Value Of Circulating Tumor Cells Research Articles

946P - Predictive and prognostic value of tumor cells and circulating plasma free DNA in advanced epithelial ovarian carcinoma

Background: In recent years there has been increasing interest in the study of circulating tumor cells (CTC) and plama cell-free circulating DNA (cfDNA) as a new biomarker in neoplastic diseases. Our work aims to clarify its clinical role in ovarian epithelial carcinoma (OEC). Methods: A prospective, multicenter, observational study has been conducted for 3 years in patients with advanced or responsive OEC. The predictive value and prognosis of CTC and cfDNA have been determined for both progression-free survival (PFS) and overall survival (OS). Their values have been compared with a control group. CTC were analysed by the CellSearch method and cfDNA by ALU-sequences-based quantitative PCR using two primers (ALU115 and ALU247); cfDNA integrity was calculated by ALU247/ALU115 ratio. This study was approved by the Central Research Ethics Committee. Updated data are presented. Results: This study was conducted from November 2013 until December 2016. We recruited 88 patients, 15 benign tumors and 16 healthy subjects. Clinical characteristics were similar to other series, with a mean age of 57 years, 68.2% serous high grade subtype and 55.7% with stage IIIC. CTC were positive in 23.8% of patients (>1CTC/7,5mL). Levels of cfDNA were significantly elevated in patients than in the group of benign tumors and healthy control. CTC proved to be a negative prognostic factor of OS, with an average of 19.8 months versus 30.4 months (log-rank 4,649, p < 0.031). A high cfDNA value was also a negative prognostic factor for OS, with 22.5 months versus 28.7 months (log-rank 7.308, p < 0.007). The reduction in the integrity of cfDNA greater than 50% after chemotherapy was an early marker of resistance, with PFS of 6.4 months versus 16.0 months (HR 0.027, p < 0.02). The baseline value of cfDNA was also a negative prognostic factor for SLP, with 10 months versus 18 months (log-rank 7.233, p < 0.007). The baseline level of cfDNA and its integrity presented a predictive value of response to chemotherapy and the outcome of surgery, respectively. Conclusions: In summary, CTC and cfDNA in advanced ovarian epithelial carcinoma have been prognostic factors for survival. In addition, cfDNA has a predictive value of response. These two biomarkers may be useful in clinical practice. Legal entity responsible for the study: University Clinical Hospital Virgen Arrixaca - Murcia/ES Funding: None Disclosure: All authors have declared no conflicts of interest.

Clinicopathological and prognostic significance of circulating tumor cells in patients with head and neck cancer: a meta-analysis.

BackgroundMany studies have assessed the clinical use of circulating tumor cells (CTCs) in head and neck cancer, but the clinicopathological and prognostic significance of CTCs is still unclear.Materials and methodsTwo authors systematically searched the studies independently with keywords in PubMed, MEDLINE, EMBASE, Science Citation Index Expanded and Cochrane Library (from inception to February 2017). The estimated hazard ratio (HR), risk ratio (RR) and their 95% confidence intervals (95% CIs) were set as effect measures. All analyses were performed by STATA 12.0.ResultsA total of 17 studies were included in this meta-analysis. Positive CTCs were significantly associated with poor overall survival (HR =2.80, 95% CI: 1.34–5.86), disease-free survival (HR =3.86, 95% CI: 2.03–7.36) and progression-free survival (HR =3.31, 95% CI: 1.71–6.42). CTC-positive patients tend to have higher recurrence (RR =2.13, 95% CI: 1.26–3.59) and regional lymph node metastasis (RR =1.18, 95% CI: 1.02–1.36) rate and a more advanced tumor stage (RR =1.16, 95% CI: 1.03–1.32).ConclusionOur meta-analysis has confirmed the significant prognostic value of CTCs in head and neck cancer patients. The presence of CTCs could be used as a monitoring tool for tumor status of head and neck cancer, especially for the early detection of the tumor recurrence and progression, advanced disease and the node metastasis.

A comprehensive study of circulating tumour cells at the moment of prostate cancer diagnosis: biological and clinical implications of EGFR, AR and SNPs.

Circulating tumor cells (CTCs) have been recently accepted as prognostic markers in metastatic prostate cancer (PCa). However, very few studies have analyzed their role in early-stage PCa. The aim of this research is to study the value of CTCs at the moment of PCa diagnosis and to identify different subpopulations of CTCs. Patients with PSA value > 4 ng/ml and clinical suspicion of PCa were included. Samples were collected immediately before prostatic biopsy. CTCs were isolated by immunomagnetic technique using a multi-CK specific antibody. Molecular expression of EGFR and AR in the tissue was analysed by real-time PCR. Up to eight different SNPs in patients’ blood DNA were studied.In a total of 86 patients, the CTC detection rate was 18.6%. The sensitivity, specificity, positive and negative predictive values of CTCs to detect PCa was 14.2%, 78.4%, 31.2% and 57.4%, respectively. Up to 75% of CTC-positive patients were AR-negative. A direct association was found between the expression of AR in the prostatic tissue and the presence of CTCs in blood (p<0.05). We observed an inverse relation between the expression of EGFR in the tissue and the expression of AR in the CTCs. No significant association between SNPs and CTCs was found.The low detection rate of CTCs in early-stage PCa limits their role as a diagnostic marker. Nevertheless, we show that they may hide important prognostic information. Overexpression of AR in the prostate may facilitate cell dissemination.

