Value Of Circulating Tumor Cells Research Articles

Circulating tumor cells et al.: towards a comprehensive liquid biopsy approach in breast cancer.

Liquid biopsy has emerged as a crucial tool in managing breast cancer (BC) patients, offering a minimally invasive approach to detect circulating tumor biomarkers. Until recently, the majority of the studies in BC focused on evaluating a single liquid biopsy analyte, primarily circulating tumor DNA and circulating tumor cells (CTCs). Despite the proven prognostic and predictive value of CTCs, their low abundance when detected using enrichment methods, especially in the early stages, poses a significant challenge. It is becoming evident that combining diverse circulating biomarkers, each representing different facets of tumor biology, has the potential to enhance the management of patients with BC. This article emphasizes the importance of considering these biomarkers as complementary/synergistic rather than competitive, recognizing their ability to contribute to a comprehensive disease profile. The review provides an overview of the clinical significance of simultaneously analyzing CTCs and other biomarkers, including cell-free circulating DNA, extracellular vesicles, non-canonical CTCs, cell-free RNAs, and non-malignant cells. Such a comprehensive liquid biopsy approach holds promise not only in BC but also in other cancer types, offering opportunities for early detection, prognostication, and therapy monitoring. However, addressing associated challenges, such as refining detection methods and establishing standardized protocols, is crucial for realizing the full potential of liquid biopsy in transforming our understanding and approach to BC. As the field evolves, collaborative efforts will be instrumental in unlocking the revolutionary impact of liquid biopsy in BC research and management.

Abstract 7493: Real-time monitoring of CTC number, biomarker expression, and kinetics as prognostic and predictive markers in patients with GI malignancies

Abstract Background: Most of the utility in the clinic regarding liquid biopsies has been around circulating tumor DNA (ctDNA). ctDNA analysis provides valuable insight into mutational burden and therapeutic targets, but the speed and cost of analysis are prohibitive especially when it comes to serial testing. Using a cohort of patients with advanced or metastatic gastrointestinal (GI) cancers, we herein show the feasibility of using circulating tumor cell (CTC) enumeration, biomarker analysis, and kinetics as complementary to or as an alternative to ctDNA testing. Methods: Using the RareCyte platform, nucleated cells from blood were collected, spread on slides, stained, and imaged with CyteFinder® for CTC identification (CK/EpCAM+, CD45-) and quantification of biomarkers (PD-L1/HER2 or Ki67/EGFR). CTC enumeration (CLIA-accredited CTC test) and biomarker status for each patient were tracked over time and compared with ctDNA mutational analysis and tumor burden. Results: CTCs were detected in 60/70 samples (86%) and in 36/38 unique patients (95%). On average, more CTCs were detected in baseline samples from colorectal cancer patients compared to other subtypes (Table 1). Moreover, multiple patients showed pronounced changes in CTC number that correlated with treatment response, as well as strong correlation between CTC and tissue biomarker expression e.g. HER2. Rapid decline and/or CTC clearance was seen as early as a few days into starting effective systemic therapy. The average turnaround time for CTC testing was 5-7 days. Table 1. CTC values in initial baseline blood draw sample Gastrointestinal cancer subtype # of patients Average # of CTCs per 7.5mL blood Median # of CTCs per 7.5mL blood Minimum # of CTCs per 7.5mL blood Maximum # of CTCs per 7.5mL blood Colorectal carcinoma 27 84 3 0 1378 Cholangiocarcinoma 3 8 3 1 23 Pancreatic cancer 3 2 2 0 3 Hepatocellular carcinoma 2 4 5 0 10 Gallbladder cancer 1 6 6 3 8 Appendix cancer 1 2 2 1 3 Anal cancer 1 0 0 0 0 Conclusions: Our study emphasizes that real-time monitoring of CTCs is complementary to traditional tumor burden monitoring via imaging and ctDNA analysis. CTCs offer multiple unique advantages vs ctDNA including early response assessment within days of initiating therapy, a substantial decrease in cost of serial testing, and the ability to assess clinically relevant cell surface protein biomarkers e.g. HER2 and PD-L1. Citation Format: Erin G. Bayer, Brock M. Bartels, Rachel N. Ponting, Areeb Lutfi, Maaz K. Afghan, Arturo B. Ramirez, Pashtoon M. Kasi. Real-time monitoring of CTC number, biomarker expression, and kinetics as prognostic and predictive markers in patients with GI malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7493.

Abstract 7500: Distribution and prognostic significance of circulating tumor cells (CTCs) and tumor-derived extracellular vesicles (tdEVs) in patients (pts) with lobular metastatic breast cancer (MBC)

