In vivofluorescenceflow cytometry reveals that the nanoparticle tumor vaccine OVA@HA-PEI effectively clears circulating tumor cells

Tumor vaccine therapy offers significant advantages over conventional treatments,including reduced toxic side effects.However,it currently functions primarily as an adjuvant treatment modality in clinical oncology due to limitations in tumor antigen selection and delivery methods.Tumor vaccines often fail to elicit a su±ciently robust immune response against progressive tumors,thereby limiting their clinical e±cacy.In this study,we developed a nanoparticle-based tumor vaccine,OVA@HA-PEI,utilizing ovalbumin(OVA)as the presenting antigen and hyaluronic acid(HA)and polyethyleneimine(PEI)as adjuvants and carriers.This formulation significantly enhanced the proliferation of immune cells and cytokines,such as CD3,CD8,interferon-,and tumor necrosis factor-,in vivo,effectively activating an immune response against B16–F10 tumors.In vivofluorescenceflow cytometry(IVFC)has already become an effective method for monitoring circulating tumor cells(CTCs)due to its direct,noninvasive,and long-term detection capabilities.Our study utilized a laboratory-constructed IVFC system to monitor the immune processes induced by the OVA@HA-PEI tumor vaccine and an anti-programmed death-1(PD-1)antibody.The results demonstrated that the combined treatment of OVA@HA-PEI and anti-PD-1 antibody significantly improved the survival time of mice compared to anti-PD-1 antibody treatment alone.Additionally,this combination therapy substantially reduced the number of CTCs in vivo,increased the clearance rate of CTCs by the immune system,and slowed tumor progression.Thesefindings greatly enhance the clinical application prospects of IVFC and tumor vaccines.

Prognostic value of circulating tumor cells combined with neutrophil-lymphocyte ratio in patients with hepatocellular carcinoma

BACKGROUND Circulating tumor cell(CTC)count and neutrophil-to-lymphocyte ratio(NLR)are both closely associated with the prognosis of hepatocellular carcinoma(HCC).AIM To investigate the prognostic value of combining these two indicators in HCC.METHODS Clinical data were collected from patients with advanced HCC who received im-mune therapy combined with targeted therapy at the Department of Oncology,the Affiliated Hospital of Southwest Medical University,Sichuan,China,from 2021 to 2023.The optimal cutoff values for CTC programmed death-ligand 1(PD-L1)(+)>1 or CTC PD-L1(+)≤1 and NLR>3.89 or NLR≤3.89 were evaluated using X-Tile software.Patients were categorized into three groups based on CTC PD-L1(+)counts and NLR:CTC-NLR(0),CTC-NLR(1),and CTC-NLR(2).The relationship between CTC-NLR and clinical variables as well as survival rates was assessed.RESULTS Patients with high CTC PD-L1(+)expression or NLR at baseline had shorter median progression-free survival(m-PFS)and median overall survival(mOS)than those with low levels of CTC PD-L1(+)or NLR(P<0.001).Mean-while,patients in the CTC-NLR(2)group showed a significant decrease in mPFS and mOS.Cox regression analysis revealed that alpha-fetoprotein(AFP),CTC PD-L1(+),and CTC-NLR were independent predictors of OS.The time-dependent receiver operating characteristic curve showed that the area under the curve of CTC-NLR at 12 months(0.821)and 18 months(0.821)was superior to that of AFP and CTC PD-L1(+).CONCLUSION HCC patients with high CTC PD-L1(+)or NLR expression tend to exhibit poor prognosis,and a high baseline CTC-NLR score may indicate low survival.CTC-NLR may serve as an effective prognostic indicator for patients with advanced HCC receiving immunotherapy combined with targeted therapy.

