Circulating Tumor Cells In Peripheral Blood Research Articles

A novel electrochemiluminescent cytosensor using dual-target magnetic probe recognition and nanozymes-catalyzed cascade signal amplification for precise phenotypic enumeration of CTCs.

The inability of surgical biopsy to monitor the dynamic evolution of cancer cells hampers its capacity to reflect real-time tumor heterogeneity. Circulating tumor cells (CTCs), as a crucial target in liquid biopsy, offer a novel approach for accurate monitoring of tumors. However, the rarity and complex phenotype resulting from epithelial-mesenchymal transition pose challenges for conventional methods such as CellSearch and immunohistochemistry, which have insufficient ability for simultaneous phenotyping and enumeration of CTCs. The enumeration of a single phenotype CTCs is insufficient for accurately assessing disease progression. Herein, we propose a strategy to address this issue by fabricating an electrochemiluminescence cytosensor via the integration of dual-target enrichment and nanozymes-catalyzed cascade signal amplification. The graphene oxide@hollow mesoporous Prussian blue/Pt (GO@HMPB/Pt) complex, possessing a large specific surface area and exceptional catalytic activity, is employed for loading a substantial amount of luminol as the signal probe. Dual-target magnetic PPy@Fe3O4/Au-antibody/aptamer is utilized for the magnetic capture of both epithelial and interstitial CTCs. Glutathione (GSH) can disrupt theAu-S bond on aptamer by a thiol exchange reaction and selectively releases a specific subset of phenotypic CTCs, thereby facilitating the efficient capture, accurate classification, and ultrasensitive detection of CTCs in peripheral blood. Using the epithelial MCF-7 and mesenchymal Hela cells as models, the ECL cytosensor demonstrates excellent performance in identifying cells spiked into whole blood. This study presents a novel approach for early detection of metastasis, tracking tumor recurrence, and monitoring therapeutic efficacy.

Circulating Tumor Cell Count and Overall Survival in Patients With Metastatic Hormone-Sensitive Prostate Cancer

In metastatic hormone-sensitive prostate cancer (mHSPC), new first-line combination therapies have enhanced overall survival (OS), but clinical outcomes for individual patients vary greatly and are difficult to predict. Peripheral blood circulating tumor cell (CTC) count is the most extensively validated prognostic liquid biomarker in metastatic castration-resistant prostate cancer (mCRPC), and recent studies have suggested that it may also be informative in mHSPC. To examine the prognostic value of CTC count in men with mHSPC. In this prognostic study, peripheral blood was drawn at registration (baseline) and at progression to mCRPC in the S1216 study (March 1, 2013, to July 15, 2017), a phase 3, prospective, randomized clinical trial in men with mHSPC. The CTCs were enumerated using a US Food and Drug Administration-cleared isolation platform. Counts were categorized as 0, 1 to 4, or 5 or more CTCs per 7.5 mL based on the prognostic value of these cut points in prior studies. The data analysis was performed between October 28, 2022, and June 15, 2023. Metastatic hormone-sensitive prostate cancer. Circulating tumor cell count was evaluated for an association with 3 prespecified trial end points: OS, progression-free survival, and 7-month prostate-specific antigen, after adjusting for other baseline covariates using proportional hazards and logistic regression models. Of 1313 S1216 participants (median [IQR] age, 68 [44-92] years), evaluable samples from 503 (median [IQR] age, 69 [46-90] years) with newly diagnosed mHSPC were collected at baseline, and 93 samples were collected at progression. Baseline counts were 5 or more CTCs per 7.5 mL in 60 samples (11.9%), 1 to 4 CTCs per 7.5 mL in 107 samples (21.3%), and 0 CTCs per 7.5 mL in 336 samples (66.8%). Median OS for men with 5 or more CTCs per 7.5 mL was 27.9 months (95% CI, 24.1-31.2 months) compared with 56.2 months (95% CI, 45.7-69.8 months) for men with 1 to 4 CTCs per 7.5 mL and not reached at 78.0 months follow-up for men with 0 CTCs per 7.5 mL. After adjusting for baseline clinical covariates, men with 5 or more CTCs per 7.5 mL at baseline had a significantly higher hazard of death (hazard ratio, 3.22; 95% CI, 2.22-4.68) and disease progression (hazard ratio, 2.46; 95% CI, 1.76-3.43) and a lower likelihood of prostate-specific antigen complete response (odds ratio, 0.26; 95% CI, 0.12-0.54) compared with men with 0 CTCs per 7.5 mL at baseline. Adding baseline CTC count to other known prognostic factors (covariates only: area under the curve, 0.73; 95% CI, 0.67-0.79) resulted in an increased prognostic value for 3-year survival (area under the curve, 0.79; 95% CI, 0.73-0.84). In this prognostic study, the findings validate CTC count as a prognostic biomarker that improved upon existing prognostic factors and estimated vastly divergent survival outcomes regardless of subsequent lines of therapy. As such, baseline CTC count in mHSPC may serve as a valuable noninvasive biomarker to identify men likely to have poor survival who may benefit from clinical trials of intensified or novel regimens.