Abstract Background: Invasive lobular carcinoma (ILC) has distinctive clinical and molecular features compared with invasive ductal carcinoma (IDC). CTCs and tdEVs are independent prognostic factors in MBC, however, liquid biopsy studies focusing on ILC are to date scarce. Higher CTC levels have been observed in ILC than IDC. One study including 28 ILC pts suggested the use of higher CTC cutoff for prognosis stratification. The clinical significance of tdEVs in ILC is unexplored. This study aimed to assess differences in the distribution and prognostic value of CTCs and tdEVs between ILC and IDC. Methods: Blood samples were collected from 304 pts with MBC before starting a new line of therapy at Northwestern University (Chicago, IL) between 2016 and 2021 (NU16B06 trial). Blood was processed with the CellSearch® system. The ACCEPT software was applied to CellSearch® images to automatically enumerate CTCs and tdEVs. Association of CTCs and tdEVs with overall survival (OS) was tested in ILC and IDC pts. A count of CTCs ≥ 5 was considered high. The cutoff levels for tdEVs were < 20 (low), 20-79 (intermediate), and ≥ 80 (high), as previously reported. Additional exploratory cutoffs for CTCs (≥ 20, ≥ 50, ≥ 80, and ≥ 100) and tdEVs (based on quartiles of tdEV distribution in the overall population and ILC) were also evaluated. Results: Of the 304 pts, 47 had ILC while 257 had IDC. 56% of pts received first-line therapy. Median CTC count was 8 [interquartile range (IQR) 2-35] in ILC and 1 (IQR 0-8) in IDC pts (P <.001). A significantly higher median tdEVs count was observed in ILC (36; IQR 18-115) than in IDC (11; IQR 2-86) pts (P =.002). High levels of CTCs (≥5) were not associated with OS in ILC (P =.88), even when higher cutoffs (≥ 20, ≥ 50, and ≥ 80) were used. Only the detection of ≥ 100 CTCs was significantly associated with OS in ILC pts (median OS 36 vs 10 months; HR 3.4; P =.028). In contrast, the prognostic effect of CTCs ≥ 5 was strong in IDC and maintained for all cutoffs. Similarly, while the prognostic value for tdEVs was confirmed for IDC using both 20-80 and exploratory cutoffs (3-105 and 18-155), no association between tdEV levels and OS was observed in ILC regardless of the cutoffs used. Conclusion: ILC is associated with higher CTCs and tdEVs compared to IDC, probably due to the impaired cell-cell adhesion that characterizes ILC. Unlike IDC, the prognostic effect for the established cutoff of ≥ 5 CTC was not observed in ILC. A higher CTC cutoff could be considered for this subtype of BC. For tdEV no association with OS was observed, regardless of the cutoff used. These findings emphasize that ILC is a distinct entity also in the liquid biopsy features. Because of the low prevalence of ILC, research efforts should be directed at combining data from ILC pts to better characterize this subtype of BC and understand the reason behind the different associations with OS as compared to IDC. Citation Format: Eleonora Nicolo, Lorenzo Gerratana, Lorenzo Foffano, Laura Munoz-Arcos, Mara S. Serafini, Maroua Manai, Letizia Pontolillo, Nadia Bayou, Elisabetta Molteni, Amanda K. Strickland, Caterina Gianni, Youbin Zhang, Paolo D'Amico, Andrew A. Davis, Jeannine Donahue, Huiping Liu, William J. Gradishar, Ami N. Shah, Giuseppe Curigliano, Carolina Reduzzi, Massimo Cristofanilli. Distribution and prognostic significance of circulating tumor cells (CTCs) and tumor-derived extracellular vesicles (tdEVs) in patients (pts) with lobular metastatic breast cancer (MBC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7500.

Circulating Tumor Cells: How Far Have We Come with Mining These Seeds of Metastasis?

Circulating tumor cells (CTCs) are cancer cells that slough off from the tumor and circulate in the peripheral blood and lymphatic system as micro metastases that eventually results in macro metastases. Through a simple blood draw, sensitive CTC detection from clinical samples has proven to be a useful tool for determining the prognosis of cancer. Recent technological developments now make it possible to detect CTCs reliably and repeatedly from a simple and straightforward blood test. Multicenter trials to assess the clinical value of CTCs have demonstrated the prognostic value of these cancer cells. Studies on CTCs have filled huge knowledge gap in understanding the process of metastasis since their identification in the late 19th century. However, these rare cancer cells have not been regularly used to tailor precision medicine and or identify novel druggable targets. In this review, we have attempted to summarize the milestones of CTC-based research from the time of identification to molecular characterization. Additionally, the need for a paradigm shift in dissecting these seeds of metastasis and the possible future avenues to improve CTC-based discoveries are also discussed.

Prognostic Significance of Excision Repair Cross-Complementation Group 1 on Circulating Tumor Cells for Nasopharyngeal Carcinoma.

Liquid biopsy, including the detection of circulating tumor cells (CTCs), has emerged as a promising tool for cancer diagnosis and monitoring. However, the prognostic value of CTCs in nasopharyngeal carcinoma (NPC) remains unclear due to the lack of phenotypic characterization. The expression of Excision Repair Cross-Complementation Group 1 (ERCC1) and CTCs epithelial-mesenchymal transition (EMT) have been associated with treatment efficacy. In this study, we aimed to evaluate the prognostic significance of ERCC1 expression on CTCs and their EMT subtypes before treatment in NPC. We retrospectively analyzed 108 newly diagnosed locally advanced NPC patients who underwent CanPatrol™ CTC testing between November 2018 and November 2021. CTCs were counted and classified into epithelial, epithelial-mesenchymal hybrid, and mesenchymal subtypes. ERCC1 expression was divided into negative and positive groups. Clinical features and survival outcomes were analyzed. The positive rate of CTCs was 92.6% (100/108), with an ERCC1 positivity rate of 74% (74/100). Further analysis of the subtypes showed that positive ERCC1 on mesenchymal CTCs was associated with a later N stage (P = .01). Positive ERCC1 expression was associated with poor overall survival (OS; P = .039) and disease-free survival (DFS; P = .035). Further analysis of subtypes showed that the positive ERCC1 on mesenchymal-type CTCs was associated with poor OS (P = .012) and metastasis-free survival (MFS; P = .001). Our findings suggest that ERCC1 expression on CTCs may serve as a new prognostic marker for NPC patients. Evaluating CTCs subtypes may become an auxiliary tool for personalized and precise treatment.