Circulating tumor cells in pancreatic cancer:The prognostic impact in surgical patients

Pancreatic cancer is associated with a poor prognosis,even in the early stages,mainly due to metastatic progression.New diagnostic techniques that predict unfavorable outcomes are needed in order to improve treatment strategies.Circulating tumor cells(CTCs)are showing promising results as a predictive biomarker for various tumors.In this editorial we comment on the article by Zhang et al,who published the first systematic review and meta-analysis evaluating the prognostic value of CTCs as biomarkers in early-stage pancreatic cancer patients undergoing surgery.CTCs were detected in peripheral or central venous system blood,before or during surgery.Positive CTCs showed a correlation with decreased overall survival and decreased relapse-free,disease-free and progression-free survival in this meta-analysis.However,the heterogeneity was significant.The authors suggest that this result was related to the separation methods used between studies,but other differences such as the margin status or the neoadjuvant and adjuvant treatments used are also important to consider.CTCs may be a potential prognostic biomarker in pancreatic cancer patients,but it is necessary to compare and standardize the platforms used to isolate CTCs,to compare different biomarkers from liquid biopsy and to determine the impact on prognosis when therapeutic changes are made based on CTCs levels.

Circulating tumor cells as prognostic marker in pancreatic cancer

In this editorial we comment on the article by Zhang et al published in the recent issue of the World Journal of Clinical Oncology.Pancreatic cancer is the fourth most common cause of cancer-related mortality and has the lowest survival rate among all solid cancers.It causes 227000 deaths annually worldwide,and the 5-year survival rate is very low due to early metastasis,which is 4.6%.Cancer survival increases with better knowledge of risk factors and early and accurate diagnosis.Circulating tumor cells(CTCs)are tumor cells that intravasate from the primary tumor or metastasis foci into the peripheral blood circulation system spontan-eously or during surgical operations.Detection of CTC in blood is promising for early diagnosis.In addition,studies have associated high CTC levels with a more advanced stage,and more intensive treatments should be considered in cases with high CTC.In tumors that are considered radiologically resectable,it may be of critical importance in detecting occult metastases and preventing unnecessary surgeries.

Circulating tumor cells and circulating tumor DNA in breast cancer diagnosis and monitoring

Liquid biopsy,including both circulating tumor cells and circulating tumor DNA,is becoming more popular as a diagnostic tool in the clinical management of breast cancer.Elevated concentrations of these biomarkers during cancer treatment may be used as markers for cancer progression as well as to understand the mechanisms underlying metastasis and treatment resistance.Thus,these circulating markers serve as tools for cancer assessing and monitoring through a simple,non-invasive blood draw.However,despite several study results currently noting a potential clinical impact of ctDNA mutation tracking,the method is not used clinically in cancer diagnosis among patients and more studies are required to confirm it.This review focuses on understanding circulating tumor biomarkers,especially in breast cancer.

Recent progress in aptamer-based microfluidics for the detection of circulating tumor cells and extracellular vesicles

Liquid biopsy is a technology that exhibits potential to detect cancer early,monitor therapies,and predict cancer prognosis due to its unique characteristics,including noninvasive sampling and real-time analysis.Circulating tumor cells(CTCs)and extracellular vesicles(EVs)are two important components of circulating targets,carrying substantial disease-related molecular information and playing a key role in liquid biopsy.Aptamers are single-stranded oligonucleotides with superior affinity and specificity,and they can bind to targets by folding into unique tertiary structures.Aptamer-based microfluidic platforms offer new ways to enhance the purity and capture efficiency of CTCs and EVs by combining the advantages of microfluidic chips as isolation platforms and aptamers as recognition tools.In this review,we first briefly introduce some new strategies for aptamer discovery based on traditional and aptamer-based microfluidic approaches.Then,we subsequently summarize the progress of aptamer-based microfluidics for CTC and EV detection.Finally,we offer an outlook on the future directional challenges of aptamer-based microfluidics for circulating targets in clinical applications.

Circulating tumor cells: Biological features and survival mechanisms

Circulating tumor cells(CTCs)are neoplastic cells that are detached from primary tumors and enter circulation.Enumeration and characterization of CTCs are of significance in cancer diagnosis,prognosis,and treatment monitoring.CTC survival in the bloodstream is a limiting step for the development of metastases in distant organs.Recent technological advances,especially in single-cell molecular analyses have uncovered heterogeneous CTC survival mechanisms.Undergoing epithelial-to-mesenchymal transition(EMT),increasing stem cell-like properties,and forming cell clusters enable CTCs to adapt to the harsh microenvironment of the circulation.Expressing and releasing several immunosuppressive molecules help CTCs escape from anti-cancer immune mechanisms.This review article summarizes the biological characteristics of CTCs and focuses on the recent understanding of the mechanisms by which CTCs survive in circulation.Additionally,the clinical and therapeutic implications of CTCs are discussed.