Single-Cell Analysis of Bone-Marrow-Disseminated Tumour Cells

Metastasis frequently targets bones, where cancer cells from the primary tumour migrate to the bone marrow, initiating new tumour growth. Not only is bone the most common site for metastasis, but it also often marks the first site of metastatic recurrence. Despite causing over 90% of cancer-related deaths, effective treatments for bone metastasis are lacking, with current approaches mainly focusing on palliative care. Circulating tumour cells (CTCs) are pivotal in metastasis, originating from primary tumours and circulating in the bloodstream. They facilitate metastasis through molecular interactions with the bone marrow environment, involving direct cell-to-cell contacts and signalling molecules. CTCs infiltrate the bone marrow, transforming into disseminated tumour cells (DTCs). While some DTCs remain dormant, others become activated, leading to metastatic growth. The presence of DTCs in the bone marrow strongly correlates with future bone and visceral metastases. Research on CTCs in peripheral blood has shed light on their release mechanisms, yet investigations into bone marrow DTCs have been limited. Challenges include the invasiveness of bone marrow aspiration and the rarity of DTCs, complicating their isolation. However, advancements in single-cell analysis have facilitated insights into these elusive cells. This review will summarize recent advancements in understanding bone marrow DTCs using single-cell analysis techniques.

The clinical application of circulating tumor cells (CTCs) for immunotherapy in advanced pancreatic cancer.

e14541 Background: Circulating tumor cell (CTCs) has the potential utility for screening, prognostic and predictive assessment, and offers the potential for personalized management. This trial aimed to evaluate the prognostic and predictive impact of programmed cell death ligand 1 (PD-L1) positive CTCs in patients with pancreatic ductal adenocarcinoma (PDAC). Methods: Twenty-three heavily treated advanced PDAC patients were randomized to receive camrelizumab combined with apatinib or capecitabine, or placebo plus capecitabine until disease progression or unacceptable toxicity, and peripheral blood CTCs samples were collected at baseline and after treatment. Forty-nine CTCs samples from 23 recruited advanced PDAC patients were detected by subtraction enrichment and immunostaining-FISH (SE-iFISH). PD-L1 expression level also was evaluated in tumor tissue samples by Immunohistochemistry (IHC). Investigator-assessed 6-month overall survival (OS) rate and 3-month progression-free survival (PFS) rate was evaluated. Results: The positive detection rate of CTCs in 23 PDAC patients was 95.7%. The cell sub-types of sizes of CTC analysis showed that 58.6% CTCs was the small CTCs (Diameter < 5μm). CTCs karyotypes analysis exhibited that triploid small CTCs were the main karyotype (41.2%) in small CTC, and pentaploid large CTCs was the major karyotype (74.7%) in large CTCs. PD-L1+ CTCs were detected in 43.5% of PDAC patients. Tissue PD-L1 positive detection rate was 50% (CPS > 1%) in available tissue samples(n=10). The presence of PD-L1+ CTCs cohort exhibited a significant increase in 6-month OS rate (80% versus 38.5%, P < 0.05) and 3-month PFS rate (70% versus 23.1%, P < 0.05) compared to PD-L1- CTCs cohort. Compared with the PD-L1- CTCs patients, the combination therapy camrelizumab markedly improved the 6-month OS rates (75% versus 50%) and 3-month PFS rate (62.5% versus 25%) in PD-L1+ CTC patients. Conclusions: PD-L1+ CTCs could be considered as a favorable prognostic marker in advanced PDAC. These results suggest that PD-L1+ CTCs might be used as a predictor of the efficacy of cancer immunotherapy in advanced PDAC, and provide a new monitoring method for the precise treatment of advanced PDAC. The expanding sample size test is now ongoing. Clinical trial information: ChiCTR1900024095 .

Comprehensive characterization of circulating tumor cells and cell-free DNA in patients with metastatic melanoma.

Advances in therapeutic approaches for melanoma urge the need for biomarkers that can identify patients at risk for recurrence and to guide treatment. The potential use of liquid biopsies in identifying biomarkers is increasingly being recognized. Here, we present a head-to-head comparison of several techniques to analyze circulating tumor cells (CTCs) and cell-free DNA (cfDNA) in 20 patients with metastatic melanoma. In this study, we investigated whether diagnostic leukapheresis (DLA) combined with multimarker flow cytometry (FCM) increased the detection of CTCs in blood compared to the CellSearch platform. Additionally, we characterized cfDNA at the level of somatic mutations, extent of aneuploidy and genome-wide DNA methylation. Both CTCs and cfDNA measures were compared to tumor markers and extracranial tumor burden on radiological imaging. Compared to the CellSearch method applied on peripheral blood, DLA combined with FCM increased the proportion of patients with detectable CTCs from 35% to 70% (P = 0.06). However, the median percentage of cells that could be recovered by the DLA procedure was 29%. Alternatively, cfDNA mutation and methylation analysis detected tumor load in the majority of patients (90% and 93% of samples successfully analyzed, respectively). The aneuploidy score was positive in 35% of all patients. From all tumor measurements in blood, lactate dehydrogenase (LDH) levels were significantly correlated to variant allele frequency (P = 0.004). Furthermore, the presence of CTCs in DLA was associated with tumor burden (P < 0.001), whereas the presence of CTCs in peripheral blood was associated with number of lesions on radiological imaging (P < 0.001). In conclusion, DLA tended to increase the proportion of patients with detectable CTCs but was also associated with low recovery. Both cfDNA and CTCs were correlated with clinical parameters such as LDH levels and extracranial tumor burden.