Advances in circulating tumor cells for early detection, prognosis and metastasis reduction in lung cancer.

Globally, lung cancer stands as the leading type of cancer in terms of incidence and is the major source of mortality attributed to cancer. We have outlined the molecular biomarkers for lung cancer that are available clinically. Circulating tumor cells (CTCs) spread from the original location, circulate in the bloodstream, extravasate, and metastasize, forming secondary tumors by invading and establishing a favorable environment. CTC analysis is considered a common liquid biopsy method for lung cancer. We have enumerated both in vivo and ex vivo techniques for CTC separation and enrichment, examined the advantages and limitations of these methods, and also discussed the detection of CTCs in other bodily fluids. We have evaluated the value of CTCs, as well as CTCs in conjunction with other biomarkers, for their utility in the early detection and prognostic assessment of patients with lung cancer. CTCs engage with diverse cells of the metastatic process, interfering with the interaction between CTCs and various cells in metastasis, potentially halting metastasis and enhancing patient prognosis.

Kinetics and Biology of Circulating Tumor Cells (CTCs) and Measurable Residual Disease (MRD): Two Dynamic High-Risk Clones in Multiple Myeloma (MM)

BACKGROUND CTCs and MRD are two small clones associated with a high risk of progression in smoldering MM (SMM) and post treatment relapse in active MM. Despite growing knowledge about their clinical significance and biological features, these questions remain unanswered: 1) What is the prognostic value of CTC and MRD kinetics? 2) Is MM dissemination and on-treatment resistance driven by specific genomic alterations? 3) Are CTCs and MRD genetically related? AIM Analyze the kinetics and molecular features of CTCs and MRD in SMM and MM patients. METHODS CTC kinetics were investigated using NGF in 164 SMM patients with ≥3 assessments collected every 6 months in the iMMunocell study. MRD kinetics were investigated using NGF in 277 MM patients with ≥4 assessments in BM aspirates after induction, transplant, consolidation, and yearly during maintenance (GEM2012MENOS65-GEM2014MAIN trials). Unsupervised clustering based on CTC and MRD kinetics was performed using CONNECTOR. Molecular features of BM tumor cells at diagnosis were compared to paired CTCs in 94 SMM and MM patients and to paired MRD in 96 MM patients. In 10 cases, genomic data from tripartite BM tumor cells, CTCs, and MRD was available. Tumor samples were isolated based on patient-specific aberrant phenotypes using FACS. Whole exome and RNA-seq were performed in 343 and 217 samples, respectively. RESULTS Based on the assessment of CTCs in 754 PB samples, SMM patients clustered into 3 groups: sustained undetectable (n = 20), stable (n = 71) and evolving (n = 73) CTC levels. With a median follow-up of 2 years, the respective 2y progression-free rates were 100%, 94% and 81% ( P =.003). Risk stratification based on CTC kinetics (C-index 0.68) outperformed a single baseline assessment (C-index 0.61) and, along with the IMWG 20/2/20 model, had independent prognostic value (HR 3.4; P = .01). Patients with high risk cytogenetics showed more frequently evolving CTC levels. Studying genetic drivers of tumor egress, we found CTCs to closely resemble BM tumor cells (88% concordance at copy number [CNA] and somatic mutational [SNV] level). No recurrent cell-specific SNV or CNA was identified. CTC's transcriptional profile revealed 268 DEGs enriched in interferon alpha, gamma response and cell-adhesion pathways. Next, the prognostic value of MRD dynamics was analyzed according to 1,759 assessments, identifying 3 patient clusters: sustained undetectable (n = 100), stable (n = 104), and evolving (n = 73) MRD levels. Respective 6-year rates of PFS after consolidation were 90%, 75% and 5% ( P <.001), and of OS were 94%, 95% and 66% ( P<.001). Risk stratification based on MRD kinetics (C-index 0.82) outperformed a single MRD assessment before maintenance (C-index 0.62), the R-ISS (C-index 0.57) and R2-ISS (C-index 0.61) at diagnosis. MRD cells showed heterogeneous genomic evolution (60% median concordance) diverging from BM tumor cells at diagnosis due to de novo CNA and SNV that emerged at a subclonal level, particularly after transplant. After induction, MM driver mutations were detected or remained clonal in 13/19 (68%) MRD, whereas after transplant, this was true for only 5/13 (38%) MRD. No recurrent cell-specific SNV or CNA was identified. The transcriptional signature of MRD was treatment dependent, with common molecular features of on-treatment resistance found in all 96 patient-paired samples analyzed after various treatment stages. The 334 DEGs were commonly enriched in KRAS signaling hallmarks. Genomic profiles of tripartite BM tumor cells, CTCs and MRD from 10 MM patients confirmed a patient-specific mutational landscape with genomic heterogeneity in MRD. For instance, one patient exhibited BRAF & DIS3 mutations and del(13q) detected at baseline in BM tumor cells and CTCs, which persisted in MRD. After induction, de novo clonal SNVs, including a NEK11 mutation, emerged, followed by the appearance of 3 subclonal CNA after transplant, possibly contributing to resistance. CONCLUSIONS This is the largest dataset analyzing the kinetics and molecular features of CTCs and MRD in SMM and active MM. We showed superior prognostic value of CTC and MRD kinetics over single assessments. Tumor dissemination seems to be driven by transcriptional priming rather than acquired secondary genetic events, while on-treatment resistance is linked to genomic and transcriptional evolution, without recurrent genetic alterations and with common molecular signatures of resistance.