Liquid biopsy in ovarian cancer: recent advances on circulating tumor cells and circulating tumor DNA.

Ovarian cancer remains the most lethal disease among gynecological malignancies despite the plethora of research studies during the last decades. The majority of patients are diagnosed in an advanced stage and exhibit resistance to standard chemotherapy. Circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) represent the main liquid biopsy approaches that offer a minimally invasive sample collection. Both have shown a diagnostic, prognostic and predictive value in many types of solid malignancies and recent studies attempted to shed light on their role in ovarian cancer. This review is mainly focused on the clinical value of both CTCs and ctDNA in ovarian cancer and, more specifically, on their potential as diagnostic, prognostic and predictive tumor biomarkers.

Circulating tumor cells detected by lab-on-a-disc: Role in early diagnosis of gastric cancer.

BackgroundThe use of circulating tumor cells (CTCs) as an early diagnostic biomarker and prognostic indicator after surgery or chemotherapy has been suggested for various cancers. This study aimed to evaluate CTCs in patients who underwent gastrectomy for gastric cancer and to explore their clinical usefulness in the early diagnosis of gastric cancer.MethodsA total of 116 patients with gastric cancer who underwent gastrectomy and 31 healthy volunteers were prospectively included between 2014 and 2015. Peripheral blood samples were collected before gastrectomy, and CTCs were examined using a centrifugal microfluidic system with a new fluid-assisted separation technique.ResultsAfter creating a receiver operating characteristic curve to identify the discriminative CTC value needed differentiate patients with gastric cancer from healthy volunteers, sensitivity and specificity were nearly optimized at a CTC threshold of 2 per 7.5 mL of blood. Of the 102 persons with a CTC level ≥2 per 7.5 mL of blood, 99 (97.1%) had gastric cancer, and of the 45 persons with a CTC level <2 per 7.5 mL of blood, 28 (62.2%) were healthy controls. Accordingly, the sensitivity and specificity for the differentiation of patients with gastric cancer from healthy controls were 85.3% and 90.3%, respectively. However, the presence of CTCs was not associated with any clinicopathologic features such as staging, histologic type, or mucin phenotype.ConclusionAlthough we could not prove the clinical feasibility of CTCs for gastric cancer staging, our results suggest a potential role of CTCs as an early diagnostic biomarker of gastric cancer.

Body mass index mediates the prognostic significance of circulating tumor cells in inflammatory breast cancer

BackgroundObesity (BMI≥30) may be an etiologic and prognostic factor in inflammatory breast cancer (IBC). We examined the relationship between BMI, pathologic complete response (pCR), and circulating-tumor-cell (CTC) levels in IBC. MethodsCohort included IBC patients diagnosed 2005–2015 who had neoadjuvant chemotherapy during a prospective trial on CTCs and pathologic review describing pCR. Chi-square, logistic regression, and Cox proportional hazards models were used to identify clinicopathologic associations with event-free survival (EFS). ResultsOf 73 patients, 61 (84%) had CTC values, 22 (30%) achieved a pCR, and 39 (53%) were obese. There was no difference between obese and non-obese patients for pCR rates (31% vs. 29%, p = 0.90) or presence of CTCs (23% vs. 26%, p = 0.80). Among non-obese patients, CTCs were associated with worse EFS (HR 11.69, p < 0.01), but among obese patients, there was no difference in EFS between those with and without CTCs. ConclusionsBMI mediates CTCs' prognostic significance in IBC.

The prognostic and diagnostic value of circulating tumor cells in bladder cancer and upper tract urothelial carcinoma: a meta-analysis of 30 published studies.

There are inconsistent conclusions in the association between circulating tumor cells (CTCs) and urothelial cancer (UC). We performed a meta-analysis to assess the prognostic and diagnostic value of CTCs in UC. We search Medline, Embase and Web of science for relevant studies. The study was set up according to the inclusion/exclusion criteria. 30 published studies with a total of 2161 urothelial cancer patients were included. Meta-analysis showed that CTC-positive was significantly associated with tumor stage (≤ II vs III, IV) (OR = 4.60, 95% CI: 2.34–9.03), histological grade (I, II vs III) (OR = 2.91, 95% CI: 1.92–4.40), metastasis (OR = 5.12, 95% CI: 3.47–7.55) and regional lymph node metastasis (OR = 2.47, 95% CI: 1.75–3.49). It was also significantly associated with poor overall survival (OS) (HR = 3.98, 95% CI: 2.20–7.21), progression/disease-free survival (PFS/DFS) (HR = 2.22, 95% CI: 1.80–2.73) and cancer-specific survival (CSS) (HR = 5.18, 95% CI: 2.21–12.13). Overall sensitivity and specificity of CTC detection assays were 0.35 (95% CI: 0.28–0.43) and 0.97 (95% CI: 0.92–0.99) respectively. In summary, our meta-analysis suggests that the presence of CTCs in the peripheral blood is an independent predictive indicator of poor outcomes for urothelial cancer patients. It can also be used as a noninvasive method for the confirmation of cancer diagnosis. More studies are required to further explore the role of this marker in clinical practice.