Detection of PD‑L1 expression and epithelial‑mesenchymal transition of circulating tumor cells in non‑small cell lung cancer.

The present study aimed to assess the roles of peripheral circulating tumor cell (CTC) count, CTC subtypes and programmed death ligand 1 (PD-L1) expression in the clinical staging and prognosis of patients with non-small cell lung cancer (NSCLC). A total of 100 patients with NSCLC with available tumor tissues were enrolled in the present study, and 7.5 ml peripheral blood was collected. Patients were divided into PD-L1-positive and PD-L1-negative groups according to PD-L1 immunohistochemical staining. Peripheral blood samples from both groups were analyzed to determine the CTC count, epithelial-type CTCs (E-CTCs), mesenchymal-type CTCs (M-CTCs) and PD-L1 expression. Clinical data were collected, and patients were followed up for a maximum of 36 months, with patient death as the endpoint event. Patients with PD-L1-positive tumors had a worse prognosis compared with those with PD-L1-negative tumors (P=0.045). The PD-L1-positive group exhibited significantly higher numbers of CTCs and M-CTCs compared with the PD-L1-negative group (P≤0.05). However, the number of E-CTCs did not differ significantly between the two groups (P>0.05). PD-L1-positive patients with higher CTC and M-CTC counts had relatively poorer prognoses (P≤0.05), while the number of E-CTCs had no significant effect on prognosis (P>0.05). Compared with the early-stage NSCLC group, the late-stage NSCLC group exhibited a significant increase in the CTC count (P≤0.05), while E-CTC and M-CTC counts did not significantly differ between the two groups (P>0.05). The PD-L1-positive group exhibited a significant increase in the number of PD-L1+ CTCs and PD-L1+ M-CTCs compared with the PD-L1-negative group (P≤0.05), while PD-L1+ E-CTC counts did not differ significantly between the two groups (P>0.05). The PD-L1-positive patients with a higher number of PD-L1+ CTCs and PD-L1+ M-CTCs had relatively poorer prognoses (P≤0.05), while the PD-L1+ E-CTC count had no significant effect on prognosis (P>0.05). Compared with the early-stage NSCLC group, the late-stage NSCLC group exhibited a significant increase in the number of PD-L1+ CTCs and PD-L1+ M-CTCs (P≤0.05), while PD-L1+ E-CTC counts did not significantly differ between the two groups (P>0.05). Based on univariate and multivariate analyses, the number of PD-L1+ M-CTCs was identified as an independent prognostic factor for NSCLC. In conclusion, the presence of CTCs in peripheral blood, particularly PD-L1+ M-CTC subtype, indicated poorer clinical staging and prognosis in patients with NSCLC. These findings suggested that CTCs, specifically the PD-L1+ M-CTC subtype, could serve as a monitoring indicator for the clinical staging and prognosis of patients with NSCLC.

Abstract PO2-24-02: Molecular characterization of primary tumors associated with various CTCs’ subpopulation