Circulating tumor cells as potential prognostic biomarkers for early-stage pancreatic cancer: A systematic review and meta-analysis.

Pancreatic cancer is difficult to be diagnosed early clinically, while often leads to poor prognosis. If optimal personalized treatment plan can be provided to pancreatic cancer patient at an earlier stage, this can greatly improve overall survival (OS). Circulating tumor cells (CTCs) are a collective term for various types of tumor cells present in the peripheral blood (PB), which are formed by detachment during the development of solid tumor lesions. Most CTCs undergo apoptosis or are phagocytosed after entering the PB, whereas a few can escape and anchor at distal sites to develop metastasis, increasing the risk of death for patients with malignant tumors. To investigate the significance of CTCs in predicting the prognosis of early pancreatic cancer patients. The PubMed, EMBASE, Web of Science, Cochrane Library, China National Knowledge Infrastructure, China Biology Medicine, and ChinaInfo databases were searched for articles published through December 2022. Studies were considered qualified if they included patients with early pancreatic cancer, analyzed the prognostic value of CTCs, and were full papers reported in English or Chinese. Researches were selected and assessed using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses protocol and the Newcastle-Ottawa Scale criteria. We used a funnel plot to assess publication bias. From 1595 publications, we identified eight eligible studies that collectively enrolled 355 patients with pancreatic cancer. Among these original studies, two were carried out in China; three in the United States; and one each in Italy, Spain, and Norway. All eight studies analyzed the relevance between CTCs and the prognosis of patients with early-stage pancreatic cancer after surgery. A meta-analysis showed that the patients that were positive pre-treatment or post-treatment for CTCs were associated with decreased OS [hazard ratio (HR) = 1.93, 95% confidence interval (CI): 1.197-3.126, P = 0.007] and decreased relapse-free/disease-free/progression-free survival (HR = 1.27, 95%CI: 1.137-1.419, P < 0.001) in early-stage pancreatic cancer. Additionally, the results suggest no statistically noticeable publication bias for overall, disease-free, progression-free, and recurrence-free survival. This pooled meta-analysis shows that CTCs, as biomarkers, can afford reliable prognostic information for patients with early-stage pancreatic cancer and help develop individualized treatment plans.

Advances in the role of circulating tumor cell heterogeneity in metastatic small cell lung cancer.

Small cell lung cancer (SCLC), a highly aggressive malignancy, is rapidly at an extensive stage once diagnosed and is one of the leading causes of death from malignancy. In the past decade, the treatment of SCLC has largely remained unchanged, and chemotherapy remains the cornerstone of SCLC treatment. The therapeutic value of adding immune checkpoint inhibitors to chemotherapy for SCLC is low, and only a few SCLC patients have shown a response to immune checkpoint inhibitors. Circulating tumor cells (CTCs) are tumor cells shed from solid tumor masses into the peripheral circulation and are key to tumor metastasis. Single-cell sequencing has revealed that the genetic profiles of individual CTCs are highly heterogeneous and contribute to the poor outcome and prognosis of SCLC patients. Theoretically, phenotypic analysis of CTCs may be able to predict the diagnostic significance of new potential targets for metastatic tumors. In this paper, we will discuss in depth the heterogeneity of CTCs in SCLC and the value of CTCs for the diagnosis and prognosis of SCLC and as relevant tumor markers in metastatic SCLC.

Analysis of Diagnostic Value of CTC and CTDNA in Early Lung Cancer.

This paper is the exploration and analysis of the diagnostic value of circulating tumor cells (CTC) and circulating tumor deoxyribonucleic acid (ctDNA) in early non-small cell lung cancer (NSCLC).A total of 125 NSCLC patients that hospitalized from March 2019 to December 2020 were selected, of which 64 subjects with stage I-II were listed as the early-stage group, and 61 patients with stage III-IV were classified as the intermediate-advanced group; In addition, 47 patients with benign pulmonary nodules hospitalized within same period were selected as a group of benign pulmonary nodules, and 50 healthy subjects were enrolled as a control group. The levels of CTC and ctDNA, serum tumor markers carcinoembryonic antigen (CEA), neuron-specific enolase (NSE), cytokeratin 21-1 (CYFRA21-1) and gastrin-releasing peptide precursor (pro-GRP) were detected. The diagnostic value of CTC and ctDNA in early NSCLC patients was analyzed by ROC curve. The area under the curve of CTC count for early-stage lung cancer was 0. 949 (P=0. 000, 95%CI: 0. 918-0. 980), and ctDNA was 0. 914 (P=0. 000, 95%CI: 0. 866~0. 963). There were significant differences in CTC count and ctDNA among subjects in each group (P<0. 05). CTC count and ctDNA of the early and intermediate-advanced group were remarkably higher than those of the control group and group of benign pulmonary nodules (P<0. 05), and CTC count and ctDNA of patients in the middle-late group were critically higher than those in the early-stage group (P<0. 05). There were significant differences in serum CEA, NSE, CYFRA21-1 and pro-GRP levels among all groups (P<0. 05). The results showed that CTC and ctDNA in NSCLC patients were significantly positively correlated with CEA, NSE, CYFRA21-1 and pro-GRP (P<0. 05). Peripheral blood CTC and ctDNA levels are significantly increased in NSCLC patients. They are positively correlated with serum tumor markers CEA, NSE, CYFRA21-1 and pro-GRP levels, and are related to tumor progression. CTC and ctDNA have high value in the clinical diagnosis of early NSCLC.

