenocarcinoma. Patients were eligible if they had advanced disease and either had received no prior therapy or had progressed after prior chemotherapy. Blood samples for CTC analysis were obtained at baseline and during the course of treatment. The CellSearch immunomagnetic CTC detection platform was used.ResultsTwenty-two patients with metastatic esophageal or gastric adenocarcinoma were enrolled. Eighteen received first-line EOX (epirubicin/oxaliplatin/capecitabine) chemotherapy (± panitumumab) and had baseline samples suitable for CTC analysis. At baseline, ≥ 2 CTCs were detected in 8 patients (44%). Overall tumor response rate was 60% in patients with < 2 CTCs and 37.5% in patients with ≥ 2 CTCs. Median progression-free and overall survival were 6.1 and 10.5 months and 5.2 and 6.1 months in the groups of patients with < 2 CTCs and ≥ 2 CTCs, respectively. The study was prematurely discontinued, owing to the withdrawal of commercial support.ConclusionThe incidence of CTCs in locally advanced or metastatic esophagogastric cancer may be clinically relevant. Investigation of the potential clinical utility of CTCs is warranted in a larger cohort of patients with esophagogastric cancer.