Abstract Background: Circulating tumor cells (CTCs) are prognostic in primary breast cancer (BC) patients and represent a heterogeneous population of cells with different phenotypes and biological value. However, molecular characterization of primary tumors associated with various CTCs’ subpopulation is lacking. This study aimed to identify expression of genes associated with the presence of different CTCs subpopulations in primary BC patients using a comprehensive genomics approach. Methods: This prospective ongoing study included 312 patients (pts) with non-metastatic BC enrolled from March 2012 to February 2015. CTCs were detected on day -1 to 0 before surgery. Isolated peripheral blood mononuclear cells (PBMC) were depleted of hematopoietic cells using RossetteSepTM kit negative selection with anti-CD45 antibody. RNA extracted from CD45-depleted (CD45-) PBMC was interrogated for expression of EMT-inducing transcription factors (TWIST1, SNAIL1, SLUG, ZEB1) and epithelial (CK19) gene transcripts by quantitative reverse transcription-PCR. Expressions of gene transcripts in CD45- PBMC of patients were compared to those of CD45- PBMC of 60 healthy donors. Formalin-fixed paraffin-embedded tumor tissue was subject for gene expression analysis performed by HTG EdgeSeq Oncology Biomarker panel (HTG Molecular Diagnostics, Inc., Tucson, USA). Results: CTCs with epithelial phenotype (CTC_EP) were detected in 30 (9.6%) pts, CTC with EMT (epithelial-to-mesenchymal transition) were detected in 54 (17.3%), while blood of 79 (25.3%) pts exhibit at least one type of CTC. Patients with detectable CTC in peripheral blood had inferior disease-free survival compared to patients without detectable CTC (HR = 0.62, 95% CI 0.37-1.06, P=0.048). There was no association between epithelial CTCs and analysed patients/tumour variables. Genes expression analysis identified 7 differentially expressed genes in patients with the CTC_EP phenotype (CHGA, MESP1, COL2A1, MT2A, COMP, MMP7 and ORM2), 200 genes associated with CTC_EMT phenotype (top 10 genes included CHGA, CECR6, SMAD2, HUS1, FGF10, HNF1A, SEC61G, CDK4, CD44 and F3). Analysis identified that low expression of CHGA and MMP7 was consistently associated with any CTC subpopulations. Conclusions: In this study for the first-time revealed gene expressions associated with various CTC subpopulations. We suppose that these genes could represent potential therapeutic target aimed to inhibit metastatic cascade in breast cancer. Citation Format: Michal Mego, Gabriel Minarik, Milan Hucko, Katarina Kalavska, Marian Karaba, Juraj Benca, Tatiana Sedlackova, Lucia Kucerova, Jozef Mardiak, Lubos Klucar, Zuzana Cierna. Molecular characterization of primary tumors associated with various CTCs’ subpopulation [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-24-02.

Precise Capture and Dynamic Release of Circulating Liver Cancer Cells with Dual-Histidine-Based Cell Imprinted Hydrogels.

Circulating tumor cells (CTCs) detection presents significant advantages in diagnosing liver cancer due to its noninvasiveness, real-time monitoring, and dynamic tracking. However, the clinical application of CTCs-based diagnosis is largely limited by the challenges of capturing low-abundance CTCs within a complex blood environment while ensuring them alive. Here, an ultrastrong ligand, l-histidine-l-histidine (HH), specifically targeting sialylated glycans on the surface of CTCs, is designed. Furthermore, HH is integrated into a cell-imprinted polymer, constructing a hydrogel with precise CTCs imprinting, high elasticity, satisfactory blood compatibility, and robust anti-interference capacities. These features endow the hydrogel with excellent capture efficiency (>95%) for CTCs in peripheral blood, as well as the ability to release CTCs controllably and alive. Clinical tests substantiate the accurate differentiation between liver cancer, cirrhosis, and healthy groups using this method. The remarkable diagnostic accuracy (94%), lossless release of CTCs, material reversibility, and cost-effectiveness ($6.68 per sample) make the HH-based hydrogel a potentially revolutionary technology for liver cancer diagnosis and single-cell analysis.

Digital Circulating Tumor Cells Quantification.

Circulating tumor cells (CTCs) are an emerging but vital biomarker for cancer management. An efficient methodology for accurately quantifying CTCs remains challenging due to their rareness. Here, we develop a digital CTC detection strategy using partitioning instead of enrichment to quantify CTCs. By utilizing the characteristics of droplet microfluidics that can rapidly generate a large number of parallel independent reactors, combined with Poisson distribution, we realize the quantification of CTCs in the blood directly. The limit of detection of our digital CTCs quantification assay is five cells per 5 mL of whole blood. By simultaneously detecting multiple genetic mutations, our approach achieves highly sensitive and specific detection of CTCs in peripheral blood from NSCLC patients (AUC = 1). Our digital platform offers a potential approach and strategy for the quantification of CTCs, which could contribute to the advancement of cancer medical management.

The detection of circulating tumor cells indicates poor therapeutic efficacy and prognosis in patients with nonsmall cell lung cancer: A systematic review and meta-analysis.