Prognostic value of tumor-derived extracellular vesicles and circulating tumor cells in metastatic breast cancer (BC): A focus on inflammatory BC.

1098 Background: Circulating tumor cells (CTCs) are an independent prognostic factor in metastatic breast cancer (MBC). The prognostic value of CTCs and the optimal cutoff for patients (pts) with inflammatory breast cancer (IBC), one of the most aggressive types of BC, has not been fully established. Recent evidence showed the complementary prognostic value of tumor-derived extracellular vesicles (tdEVs) to CTCs in MBC. The significance of tdEVs in IBC is unexplored. This study aimed to assess the prognostic value of CTCs and tdEVs in metastatic IBC. Methods: This study retrospectively analyzed 308 pts with MBC enrolled at Northwestern University (Chicago, IL) before starting a new line of therapy between 2016 and 2021 (NU16B06 trial). Blood samples were processed for CTCs using the CellSearch system. We applied the open source ACCEPT software to archived CellSearch images to enumerate CTCs and tdEVs. TdEVs cutoff levels were &lt;20 (low), 20-79 (intermediate), and ≥80 (high), as previously reported. The association of CTCs and tdEVs with overall survival (OS) was tested in the overall population (OvP) and IBC pts. Results: Of the 308 pts, 69 were diagnosed with IBC. 51% of pts received first-line therapy. CTCs enumerated by ACCEPT were strongly correlated with manual count (r=0.86) and hence used for this analysis. Median CTC count was 1 [interquartile range (IQR) 0-7] in IBC pts and 3 (IQR 0-14) in non-IBC (p=0.11). A significantly lower median tdEVs count was observed in IBC (7; IQR 3-45) than in non-IBC (22; IQR 4-137) pts (p=0.03). In IBC, higher CTC and tdEV counts were seen in the triple negative subtype (p=0.03) and non-visceral involvement (p=0.02), respectively. In the OvP median OS (mOS) was worse among pts with ≥5 CTCs (HR 2.6; p=0.001) and with elevated tdEVs (HR 3.3; p&lt;0.001). Only tdEVs were independently associated with poorer OS in multivariable analysis. In pts with &lt;5 CTCs there was a stepwise decrement in OS with increased tdEVs count (p=0.3); in pts with ≥5 CTCs, elevated tdEVs levels were associated with significantly worse OS (p=0.03). IBC pts with ≥5 CTCs had shorter mOS (8 vs 47 months; HR 3.6; p&lt;0.001); this association was weaker using ≥1 as CTC cutoff (HR 2.4; p=0.01). OS was adversely associated with increasing tdEVs levels (33 vs 23 vs 7 months for low, intermediate, and high subgroups, respectively; HR 3.7; p=0.001). Furthermore, elevated tdEVs were associated with shorter mOS in pts with ≥1 (p=0.055) and ≥5 CTCs (p=0.6). Among pts with &lt;5 CTCs no significant difference emerged among tdEVs subgroups (p=0.6). Conclusions: This study confirmed the strong prognostic significance of CTCs and tdEVs in MBC, including IBC. Pts with IBC had fewer CTCs and tdEVs compared to non-IBC, probably due to the predominant lymphatic spread of IBC. tdEVs offer the possibility to better define prognosis of IBC pts. Further studies are needed to evaluate their complementarity to CTCs.

Abstract P1-05-28: Decreases in Circulating Tumor Associated Cells Predicts PFS and OS In A Pooled Analysis Of Phase I Clinical Trials Using SV-BR-1-GM Therapy With Or Without Immune Check-Point Inhibitors In Metastatic Breast Cancer Patients