Combined peripheral natural killer (NK) cell and circulating tumor cell (CTC) enumeration to enhance prognostic efficiency in patients (pts) with triple-negative breast cancer (TNBC).

1105 Background: CTCs have emerged as an independent prognostic factor for metastatic breast cancer. However, the prognostic value of baseline CTCs regarding PFS in TNBC is still controversial, especially beyond first-line. We evaluated a novel combined NK cell/CTC detection system for enumeration to better understand the impact of cell count on prognosis in TNBC. Methods: 83 consecutive patients with metastatic TNBC were enrolled and received a new line of therapy (median=2, range: 1~5). Baseline circulating CTCs and NK cells were isolated and enumerated simultaneously prior to starting a new line of therapy using our previously published method targeting EpCAM (Bai et al. J Mater Chem B, 2014) and flow cytometry (antibodies against CD3-, CD45+, CD56+) respectively. The NK cell level was expressed as the proportion of total lymphocytes (%). Patients with normal to high NK cell proportion (≥ 9.15%) and &gt; 2 CTC/2 ml were defined as NK resistant CTC, the rest were defined as non-NK resistant CTC. Results: 33 out of 83 TNBCs had first-line of therapy. Pts with ≤ 2 CTC/2 ml had a significantly longer median PFS than those with &gt; 2 CTC/2 ml ( P = 0.028). The differences in median PFS were not significant ( P=0.110) for the entire group of pts with TNBC receiving different lines of therapy. Pts with non-NK resistant CTCs had a significantly longer median PFS than those with NK resistant CTCs ( P= 0.025), regardless of line of therapy. Conclusions: Including peripheral NK cell level into consideration can improve CTC prognostic efficiency in predicting PFS for TNBCs receiving different line of therapy. Further analysis of the unique biological features of NK-resistant CTCs and associated clinical consequence holds promise in guiding clinical practice. [Table: see text]

Dynamic evaluation of circulating tumour cells in patients with advanced gastric and oesogastric junction adenocarcinoma: Prognostic value and early assessment of therapeutic effects

BackgroundThe identification of dynamic biomarkers in advanced gastric and oesogastric junction adenocarcinoma (GOA) could help to tailor strategies for each patient. Enumeration of circulating tumour cells (CTCs) is approved by the US Food and Drug Administration in breast, colon and prostate cancer but is not in advanced GOA. Our study aims to establish the optimal threshold and the clinical significance of CTC count in advanced GOA before and during treatment. MethodsOne hundred six patients with untreated advanced GOA were included in the ancillary study of the PRODIGE 17-ACCORD 20 trial. CTCs were detected in the peripheral blood using the CellSearch system on day 0 (D0) and day 28 (D28). The prognostic value of CTCs at D0 and D28 was analysed by testing several thresholds. ResultsAt baseline, median CTC count was 1 (range, 0–415). While CTCs ≥1, 2 or 3 at D0 were all significantly associated with worse overall survival (OS) and progression-free survival (PFS), CTCs ≥2 were the optimal threshold, on D0 or D28. CTCs ≥2 at D28 were also predictive of disease control. Taking into account both D0 and D28 CTC count defined 3 groups (low/low, high/low and low-high/high) with significantly different PFS (p = 0.0002) and OS (p = 0.003). ConclusionQuantification of CTCs at baseline and during treatment may be a useful prognostic tool in advanced GOA, as it is associated with worse PFS and OS. A threshold ≥2 CTCs seems to have the best discriminant value. Change in CTC count between baseline and D28 could help to tailor treatment to each individual patient.

Detection of Circulating Tumor Cells and Circulating Tumor Microemboli in Gastric Cancer