The efficacy and prognostic value of circulating tumor cells (CTCs) in nonsmall cell lung cancer (NSCLC) are controversial based on the existing research. This systematic review and meta-analysis evaluated the significance of CTCs in NSCLC therapy monitoring and prognosis prediction, supporting their potential as clinical biomarkers. We conducted a comprehensive search of PubMed, Embase, Web of Science, The Cochrane Library, WanFang Data, CNKI, and VIP through September 20, 2023. Inclusion criteria were cohort studies involving NSCLC patients, focusing on peripheral blood CTCs, and assessing outcomes such as pre- and posttreatment CTC rates or levels, progression-free survival (PFS), and overall survival (OS). Two reviewers independently extracted the data and assessed risk of bias using the Newcastle-Ottawa Scale. We utilized Review Manager 5.4.1 for meta-analysis, calculating pooled odds ratios (ORs) for dichotomous outcomes, mean differences for continuous variables and hazard ratios (HRs) for survival data, applying fixed- or random-effects models based on heterogeneity assessed by the I2 statistic. This study was registered in PROSPERO (No. CRD42023450035). Twenty-two eligible studies with a total of 1674 NSCLC patients were included. Meta-analysis results showed that the CTCs-positive rate (OR=0.59, 95% CI 0.45 to 0.77, p=0.0001) and CTCs count (mean difference=-3.10, 95% CI -5.52 to -0.69, p=0.01) were significantly decreased after antitumor treatment. Compared with the CTCs nonreduced group, the CTC-reduced group showed better PFS (HR=1.71, 95% CI 1.35 to 2.17, p<0.00001) and OS (HR=1.50, 95% CI 1.21 to 1.86, p=0.0003) after treatment. PFS and OS in CTC-positive groups were lower than those in the CTCs-negative group pretreatment (HR=2.49, 95% CI 1.78 to 3.47, p<0.00001; HR=1.80, 95% CI 1.29 to 2.52, p=0.0006) and posttreatment (HR=3.36, 95% CI 2.12 to 5.33, p<0.00001; HR=3.31, 95% CI 1.75 to 6.27, p=0.0002). CTCs can be used as a biomarker to monitor NSCLC efficacy, predict prognosis and guide follow-up treatment.

Multi-DNA-Modified Double-Network Hydrogel with Customized Microstructure: A Novel System for Living Circulating Tumor Cells Capture and Real-Time Detection.

The precise and effective isolation of living circulating tumor cells (CTCs) from peripheral blood, followed by their real-time monitoring, is crucial for diagnosing cancer patients. In this study, a cell-imprinted double-network (DN) hydrogel modified with circular multi-DNA (CMD), coined the CMD-imprinted hydrogel with fixed cells as templates (CMD-CIDH), was developed. The hydrogel featured a customized surface for proficient capture of viable CTCs and in situ real-time fluorescent detection without subsequent release. The customized surface, constructed using polyacrylamide/chitosan DN hydrogel as the matrix on the cell template, had a dense network structure, thereby ensuring excellent stability and a low degradation rate. Optimal capture efficiencies, recorded at 93 ± 3% for MCF-7 cells and 90 ± 2% for Hela cells, were achieved by grafting the CMD and adjusting the nodule size on the customized surface. The capture efficiency remained significantly high at 67 ± 11% in simulated breast cancer patient experiments even at a minimal concentration of 5 cells mL-1. Furthermore, CMD grafted onto the surface produced a potent fluorescence signature, enabling in situ real-time fluorescent detection of the target cell's growth state even in complex environments. The customized surface is highly efficient for screening CTCs in peripheral blood and has promising potential for setting up the CTCs culture.

A novel method for detecting circulating tumor cells immunity based on micro-nano technique

This study was to develop a new method for detecting circulating tumor cells (CTCs) with high sensitivity and specificity, therefore to detect the colorectal cancer as early as possible for improving the detection rate of the disease. To this end, we prepared some micro-column structure microchips modified with graphite oxide-streptavidin (GO-SA) on the surface of microchips, further coupled with a broad-spectrum primary antibody (antibody1, Ab1), anti-epithelial cell adhesion molecule (anti-EpCAM) monoclonal antibody to capture CTCs. Besides, carboxylated multi-walled carbon nanotubes (MWCNTs-COOH) were coupled with colorectal cancer related antibody as specific antibody 2 (Ab2) to prepare complex. The sandwich structure consisting of Ab1-CTCs-Ab2 was constructed by the microchip for capturing CTCs. And the electrochemical workstation was used to detect and verify its high sensitivity and specificity. Results showed that the combination of immunosensor and micro-nano technology has greatly improved the detection sensitivity and specificity of the immunosensor. And we also verified the feasibility of the immunosensor for clinical blood sample detection, and successfully recognitized detection and quantization of CTCs in peripheral blood of colorectal cancer patients by this immunosensor. In conclusion, the super sandwich immunosensor based on micro-nano technology provides a new way for the detection of CTCs, which has potential application value in clinical diagnosis and real-time monitoring of disease.

Correlation analysis of circulating tumor cells and Claudin-4 in breast cancer.