Abstract Background: In metastatic Breast Cancer (mBC), Circulating Tumor Cells (CTCs) are clinical indicators of worse prognosis and indicate patients (pts) not responding to current therapy. However CTCs are rare, found in &amp;lt; 20% of mBC pts, and many pts without CTCs may also progress. Recently an inflammatory pro-tumorigenic PD-L1 expressing macrophage (Cancer associated macrophage-like cell [CAML])was identified in the blood, which was found in &amp;gt;90% of mBC pts and may indicate tumor response to new therapies (JCO 40[16_Suppl] 2022). SV-BR-1-GM is a mBC cell line derived with antigen presenting characteristics was developed for treatment of mBC as a monotherapy (monoTx), or in combination with checkpoint inhibitors (comboTx). We report preliminary post-hoc results of a pooled analysis of n=18 monoTx mBCs pts and interim results of n=15 comboTx to analyze the predictive value of CTCs &amp; CAMLs, as well as CAML PD-L1 expression, isolated from pt peripheral blood pre &amp; post treatment to predict drug response, with end point outcomes of Progression Free Survival (PFS) and Overall Survival (OS) at 24 months. Methods: The SV-BR-1-GM regimen includes low pre-dose cyclophosphamide, intradermal inoculation of ~20 million irradiated SV-BR-1-GM cells and post-dose local interferon-α with cycles every 2 weeks x 3, then monthly. ComboTx adds an anti-PD-1 antibody with cycles every 3 weeks. Blinded blood samples were taken at baseline (BL), prior to starting SV-BR-1-GM therapy (n=33), and a 2nd sample (T1) taken after therapy initiation (~52 days) obtained as part of the exploratory portion of 2 prospective phase I clinical drug studies, NCT03066947 &amp; NCT03328026, to evaluate the predictive value of CTCs/CAMLs and CAML PD-L1 measured by LifeTracDx liquid biopsy. The quantities of CTCs &amp; CAMLs were analyzed based on PFS using RECIST v1.1 and OS hazard ratios (HRs) by censored univariate analysis at 24 months. Results: A total of 33 mBC pts were pooled from monoTx (n=18), or comboTx (n =15), all with available blood samples at BL. CTCs were found in 30% (n=10/33) of pts at BL, and CAMLs were found in 94% (n=31/33) at BL. Presence of CTCs at BL did not correlate with worse PFS (HR=1.0, p=0.8550), but did trend for pts with worse survival (HR=5.1, p=0.0641). T1 samples were available from 61% (n=20/33) pts. A drop in CAMLs or CTCs after treatment at T1 was observed in 50% of pts, which correlated with a significantly improved PFS HR=11.8, p=0.0019 and OS HR=226.3, p=0.0397. Overall, pts with a decrease in CTCs/CAMLs after induction of SV-BR-1-GM therapy had a ~350% improvement in median PFS (1.9 mo. vs 6.6 mo.) and a ~200% improvement in median OS (6.3 mo. vs 12.4 mo). When stratified between monoTx and comboTx, pts with a decrease in CTCs/CAMLs had an improved PFS (HR 11.9, p=0.0136) in the monoTx group and an improved PFS (HR 17.5, p=0.0017) in the comboTx group. Further, while expression of CAML PDL1 at BL was not correlated with improved PFS (HR=1.0, p=0.9078), CAML PD-L1 expression at BL was correlated with significantly better OS HR=9.5, p=0.0116, consistent with long term benefit of SV-BR-1-GM therapy in this group of pts. Conclusions: We observed that treatment with the SV-BR-1-GM regimen was associated with decreases in the presence of CTCs and CAMLs in 50% of patients, which also significantly correlated with ~350% better median PFS and ~200% better median OS within 2 years. SV-BR-1-GM therapy alone, or as a combination treatment with anti-PD-1, appears to have improved long term clinical outcomes in a large portion of heavily pre-treated mBC patients compared to other typical standard of care published results. Citation Format: Daniel Adams, Mingjin Chang, Miguel Lopez-Lago, Cha-Mei Tang, William Williams, Giuseppe Del Priore. Decreases in Circulating Tumor Associated Cells Predicts PFS and OS In A Pooled Analysis Of Phase I Clinical Trials Using SV-BR-1-GM Therapy With Or Without Immune Check-Point Inhibitors In Metastatic Breast Cancer Patients [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P1-05-28.

Circulating Tumor Cell Detection by Liquid Biopsy during Early-Stage Endometrial Cancer Surgery: A Pilot Study

The recurrence of non-metastatic endometrial carcinoma (EC) (6 to 21%) might be due to disseminated tumor cells. This feasibility study investigated whether circulating tumor cells (CTCs) were detectable in blood samples from the peripheral and ovarian veins of 10 patients undergoing laparoscopic resection of stage I-II EC between July 2019 and September 2021. CTCs were detected using the CellSearch® system (i) preoperatively (T0) in peripheral blood, (ii) after ovary suspensory ligament pediculation in ovarian vein blood (T1), and (iii) before colpotomy in peripheral blood (T2). CTCs were detected only in ovarian vein samples in 8/10 patients. The CTC median number did not differ with patient age (37 (min-max: 0-91) in <70-year-old vs. 11 (0-65) in ≥70 year-old women, p = 0.59), tumor grade (15 (0-72) for grade 1 vs. 15 (0-91) for grade 2, p = 0.97), FIGO stage (72 (27-91) vs. 2 (0-65) vs. 3 (0-6]) for stage IA, B, and II, respectively; p = 0.08), and tumor size (40 (2-72) for size < 30 mm vs. 4 (0-91) for size ≥ 30 mm, p = 0.39). Estrogen receptor-positive CTCs and CTC clusters were identified. The prognostic and therapeutic values of CTCs released during EC surgery need to be determined.

Prognostic value of stem-like circulating tumor cells in patients with cancer: a systematic review and meta-analysis.

Despite increasing interest in the study of circulating tumor cells (CTC) subsets, especially epithelial-mesenchymal transition(EMT) and stem cells subsets of CTC that play a key role in tumor recurrence and metastasis, there is no evidence from meta-analyses that shows the correlation between stem-like CTCs and prognosis in cancer patients. Thus, we performed a meta-analysis to assess its prognostic value. Sixteen articles were screened by searching the PubMed, Embase, Cochrane, China National Knowledge Internet (CNKI) and Wanfang databases. The hazard ratio (HR) and 95% confidence interval (95% CI) extracted from each article were summarized. Patients with positive stem-like CTCs in peripheral blood had significantly shorter overall survival (OS, HR: 2.58, 95% CI 1.76-3.79, P < 0.00001), progression-free survival (PFS, HR: 2.21, 95% CI 1.26-3.89, P = 0.006) and disease-free survival (DFS, HR: 2.53, 95% CI: 1.12-5.70, P = 0.03). This study provides the first meta-analysis evidence for the prognostic value of stem-like CTCs, demonstrating that these cells are associated with poor prognosis in cancer patients.Systematic review registrationCRD42022322062.