PURPOSE: Gastric cancer studies indicated a potential correlation between circulating tumor cells (CTCs) in peripheral blood and tumor relapse/metastasis. The prevalence and significance of circulating tumor microemboli (CTM) in gastric cancer remain unknown. We investigated the prevalence and prognostic value of CTCs and CTM for progression-free survival (PFS) and overall survival (OS) in gastric cancer patients. METHODS:Eighty-one gastric cancer patients consented to provide 5ml of peripheral blood before systematic therapy. CTCs and CTM were isolated using isolation by size of epithelial tumor cells and characterized by cytopathologists. For 41 stage IV gastric cancer patients, CTM was investigated as a potential biomarker to predict prognosis. RESULTS:CTCs were detected in 51 patients; the average count was 1.81. In clinical stage I, II, III, and IV patients, the average CTC counts were 1.40, 0.67, 1.24, and 2.71, respectively. CTM were detected in 3 of 33 clinical stage I to IIIb patients, at an average of 0.12 (0-2). CTM were detected in 13 of 53 clinical stage IIIc to IV patients, at an average of 1.26 (0-22). In stage IV patients, CTM positivity correlated with the CA125 level. PFS and OS in CTM-positive patients were significantly lower than in CTM-negative patients (P<.001). CTM positivity was an independent factor for determining the PFS (P=.016) and OS (P=.003) of stage IV patients in multivariate analysis. Using markers of the epithelial-mesenchymal transition, single CTCs were divided into three phenotypes including epithelial CTCs, biphenotypic epithelial/mesenchymal CTCs, and mesenchymal CTCs. For CTM, CK−/Vimentin+/CD45− and CK+/Vimentin+/CD45− phenotypes were observed, but the CK+/Vimentin−/CD45− CTM phenotype was not. CA125 was detected in gastric cancer cell lines BGC823 and MGC803. CONCLUSIONS: In stage IV patients, CTM positivity was correlated with serum CA125 level. CTM were an independent predictor of shorter PFS and OS in stage IV patients. Thus, CTM detection may be a useful tool to predict prognosis in stage IV patients.

CIRCULATING TUMOR CELLS IN BREAST CANCER: FUNCTIONAL HETEROGENEITY, PATHOGENETIC AND CLINICAL ASPECTS

Each patient has a unique history of cancer ecosystem development, resulting in intratumor heterogeneity. In order to effectively kill the tumor cells by chemotherapy, dynamic monitoring of driver molecular alterations is necessary to detect the markers for acquired drug resistance and find the new therapeutic targets. To perform the therapeutic monitoring, frequent tumor biopsy is needed, but it is not always possible due to small tumor size or its regression during the therapy or tumor inaccessibility in advanced cancer patients. Liquid biopsy appears to be a promising approach to overcome this problem, providing the testing of circulating tumor cells (CTC) and/or tumor-specific circulating nucleic acids. Their genomic characteristics make it possible to assess the clonal dynamics of tumors, comparing it with the clinical course and identification of driver mutation that confer resistance to therapy. The main attention in this review is paid to CTC. The biological behavior of the tumor is determined by specific cancer-promoting molecular and genetic alterations of tumor cells, and by the peculiarities of their interactions with the microenvironment that can result in the presence of wide spectrum of circulating tumor clones with various properties and potentialities to contribute to tumor progression and response to chemotherapy and prognostic value. Indeed, data on prognostic or predictive value of CTC are rather contradictory, because there is still no standard method of CTC identification, represented by different populations manifesting various biological behavior as well as different potency to metastasis. Circulating clasters of CTC appear to have essentially greater ability to metastasize in comparison with single CTC, as well as strong association with worse prognosis and chemoresistance in breast cancer patients. The Food and Drug Administration (USA) has approved the CTC-based prognostic test for clinical application in patients with advanced breast cancer. Prospective clinical trials have demonstrated that measuring changes in CTC numbers during treatment is useful for monitoring therapy response in breast cancer patients. Molecular and genetic analysis of CTC gives the opportunity to have timely information on emergence of resistant tumor clones and may shed light on the new targets for pathogenetic antitumor therapy.

Research progress of circulating tumor cells in breast cancer

Breast cancer is the most common malignant tumor in female, and the recurrence and metastasis is the leading cause of death. Previous image evaluation of breast cancer could not detect the tumor at its early stage timely. The circulating tumor cells in the peripheral blood are like the seeds buried in the body, which cause the later recurrence and metastasis. So we can detect circulating tumor cells in peripheral blood to evaluate breast cancer progression. In present study, circulating tumor cells can be implied more than three ways in clinical application of breast cancer. They could be used to monitor the therapeutic effect in breast cancer therapy timely, to analyze molecular classification of breast cancer and conduct clinical drug sensitivity tests. Although the value of circulating tumor cells in breast cancer has been widely recognized, many factors limited their application in clinical. Thus, we conduct this review about the current progress of circulating tumor cells in breast cancer. Key words: Neoplastic cells, circulating; Breast neoplasms; Therapeutic uses; Review

The role of circulating tumor cells in evaluation of prognosis and treatment response in advanced non-small-cell lung cancer.