Objective: We aimed to explore the relationship between peripheral blood circulating tumor cells (CTCs) and the expression of Claudin-4 in patients with breast cancer, and further explore the potential impact on clinical prognosis and risk assessment. Methods: We classified and enumerated circulating tumor cells in the blood of breast cancer patients by CTC-enriched in situ hybridization and the detection of Claudin-4 expression by immunohistochemistry. We carried out an analysis of the correlation between the two and the comparison of their impact on clinical parameters and prognosis. Results: There were 38 patients with a low expression of Claudin-4 and 27 patients with a high expression of Claudin-4. Compared with Claudin-4 low-expression patients, the number of CTCs was higher in patients with high Claudin-4 expression (11.7 vs. 7.4, p < 0.001). High Claudin-4 expression was associated with a lower count of epithelial CTCs (E-CTCs) (3.4 vs. 5.0, p = 0.033), higher counts of mesenchymal CTCs (M-CTC) (4.4 vs. 1.1, p < 0.001), and epithelial/mesenchymal CTCs (E/M-CTCs) (4.0 vs. 3.5, p = 0.021). The intensity of Claudin-4 was positively correlated with CTC (rs = 0.43, p = 0.001). Multivariate COX regression analysis showed that CTC counts (HR = 1.3, p < 0.001), Claudin-4 (HR = 4.6, p = 0.008), and Lymphatic metastasis (HR = 12.9, p = 0.001) were independent factors for poor prognosis. COX regression of CTC classification showed that epithelial/mesenchymal CTCs (E/M-CTC) (HR = 1.9, p = 0.001) and mesenchymal CTCs (M-CTC) (HR = 1.5, p = 0.001) were independent influencing factors of adverse reactions in breast cancer patients. Conclusion: The number of CTC in breast cancer is positively correlated with the expression of Claudin-4. High CTC counts and a high proportion of M-CTCs correlated with Claudin-4 expression. CTC counts and Claudin-4 expression were independent predictors of poor prognosis in breast cancer patients.

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.

Liquid Biopsy in Hepatocellular Carcinoma: The Significance of Circulating Tumor Cells in Diagnosis, Prognosis, and Treatment Monitoring.

Hepatocellular carcinoma (HCC) is an aggressive malignancy with poor outcomes when diagnosed at an advanced stage. Current curative treatments are most effective in early-stage HCC, highlighting the importance of early diagnosis and intervention. However, existing diagnostic methods, such as radiological imaging, alpha-fetoprotein (AFP) testing, and biopsy, have limitations that hinder early diagnosis. AFP elevation is absent in a significant portion of tumors, and imaging may have low sensitivity for smaller tumors or in the presence of cirrhosis. Additionally, as our understanding of the molecular pathogenesis of HCC grows, there is an increasing need for molecular information about the tumors. Biopsy, although informative, is invasive and may not always be feasible depending on tumor location. In this context, liquid biopsy technology has emerged as a promising approach for early diagnosis, enabling molecular characterization and genetic profiling of tumors. This technique involves analyzing circulating tumor cells (CTCs), circulating tumor DNA (ctDNA), or tumor-derived exosomes. CTCs are cancer cells shed from the primary tumor or metastatic sites and circulate in the bloodstream. Their presence not only allows for early detection but also provides insights into tumor metastasis and recurrence. By detecting CTCs in peripheral blood, real-time tumor-related information at the DNA, RNA, and protein levels can be obtained. This article provides an overview of CTCs and explores their clinical significance for early detection, prognosis, treatment selection, and monitoring treatment response in HCC, citing relevant literature.

Exploring the Potential Driver Gene Mutations That Promote Renal Cancer Cell Metastasis and Implantation Based on Circulating Tumor Cells Culture.

Studies have shown that the circulating tumor cell (CTC) is a necessary condition for the invasion and distant metastasis of renal cell carcimona (RCC). However, few CTCs-related gene mutations have been developed which could promote the metastasis and implantation of RCC. The objective of this study is to explore the potential driver gene mutations that promote RCC metastasis and implantation based on CTCs culture. Fifteen patients with primary mRCC and three healthy subjects were included, and peripheral blood was obtained. After the preparation of synthetic biological scaffolds, peripheral blood CTCs were cultured. Successful cultured CTCs were applied to construct CTCs-derived xenograft (CDX) models, followed by DNA extraction, whole exome sequencing (WES) and bioinformatics analysis. Synthetic biological scaffolds were constructed based on previously applied techniques, and peripheral blood CTCs culture was successfully performed. We then constructed CDX models and performed WES, and explored the potential driver gene mutations that may promote RCC metastasis and implantation. Bioinformatics analysis showed that KAZN and POU6F2 may be closely related to the prognosis of RCC. We successfully performed the culture of peripheral blood CTCs and, on this basis we initially explored the potential driver mutations for the metastasis and implantation of RCC.

Analysis of different protein expression levels in peripheral blood circulating tumor cells from patients with diffuse large B-cell lymphoma and their predictive efficiency for recurrence