Current Status of Circulating Tumor Cells in Head and Neck Squamous Cell Carcinoma: A Review.

Circulating tumor cells (CTCs) are found in the blood of patients with cancer, including head and neck squamous cell carcinomas (HNSCCs). The aim is to review the most up-to-date status of CTCs for applications in patients with HNSCC. English articles in PubMed. All the studies on CTCs in HNSCCs in the literature were reviewed. There is emerging information on the diagnostic and prognostic value of CTCs in HNSCCs. Evidence also highlights the advantages of various downstream analysis approaches over circulating tumor DNA (ctDNA), such as single-CTC analysis, ex vivo, and in vivo expansion of CTCs. Multiple phenotypic surface markers (cytokeratins, EpCAM, vimentin, etc.), used for CTCs characterization using different immunoassays, could predict disease progression as well as patients' response to treatment efficacy. Immune checkpoint inhibitors' status in CTCs could also provide better insight into treatment. Clonal expansion of CTCs and single-cell analysis of CTCs are the most emerging fields nowadays which may offer an understanding of the mechanism of tumor evolution as well as therapeutic efficacy. Although several clinical trials are ongoing, limitations still exist in the detection and characterization of CTCs. Due to the lack of a gold standard protocol, the sensitivity and specificity of CTC enumeration methods vary. Prospective clinical trials are still needed before CTCs can be employed as diagnostic and prognostic markers in the clinical management of patients with HNSCC.

Liquid Biopsy Detecting Circulating Tumor Cells in Patients with Non-Small Cell Lung Cancer: Preliminary Results of a Pilot Study.

Lung cancer is still the leading cause of cancer-related death worldwide. Interest is growing towards early detection and advances in liquid biopsy to isolate circulating tumor cells (CTCs). This pilot study aimed to detect epithelial CTCs in the peripheral blood of early-stage non-small cell lung cancer (NSCLC) patients. We used Smart BioSurface® (SBS) slide, a nanoparticle-coated slide able to immobilize viable nucleated cellular fraction without pre-selection and preserve cell integrity. Forty patients undergoing lung resection for NSCLC were included; they were divided into two groups according to CTC value, with a cut-off of three CTCs/mL. All patients were positive for CTCs. The mean CTC value was 4.7(± 5.8 S.D.) per ml/blood. In one patient, next generation sequencing (NGS) analysis of CTCs revealed v-raf murine sarcoma viral oncogene homolog B(BRAF) V600E mutation, which has also been identified in tissue biopsy. CTCs count affected neither overall survival (OS, p = 0.74) nor progression-free survival (p = 0.829). Multivariable analysis confirmed age (p = 0.020) and pNodal-stage (p = 0.028) as negative predictors of OS. Preliminary results of this pilot study suggest the capability of this method in detecting CTCs in all early-stage NSCLC patients. NGS on single cell, identified as CTC by immunofluorescence staining, is a powerful tool for investigating the molecular landscape of cancer, with the aim of personalized therapies.

Association between circulating tumor cells in the peripheral blood and the prognosis of gastric cancer patients: a meta-analysis.

Research on the correlation between circulating tumor cells (CTCs) and gastric cancer (GC) has increased rapidly in recent years. However, whether CTCs are associated with GC patient prognosis is highly controversial. This study aims to evaluate the value of CTCs to predict the prognosis of GC patients. A meta-analysis. We searched the PubMed, Embase, and Cochrane Library databases for studies that reported the prognostic value of CTCs in GC patients before October 2022. The association between CTCs and overall survival (OS) and disease-free survival (DFS)/recurrence-free survival (RFS) and progression-free survival (PFS) of GC patients was assessed. Subgroup analyses were stratified by sampling times (pre-treatment and post-treatment), detection targets, detection method, treatment method, tumor stage, region, and HR (Hazard Ratio) extraction methods. Sensitivity analysis was performed by removing individual studies to assess the stability of the results. Publication bias was evaluated using funnel plots, Egger's test, and Begg's test. We initially screened 2000 studies, of which 28 were available for further analysis, involving 2383 GC patients. The pooled analysis concluded that the detection of CTCs was associated with poor OS (HR = 1.933, 95% CI 1.657-2.256, p < 0.001), DFS/RFS (HR = 3.228, 95% CI 2.475-4.211, p < 0.001), and PFS (HR = 3.272, 95% CI 1.970-5.435, p < 0.001). Furthermore, the subgroup analysis stratified by tumor stage (p < 0.01), HR extraction methods (p < 0.001), detection targets (p < 0.001), detection method (p < 0.001), sampling times (p < 0.001), and treatment method (p < 0.001) all showed that CTC detection was associated with poor OS and DFS/RFS for GC patients. Furthermore, the study showed that CTCs were associated with the poor DFS/RFS of GC when CTCs were detected for patients from Asian or No-Asian regions (p < 0.05). In addition, higher CTCs predicted poorer OS for GC patients who are from Asian regions (p < 0.001), but without statistical difference for GC patients from No-Asian regions (p = 0.490). CTC detection in peripheral blood was associated with poor OS, DFS/RFS, and PFS in patients with GC.