Non-small-cell lung cancer (NSCLC) lacks validated biomarkers to predict the prognosis and treatment response. This study investigated whether circulating tumor cells (CTCs) detectable could reminder high risk of distant metastasis, provide prognostic information, and early indicate the response to the conventional therapy in patients with advanced NSCLC. In this single-center prospective study, blood samples for CTC analysis were obtained from 59 patients with previously untreated, stage III or IV NSCLC both before and after administration of two cycles of chemotherapy. CTCs took in peripheral blood were measured by Cell Search detect technique. Carcino-embryonic antigen and count of metastatic sites were positively related to CTC count analyzed by multiple linear regression (P < 0.05). The median overall survival was 11.2 months (95% CI: 10.37-12.03months) for the baseline CTC ≥ 2 group compared with 8.3months (95% CI: 7.72-8.88months) for the CTC < 2 group (log-rank test P < 0.05). Similarly, patients with CTC ≥ 2 at baseline had a significantly shorter median PFS (4.3 months, 95% CI: 3.7-4.9months) compared with patients with CTC < 2 (6.2months, 95% CI: 5.59-6.82months) (log-rank test P < 0.05). For the disease control (stable disease, partial response, or complete response), group CTC value before treatment did not present difference with that after therapy compared by pared-samples T test (t = 1.455, P = 0.154), similar to the result of progressed group (progressive disease) (t = -0.987, P = 0.335). The CTC value of progressed group was higher than that of disease control group either at baseline or post chemotherapy. These data provide an evidence of positive correlation between CTC counts with CEA, as well as count of metastatic sites. Meanwhile, CTCs could be an effective predictor of distant metastasis and poor prognosis. In this study, CTCs are poorly related to treatment response. Whether CTCs could be a predictor of curative effect in advanced NSCLC should be validated by more researches in the future.

Association of changes in circulating cell-free plasma DNA (cfDNA) and circulating tumor cells (CTC) during treatment with clinical outcome from olaparib in castration-resistant prostate cancer (CRPC): Exploratory analyses from the TOPARP-A trial.

141 Background: Response biomarkers are needed to optimize treatment switch decisions in CRPC patients. CTC and cfDNA may have clinical utility as response biomarkers; we studied them during olaparib treatment in a phase II trial in CRCP (Mateo et al NEJM 2015). Methods: CTC were enumerated using CellSearch (Jannsen Diagnostics) and cfDNA was extracted with the QIASymphony circulating DNA kit (Qiagen) from blood samples taken at baseline, 4- and 8-weeks (wk) of therapy. Radiological progression-free survival (rPFS) was defined as time from starting treatment to progression by RECIST 1.1, bone scan (PCWG2) or death. Overall survival (OS) was defined as time from starting treatment to death. CTC changes were categorized based on conversion from ≥ 5 to &lt; 5 CTC/7.5ml blood and on ≥ 30% decline (Lorente et al Eur Urol 2016). cfDNA changes were evaluated as percentage change from baseline (continuous and binary). The prognostic value of CTC and cfDNA changes were assessed by Landmark analysis and Cox models with time-varying covariates; p-value &lt; 0.01 were considered significant to account for multiple tests. Results: Overall, 13/47 (28%) and 16/42 (38%) evaluable patients had a CTC conversion at 4- and 8-wk respectively. A CTC conversion after 4-wk of olaparib associated with longer rPFS (median 8.9 vs 2.7 months [m], p = 0.001); a similar association was found at 8-wk. A 30% CTC decline at 4-wk also associated with longer rPFS (median 4.4 vs 2.6 m, p = 0.004). CTC conversion as a time-varying covariate associated with longer OS (HR 0.26, 95%CI 0.14-0.50, p &lt; 0.001). Median baseline cfDNA was 31.6 ng/ml (IQR 19.4-57.1); 46 and 42 patients were evaluable for cfDNA changes at 4- and 8-wk. Percentage changes in cfDNA at 4- and 8- wk associated with rPFS (HR 1.01 and 1.005; p = 0.005 and 0.002 respectively) but association with OS was not significant. cfDNA declines ≥ 50% at 8- wk associated with longer rPFS (median 8.9 vs 2.7 m, p = 0.007) and OS (17.0 vs 10.1 m, p = 0.004). Conclusions: Decreases in CTC counts and cfDNA concentration associate with improved outcome from olaparib (rPFS, OS) in the TOPARP-A trial. Clinical trial information: NCT01682772.

Abstract P1-01-16: Circulating tumor cells (CTCs) biomarker evaluation from patients with metastatic breast cancer (MBC) utilizing the TargetSelectorTM platform