Objective: To analyze the expression levels of differentiation cluster 47 (CD47), signal regulatory protein α (SIRP-α), proto-oncogene (MYC) and proliferating cell associated antigen (Ki67) proteins in peripheral blood circulating tumor cells (CTC) from patients with diffuse large B-cell lymphoma and their predictive efficiency for tumor recurrence. Methods: The data of 82 patients with diffuse large B-cell lymphoma who were confirmed by histopathology and were in remission after chemotherapy in the Hematology Department of Linyi People's Hospital from January 2018 to January 2021 were retrospectively analyzed. There were 44 males and 38 females, and aged from 50 to 75 (63.8±4.6) years. The patients were divided into recurrent group (n=36) and non-recurrent group (n=46) according to their recurrence within 1 year after remission. The fasting peripheral venous blood samples (4 ml) from patients in the morning were collected, and the CTC were isolated. The expression levels of CD47, SIRP-α, MYC and Ki67 proteins in CTC were detected by Western blotting. The correlations between CD47 expression level and SIRP-α, MYC and Ki67 expression levels were analyzed by Pearson correlation analysis. The predictive efficiency of CD47, SIRP-α, MYC and Ki67 expression levels on tumor recurrence was evaluated by receiver operating characteristic (ROC) curves, and the areas under the curve (AUC) were calculated. Results: The expression levels of CD47, SIRP-α, MYC and Ki67 in recurrent group were 2.24±0.23, 1.17±0.12, 1.98±0.20 and 2.63±0.27, while those in non-recurrent group were 2.04±0.21, 1.31±0.13, 1.53±0.16 and 2.24±0.25. The expression levels of CD47, MYC and Ki67 in the recurrent group were higher than those in the non-recurrent group, while the expression levels of SIRP-α were lower than those in the non-recurrent group (all P<0.001). In 82 patients, the expression levels of CD47, SIRP-α, MYC and Ki67 were 2.13±0.22, 1.25±0.13, 1.73±0.18 and 2.41±0.26, respectively. The expression level of CD47 was negatively correlated with the expression level of SIRP-α (r=-0.308, P=0.005), but positively correlated with the expression level of MYC and Ki67 (r=0.484 and 0.332, P=0.012 and 0.003). The sensitivity of CD47, SIRP-α, MYC and Ki67 expression levels in predicting recurrence of diffuse large B-cell lymphoma was 66.7%, 72.2%, 72.2% and 66.7%, with the specificity of 67.4%, 71.7%, 67.4% and 71.7%, and AUC (95%CI) of 0.694 (0.582-0.791), 0.693 (0582-0.790), 0.714 (0.603-0.808) and 0.709 (0.598-0.804), respectively. The sensitivity of the combined detection of the above four indicators was 83.3%, with the specificity of 78.3% and the AUC (95%CI) of 0.864 (0.771-0.930), which was higher than those of the individual detection of each indicator (all P<0.05). Conclusions: The expression level of CD47 was negatively correlated with the expression level of SIRP-α, but positively correlated with the expression level of MYC and Ki67. The expression levels of CD47, SIRP-α, MYC and Ki67 have certain predictive value for tumor recurrence in patients with diffuse large B-cell lymphoma, and the predictive efficiency of combined detection is higher than single indicator detection.

Value of peripheral blood circulating tumor cell detection in the diagnosis of thoracic diseases and the prediction of severity.

Circulating tumor cell (CTC) detection, as a noninvasive liquid biopsy method, has been used in the diagnosis, prognostic indication, and monitoring of a variety of cancers. In this study, we aimed to investigate whether CTC detection could be used in the early diagnosis and prediction of severity of thoracic diseases. We enrolled 168 thoracic disease patients, all of whom underwent pathological biopsy. Carcinoembryonic antigen (CEA) and neuron-specific enolase (NSE) measurement was also performed in 146 patients. There were 131 cases of malignant thoracic diseases and 37 cases of benign lesions. We detected CTCs in a 5ml peripheral blood sample with the CTCBiopsy® system and analyzed the value of CTC count for predicting disease severity. Of 131 patients with a diagnosis of thoracic malignancy, CTCs were found in blood samples from 122 patients. However, only 2 out of 37 patients with benign thoracic disease had no detectable CTCs. There was no significant correlation between CTC count and benign and malignant lesions (P = 0.986). However, among 131 patients who had been diagnosed with malignant lesions, 33 had lymph node metastasis or distant metastasis. The presence of CTCs was significantly correlated with metastasis (P = 0.016 OR = 1.14). The area under the receiver operating characteristic (ROC) curve was 0.625 (95% confidence interval (CI), 0.519 to 0.730 P = 0.032). In addition, with stage IA1 as the cutoff, all patients were further divided into an early-stage group and a late-stage group. CTC count was significantly correlated with disease progression (P = 0.031 OR = 1.11), with an area under the curve (AUC) of 0.599 (95% CI, 0.506-0.692 P = 0.47). The sensitivity and specificity of CTC detection for the diagnosis of disease stage were 72.3% and 45.5%, respectively. In addition, the cutoff of 2.5 CTCs was the same when predicting disease metastasis and staging. Furthermore, the combination of CTC count, demographic characteristics and tumor markers had better predictive significance for disease staging. CTC count can effectively indicate the stages and metastasis of thoracic diseases, but it cannot differentiate benign and malignant diseases.