Liposome-tethered supported lipid bilayer platform for capture and release of heterogeneous populations of circulating tumor cells.

Because of scarcity, vulnerability, and heterogeneity in the population of circulating tumor cells (CTCs), the CTC isolation system relying on immunoaffinity interaction exhibits inconsistent efficiencies for all types of cancers and even CTCs with different phenotypes in individuals. Moreover, releasing viable CTCs from an isolation system is of importance for molecular analysis and drug screening in precision medicine, which remains a challenge for current systems. In this work, a new CTC isolation microfluidic platform was developed and contains a coating of the antibody-conjugated liposome-tethered-supported lipid bilayer in a developed chaotic-mixing microfluidic system, referred to as the "LIPO-SLB" platform. The biocompatible, soft, laterally fluidic, and antifouling properties of the LIPO-SLB platform offer high CTC capture efficiency, viability, and selectivity. We successfully demonstrated the capability of the LIPO-SLB platform to recapitulate different cancer cell lines with different antigen expression levels. In addition, the captured CTCs in the LIPO-SLB platform can be detached by air foam to destabilize the physically assembled bilayer structures due to a large water/air interfacial area and strong surface tension. More importantly, the LIPO-SLB platform was constructed and used for the verification of clinical samples from 161 patients with different primary cancer types. The mean values of both single CTCs and CTC clusters correlated well with the cancer stages. Moreover, a considerable number of CTCs were isolated from patients' blood samples in the early/localized stages. The clinical validation demonstrated the enormous potential of the universal LIPO-SLB platform as a tool for prognostic and predictive purposes in precision medicine.

Novel size-based design of spiral microfluidic devices with elliptic configurations and trapezoidal cross-section for ultra-fast isolation of circulating tumor cells

Investigation and analysis of circulating tumor cells (CTCs) have been valuable resources for detecting and diagnosing cancer in its early stages. Recently, enumeration and separation of CTCs via microfluidic devices have attracted significant attention due to their low cost and easy setup. In this study, novel microfluidic devices based on size-dependent cell-sorting with a trapezoidal cross-section and elliptic spiral configurations were proposed to reach label-free, ultra-fast CTCs enrichment. Firstly, the possibility and quality of separation in the devices were evaluated via a numerical simulation. Subsequently, these devices were fabricated to investigate the effects of the altering curvature and the trapezoidal cross-section on the isolation of CTCs from the peripheral blood sample at varying flow rates ranging from 0.5 mL/min to 3.5 mL/min. The experimental results indicated that the flow rate of 2.5 mL/min provided the optimal separation efficiency in the proposed devices, which was in fine agreement with the numerical analysis results. In this experiment, the purity values of CTCs between 88% and 90% were achieved, which is an indicator of the high capability of the proposed devices for the isolation and enrichment of CTCs. This strategy is hoped to overcome the limitations of classical affinity-based CTC separation approaches in the future.

The Number of Intraoperative Intestinal Venous Circulating Tumor Cells Is a Prognostic Factor for Colorectal Cancer Patients.

Purpose To assess the association between intestinal venous blood (IVB) circulating tumor cells (CTCs) and clinicopathological parameters in stage I-III colorectal cancer (CRC) patients. Methods Participants were retrospectively retrieved, who were admitted to our hospital or took annual physical exams between December 1, 2015 and December 31, 2018. A negative enrichment-immunofluorescence in situ hybridization (NE-imFISH) technique was used to isolate and identify CTCs. Receiver operating characteristic (ROC) curves and Youden index values were used to determine the critical CTC cutoff value for the diagnosis of CRC. Kaplan-Meier and log-rank methods were used to conduct survival analyses, and multivariate Cox regression analyses were employed for multivariate corrections to comprehensively evaluate the value of CTCs in the diagnosis of CRC. Relationships between IVB CTCs, clinicopathological parameters, and prognosis were then analyzed based upon patient postoperative follow-up data. Results In total, we retrieved 282 patients including 48 healthy controls, 72 patients with benign colorectal tumors, and 162 CRC patients. CRC patients exhibited significantly higher numbers of CTCs relative to control patients or those with benign disease. CTC numbers in CRC patient peripheral blood (PB) and IVB were closely associated with tumor node metastasis (TNM) staging (P < 0.01), carbohydrate antigen-125 (CA-125) levels (P < 0.001), and KRAS (Kirsten rat sarcoma virus oncogene) mutation status (P < 0.001). The disease-free survival (DFS) of patients in the CTC-negative group was significantly longer than that of patients in the CTC-positive group (24.60 ± 13.31 months vs. 18.70 ± 10.19 months, P < 0.05), with the same being true with respect to their overall survival (OS) (30.60 ± 12.44 months vs. 35.25 ± 11.57 months, P < 0.05). A multivariate analysis revealed that the detection ≥2 CTCs/3.2 ml was independently associated with poorer DFS and OS. CTC counts were independently predictive of CRC patients TNM staging, CA-125, and KRAS mutation status in both univariate and multivariate Cox proportional hazards regression analyses. Conclusion CTCs are valuable biomarkers that can be monitored to predict CRC patient disease progression.