Abstract Background: Circulating blood biomarkers represent the promise of non-invasive, real-time surrogates for tumor tissue-based biomarkers as well as afford monitoring opportunities over the course of therapy as tumors evolve and acquire resistance to treatment. Circulating tumor cells (CTCs) are cells that disseminate from tumors and can be identified in peripheral blood. CellSearch® is an FDA approved methodology for detecting and enriching this rare CTC population. The prognostic value of CTCs has been established, however the potential to characterize biomarkers on CTCs to inform treatment decisions remains an active area of investigation. Among the emerging platforms for detection and characterization of CTCs is the TargetSelectorTM system. While CellSearch® is limited to capture and detection of epithelial derived CTCs based on EpCAM and cytokeratin (CK) respectively, Biocept's TargetSelectorTM platform utilizes a novel microfluidic system for CTC enrichment based on an antibody capture cocktail that allows for enumeration of CTCs with variable phenotypes (including CK- CTCs). The CTCs are captured in transparent microfluidic chambers and cells can be viewed in situ by fluorescence microscopy and analyzed via immunocytochemistry (ICC), fluorescence in situ hybridization (FISH) and PCR analyses. Methods: Sixty-one patients with metastatic breast cancer consented and provided blood for utilization in the TargetSelectorTM platform. Based on the molecular analysis of tissue biopsies, 92% of these patients had ER+ breast tumors. Biomarker expression on captured CTCs was determined by ICC for ER and by FISH for HER2. Concordance between these results and biomarker expression on archival tumor tissue from these patients was calculated. Results: CTCs were detected in 60 of 61 patient blood samples (range 2–4471); 68% had both CK+ and CK- CTCs, and 32% had only CK- CTCs. None had only CK+ CTCs. Of those with CK+ CTCs, concordance for ER expression between the tissue and blood analyses was 85% (35/41). Concordance was much lower for patients with only CK- CTCs (32%, 6/19). Concordance for HER2 amplification in CK+ patients was 93% (38/41), and 68% (13/19) in CK- patients. For this study, the liquid biopsy was obtained over an extended period of time from when solid tumor ER and HER2 assessments were obtained; this latency period may have influenced concordance levels. For HER2, there was a significantly longer time interval between biopsies for non-concordant than for concordant pairs of samples (58.1 ± 19.7 vs. 30.9 ± 4.3 months) regardless of the CK status. No such difference was seen for the ER analysis. Conclusions: In this exploratory analysis of 61 patients with MBC, we observed a high rate of detectable CTCs as well as CTC concordance for ER (85%) and HER2 amplification (93%) for patients who had CK+ CTCs. Concordance was less if patients only had CK- CTCs. This may be attributable to heterogeneity in the breast cancer phenotypes associated with these CK- CTCs in addition to inherent issues with testing cell surface markers in this population of cells. The variable latency between the between the collection of tissue and blood samples for these analyses may account for the discrepancies observed. Citation Format: Hamilton EP, Yardley DA, Burris III HA, Shastry M, Huynh L, Bhikha L, Singh VM. Circulating tumor cells (CTCs) biomarker evaluation from patients with metastatic breast cancer (MBC) utilizing the TargetSelectorTM platform [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P1-01-16.

Clinical significance of circulating tumor cells in patients with small-cell lung cancer.

This study investigated the correlation of the presence of circulating tumor cells (CTCs) with clinical characteristics, and the predictive value of CTCs for progression-free survival (PFS) in patients with small-cell lung cancer (SCLC). Samples were obtained from 42 patients with SCLC before and after the first cycle of chemotherapy. CTCs were quantitated by negative immunomagnetic enrichment and immunocytochemistry using anti-CD45 and anti-pancytokeratin antibodies. CTCs were positive (≥2) in 76.19% of patients with SCLC and negative in the control group. The presence of CTCs was positively correlated with 6 clinical characteristics. PFS was 6.055 and 10.670 months for patients with ≥2 and <2 CTCs/7.5 mL of blood before chemotherapy; after chemotherapy PFS was 4.862 and 10.535 months, respectively. This study showed that both baseline CTC numbers and the change in CTC numbers after 1 cycle of chemotherapy are significant prognostic factors of PFS for SCLC.

Abstract S3-01: IMENEO: International MEta-analysis of circulating tumor cell detection in early breast cancer patients treated by NEOadjuvant chemotherapy