Gelatin‐Coated ZnO Array Nanoplatform for Capture, Drug Resistance Identification, and Recovery of Circulating Tumor Cells

AbstractDrug resistance causes failure of cancer treatment. Identifying drug sensitivity of tumors can guide therapeutic administration for cancer patients, however this approach lacks a facile, fast, and noninvasive evaluation platform. Herein, a gelatin‐coated ZnO array nanoplatform to isolate circulating tumor cells (CTCs) is developed, and their drug resistance for inspecting tumor sensitivity is assessed. The nanoplatform is comprised of a nanosubstrate of ZnO submicropillars (ZnO SMPs) for efficiently isolating cancer cells, and Au nanoparticles (AuNPs) based electrochemical probes for evaluating drug resistance of the captured cells. Gelatin, an extracellular matrix‐like and enzyme‐degradable material, is used to coat ZnO SMPs and AuNPs, endowing the nanoplatform with the ability to maintain cancer cell viability and realize enzyme‐responsive cell recovery for downstream bioanalysis. This nanoplatform is expected to noninvasively evaluate drug sensitivity of cancer patients through simple liquid biopsy of CTCs in peripheral blood and benefit the treatment.

Abstract P3-07-12: Allogeneic, Antigen-Presenting, GM-CSF-secreting, SV-BR-1-GM Whole Cell Therapeutic Vaccine in Advanced Metastatic Breast Cancer

Abstract Background: SV-BR-1-GM is an irradiated allogeneic cell line derived from a hormone receptor-negative (HR-) HER2-positive (HER2+) breast cancer (BC), now engineered to express HLA class I and II antigens, secrete GM-CSF and function as an antigen-presenting cell. Here we report post-hoc exploratory data for metastatic BC (MBC) patients treated with the SV-BR-1-GM regimen (SV) alone (NCT03066947) and in combination (CO) with immune checkpoint inhibitors (ICIs) (NCT03328026). Methods: SV includes cyclophosphamide 300 mg/m2 i.v. 48-72 hours prior to SV-BR-1-GM (20 × 106 cells) intradermally followed by interferon-alpha-2b at the SV-BR-1-GM inoculation sites on days 2 and 4. SV was given every 2 weeks for 3 cycles and then monthly in NCT03066947, and every 3 weeks in combination with PD-1 inhibitors in NCT03328026. Treatment was continued until disease progression or unacceptable toxicities. Results: A total of 34 refractory MBC patients (pts) were treated, including 24 with SV, and 14 with CO, including 4 who crossed over from SV to CO plus one who restarted the SV after a protocol specified administrative interruption in treatment. Median prior regimens were 5; 55% of pts had hormone receptor-positive (HR+) BC, 42% had HER2+ BC, and 16% were triple-negative BC (TNBC). Objective response rates (ORR) were 4% for monotherapy and 7% for combination. Clinical benefit rate (CBR) was 21% for monotherapy and 29% for combination. Responses and clinical benefits were seen across most BC subtypes (see table). Notably, CBR among 10 pts with HR+ BC (any HER2) who received the combination was 50% (5/10). The duration of response (DOR) among 4 patients treated with monotherapy was 49-223 days and DOR among 6 patients treated with combination was 72-292 days. Median PFS was 2.8 months for monotherapy and 4.2 months for combination. Median OS was 7.0 months for monotherapy, and 12.0 months for combination. Of 28 pts with available prior treatment data, 12 (43%) had PFS on study treatment exceeded PFS on prior treatment, of these 8/12 had &amp;gt;1 HLA match and 5/8 had ≥ 2 loci HLA-matched with SV-BR-1-GM. PFS ratio improvement was independent of the prior number of lines of therapy or BC subtypes. The SV-BR-1GM regimen was well tolerated with the most common treatment-related adverse events (AEs) being injection site reaction in 67%. There was no dose-limiting toxicity, special interest AEs such as cytokine storm, and no grade 5 observed. There were 2 non-treatment-related grade 4 AEs: worsening pleural effusion and altered mental status. Significant improvement in PFS was observed in patients with matched HLA and in association with the delayed-type hypersensitivity skin test, peripheral blood circulating tumor cells, and cancer-associated macrophages. Conclusions: SV-BR-1-GM demonstrated promising activity in patients with MBC. Treatment was well tolerated with no concerning AEs. The PFS ratio compares favorably with prior penultimate standards of care, more notably in patients with matched HLA. Clinical benefits were observed across multiple subtypes of BC, particularly in patients with HR+ disease receiving combination therapy. Phase II clinical trial is currently ongoing to evaluate the efficacy of SV-BR-1-GM in combination with ICIs. Clinical benefit and PFS on SV-BR-1-GM SV-BR-1-GM Whole Cell Therapeutic Vaccine in Heavily Pre-Treated Metastatic Breast Cancer Results Stratified by HER2 and HLA Matching Citation Format: Saranya Chumsri, William Williams, Mingjin Chang, Miguel Lopez-Lago, Charles Wiseman, Jarrod Holmes, Chaitali Nangia, Karim Mohammed, Minal Barve, Shaker Dakhil, Bonnie Guerin, Giuseppe Del Priore, Carmen Calfa. Allogeneic, Antigen-Presenting, GM-CSF-secreting, SV-BR-1-GM Whole Cell Therapeutic Vaccine in Advanced Metastatic Breast Cancer [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 P3-07-12.