Abstract Background We performed an international meta-analysis of individual patient data to assess the clinical validity of circulating tumor cell (CTC) count in non-metastatic breast cancer (BC) patients (pts) treated by neoadjuvant chemotherapy (NCT). Methods A protocol pre-specified the study objectives. We performed a literature &amp; abstracts search up to Dec 2014, then contacted all centers deemed to have eligible data (published or not): early BC pts treated with NCT with CTC count by CellSearch®. The primary endpoint was overall survival (OS); secondary endpoints included distant disease-free survival (DDFS), locoregional relapse-free interval (LRFI) and pathological complete response (pCR). Non-overlapping CTC time points were: baseline (5-0 weeks before NCT), 1-8 weeks after NCT start, 5-0 weeks before surgery and 1-52 weeks after surgery. We used Cox regression models, stratified by study, and the landmark method to establish the prognostic value of CTC count/changes during treatment and survival. Results We collected 2,156 individual pt data from 21 studies and 16 centers worldwide. With ≥1/≥2/≥5 CTC/7.5ml as thresholds, CTC positivity rate was 25/13/6% at baseline, 17/6/3% after NCT start, 15/5/1% before surgery and 11/4/1% after surgery (decrease, p&amp;lt;0.0001). Before NCT, ≥1 CTC was found in 19%, 22%, 24%, 29% and 41% of cT1, T2, T3, T4a-c and T4d BC, respectively (p&amp;lt;0.0001) and was also marginally associated with hormone-receptors negativity (p=0.04). Later CTC detection rates were not associated with any of the baseline characteristics. pCR (assessed in 2,072 pts; ypT0/isN0 used as pCR definition in 92% of pts) was observed in 24% of pts but was not associated with CTC count, at any time point. 301, 418 and 157 events were reported for OS, DDFS and LRFI, respectively. In univariate analyses, ≥1 CTC at baseline was a prognostic factor for OS (HR=2.6 [1.9-3.4], p&amp;lt;0.0001), DDFS (HR=2.4 [1.9-3.1], p&amp;lt;0.0001) and -importantly- for LRFI (HR=1.8 [1.2-2.7], p=0.001). Similar results were obtained using other thresholds (≥2 &amp; ≥5 CTC) and/or later time points (after NCT start, before surgery). There was no interaction between the prognostic impact of CTC count and tumor subtypes. Although rare, pts with persistently elevated CTC count (≥1CTC) before NCT and before surgery (5% of pts) had a worse OS than patients with persistently null CTC count (HR=6.2 [3.4-11], p&amp;lt;0.0001). Finally, in multivariate analyses, baseline CTC detection (whatever the CTC threshold used : ≥1/≥2/≥5 CTC) was an independent prognostic factor for OS, DDFS and LRFI, together with pCR, cT, cN and tumor subtype, (e.g. for OS: CTC≥2 HR=4.2 [3.0-5.9] p&amp;lt;0.0001, No pCR HR=6.2 [3.7-11] p&amp;lt;0.0001, cT4d HR=2.6 [1.1-6.6] p=0.02, cN+ HR=1.7 [1.2-2.4] p=0.003, triple negative BC HR=3.2 [2.1-5.1]). Similar results were obtained with later time points (after NCT start, before surgery). Conclusions Our study demonstrates with the highest level of evidence that CTCs are a prognostic biomarker in early BC treated by NCT. This impact was independent to that of pCR and was observed on OS, DDFS and also -for the first time- on LRFI. CTC count can usefully complement standard prognostic factors and pCR to improve the prognostication of early BC pts. Citation Format: Bidard F-C, Michiels S, Mueller V, Riethdorf S, Esserman LJ, Lucci A, Naume B, Horiguchi J, Gisbert-Criado R, Sleijfer S, Toi M, Garcia-Saenz JA, Hartkopf A, Generali D, Rothe F, Smerage J, Muinelo L, Stebbing J, Viens P, Magbanua M, Hall CS, Engebråtenm O, Takata D, Vidal-Martínez J, Onstenk W, Fujisawa N, Diaz-Rubio E, Taran F-A, Cappelletti MR, Ignatiadis M, Name N, Proudhon C, Wolf D, Bowman Bauldry J, Borgen E, Nagaoka R, Carañana V, Kraan J, Maestro M, Brucker SY, Weber K, Reyal F, Amara D, Gopalkrishna Karhade M, Ruud Mathiesen R, Tokiniwa H, Llombart-Cussac A, D'Hollander K, Cottu P, Park JW, Loibl S, Pierga J-Y, Pantel K. IMENEO: International MEta-analysis of circulating tumor cell detection in early breast cancer patients treated by NEOadjuvant chemotherapy [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr S3-01.

Abstract P1-01-15: DETECT V - Detection of the expression of estrogen receptor and human epidermal growth factor receptor 2 on circulating tumor cells from metastatic breast cancer patients

Abstract Background: The prognostic relevance of circulating tumor cells (CTCs) in metastatic breast cancer (MBC) has constantly been shown in several clinical studies. Nevertheless, the predictive value of CTCs still remains unclear. The DETECT study concept evaluates the impact of CTCs on therapeutic decisions. Patients with hormone-receptor positive, HER2-positive MBC participating in DETECT V are randomized to a dual HER2 targeted therapy (Trastuzumab and Pertuzumab) combined with either endocrine therapy or chemotherapy. Based on the expression of Estrogen Receptor (ER) and Human Epidermal Growth Factor Receptor 2 (HER2) on detected CTCs, an “Endocrine Responsiveness Score (ERS)” is calculated for prediction of endocrine treatment response. Aim of this project was the optimization of staining procedures using immunofluorescence. Methods: CTCs are enumerated and characterized using CELLSEARCH® CXC Kit which is based on the immunomagnetic enrichment of EPCAM-positive cells and immunofluorescent characterization of these cells. Comparison of staining intensities in different cell lines with known expression of ER and HER2 was used to validate the staining protocol. To validate ER-staining, staining intensities of the following cell lines were compared: SKBR-3 (negative control), MCF-7 (positive control) and T47D (intermediate expression). HER2-staining was validated comparing staining intensities of SKBR3 (positive control), MDA-MB 453 (intermediate expression) as well as MCF-7 (negative control). Results: Staining intensities were divided into the following categories: negative, weak, moderate and strong. Signal intensities using CELLSEARCH® system correlated with the known expression of HER2 and ER in the different cell lines. No expression of ER could be shown in SKBR3 cells, T47D cells showed weak expression of ER. MCF-7 cells showed moderate to strong expression of ER but no expression of HER2. MDA-MB 453 and SKBR3 showed moderate to strong and strong expression of HER2, respectively. Expression of HER2 and ER could be depicted accurately using CELLSEARCH® CXC Kit. Results were validated in a ring test in all participating laboratories. First patient samples are currently analyzed. Conclusion: Establishment and validation of the ERS by characterizing CTCs within the framework of the DETECT V study pursues the objective of assessing the potential benefit of endocrine therapy. Validation results and first patient results will be shown. Citation Format: Fehm T, Meier-Stiegen F, Riethdorf S, Schramm A, Polasik A, Niederacher D, Rack B, Taran F-A, Müller V, Janni W, Huober J. DETECT V - Detection of the expression of estrogen receptor and human epidermal growth factor receptor 2 on circulating tumor cells from metastatic breast cancer patients [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P1-01-15.

Detection of circulating tumor cells (CTC) using a non-EpCAM-based, high-definition, single-cell assay in advanced hepatocellular carcinoma (HCC) for patients enrolled on phase I and II trials of sorafenib plus temsirolimus.

311 Background: Noninvasive biomarkers are needed for diagnosis, prognosis, and molecular profiling of HCC due to scarcity of tumor tissue and heterogeneous tumor biology. CTC are detectable in metastatic HCC using methods which enrich for expression of the cell adhesion molecule, EpCAM. Because a subset of HCC does not express EpCAM, however, non-EpCAM enrichment methods are needed for CTC studies in HCC. Methods: This CTC cohort was derived from patients enrolled on multicenter phase 1 and 2 trials of the combination of sorafenib plus temsirolimus in advanced HCC (NCT01008917, NCT01687673) with approval and funding from the National Comprehensive Cancer Network (NCCN) Oncology Research Program. Eligibility required histologic diagnosis of incurable HCC with no prior systemic therapy. All patients in the cohort were treated at the recommended phase 2 dose of sorafenib 200 mg PO BID and temsirolimus 10 mg IV weekly. Whole blood samples were collected at baseline and on treatment. CTC were enumerated and analyzed cytomorphologically using a high-definition, single cell assay without EpCAM enrichment and blinded to clinical outcomes. Results: The CTC cohort was comprised of 36 patients (phase 1 n = 9, phase 2 n = 27). Characteristics: male 89%; white 64%, Asian/PI 28%, black 6%; HBsAg+ 31%, HCV+ 44%; Child Pugh A 92%, B7 8%; BCLC C 83%; tumor vascular invasion 47%; median AFP 74 ng/mL. Median OS from start of treatment was 392 days (95% CI: 214, 569). CTC ≥ 1/mL were detectable at baseline in 23/36 (64%) overall (95% CI: 47%, 80%), with similar findings in the phase 1 (56%) and phase 2 (67%) subsets. There was no significant relationship between baseline CTC values ≥ 1, 2, or 5/mL and overall survival (OS) on univariate analysis. Analyses of CTC relationship to clinical characteristics and time to progression, changes on treatment, and multivariable analysis for relationship to OS will be presented. Conclusions: CTC were detected in over 60% of patients in this advanced HCC clinical trial cohort using a non-EpCAM, high-definition single cell assay, suggesting future potential for noninvasive molecular profiling of HCC.

Circulating tumor microemboli (CTM) and vimentin+ circulating tumor cells (CTCs) detected by a size-based platform predict worse prognosis in advanced colorectal cancer patients during chemotherapy

BackgroundCirculating tumor cells (CTCs) detected in peripheral blood (PB) of cancer patients can be identified as isolated CTCs and circulating tumor microemboli (CTM). This study aimed to evaluate the prognostic value of CTM detection and CTC phenotype in advanced colorectal cancer (CRC) patients during chemotherapy.MethodsA size-based platform for CTC isolation was applied. PB samples (5 ml) from 98 advanced CRC patients during 2–6 cycles chemotherapy were collected for CTC detection, and CTC count was correlated to patient’s clinicopathological characteristics and clinical outcome. And CTC phenotype was measured by immunofluorescent staining and evaluate the predictive significance on survival in 32 CTCs-positive patients with advanced CRC.ResultsForty-eight of 98 patients were CTCs-positive, including 18 CTM-positive patients, and CTC detection was positively correlated with lymphatic invasion (P = 0.049), TNM stage (P = 0.023), and serum CEA level (P = 0.014). Moreover, Kaplan–Meier survival and Cox regression analyses revealed that the presence of CTCs was an independent factor for poor PFS and OS (P < 0.05) in advanced CRC patients during chemotherapy, and CTM-positive patients had shooter survival than isolated CTCs-positive patients (P < 0.05). Furthermore, patients with vimentin+ isolated CTCs/CTM had shorter PFS and OS compared with CK+ CTCs (P < 0.05).ConclusionsThis study provided evidence that the presence of CTCs was positively correlated with poor prognosis, and furthermore, CTM and vimentin+ CTCs predicted poorer survival, which indicated that CTM and vimentin+ CTCs detected by a sensitive platform could be used to improve prognostic value of CTCs in advanced CRC patients under treatment.