Microfluidic platform for circulating tumor cells isolation and detection

Circulating tumor cells(CTCs)are essential biomarkers for liquid biopsies,which are important in the early screening,prognosis,and real-time monitoring of cancer.However,CTCs are less abundant in the peripheral blood of patients,therefore,their isolation is necessary.Recently,the use of microfluidics for CTC sorting has become a research hotspot owing to its low cost,ease of integration,low sample consumption,and unique advantages in the manipulation of micron-sized particles.Herein,we review the latest research on microfluidics-based CTC sorting.Specifically,we consider active sorting using external fields(electric,magnetic,acoustic,and optical tweezers)and passive sorting using the flow effects of cells in specific channel structures(microfiltration sorting,deterministic lateral displacement sorting,and inertial sorting).The advantages and limitations of each method and their recent applications are summarized here.To conclude,a forward-looking perspective is presented on future research on the microfluidic sorting of CTCs.

A pilot study of the relative number of circulating tumor cells and leukocytes containing actin-binding proteins in head and neck cancer patients

Circulating tumor cells(CTCs)play an important role in tumor metastases,which is positively correlated with an increased risk of death.Actin-binding proteins,including cofilin(CFL1),profilin 1(PFN1),and adenylate cyclase-associated protein 1(CAP1),are thought to be involved in tumor cell motility and metastasis,specifically in head and neck squamous cell carcinoma(HNSCC).However,currently,there are no published studies on CFL1,PFN1,and CAP1 in CTCs and leukocytes in HNSCC patients.We assessed serum levels of CFL1,PFN1,and CAP1 and the number of CTCs and leukocytes containing these proteins in blood from 31 HNSCC patients(T1-4N0-2M0).The analysis used flow cytometry and an enzyme-linked immunosorbent assay kit.We found that CAP1+CTCs and CAP1+leukocyte subpopulations were prevalent in these HNSCC patient samples,while the prevalence rates of CFL1+and PFN1+CTCs were relatively low.Patients with stage T2-4N1-2M0 had CFL1+and PFN1+CTCs with an elevated PFN1 serum level,compared with the T1-3N0M0 group.In summary,the PFN1 serum level and the relative number of PFN1+CD326+CTCs could be valuable prognostic markers for HNSCC metastases.The current study is the first to obtain data regarding the contents of actin-binding proteins(ABPs)in CTCs,and leukocytes in blood from HNSCC patients.This is also the first to assess the relationship between the number of CTCs subgroups and disease characteristics.

Clinical implications and perspectives of portal venous circulating tumor cells in pancreatic cancer

Despite recent improvements in the diagnosis and treatment of pancreatic cancer(PC),clinical outcomes remain dismal.Moreover,there are no effective prognostic or predictive biomarkers or options beyond carbohydrate antigen 19-9 for personalized and precise treatment.Circulating tumor cells(CTCs),as a member of the liquid biopsy family,could be a promising biomarker;however,the rarity of CTCs in peripheral venous blood limits their clinical use.Because the first venous drainage of PC is portal circulation,the portal vein can be a more suitable location for the detection of CTCs.Endoscopic ultrasound-guided portal venous sampling of CTCs is both feasible and safe.Several studies have suggested that the detection rate and number of CTCs may be higher in the portal blood than in the peripheral blood.CTC counts in the portal blood are highly associated with hepatic metastasis,recurrence after surgery,and survival.The phenotypic and genotypic properties measured in the captured portal CTCs can help us to understand tumor heterogeneity and predict the prognosis of PC.Small sample sizes and heterogeneous CTC detection methods limit the studies to date.Therefore,a large number of prospective studies are needed to corroborate portal CTCs as a valid biomarker in PC.

Circulating tumor cells(CTCs)in breast cancer:a diagnostic tool for prognosis and molecular analysis

Metastatic breast cancer （MBC） is characterized by a combination of tumor growth, proliferation and metastatic progression and is typically managed with palliative intent. The benefit of standard systemic therapies is relatively limited and the disease is considered incurable suggesting the need to investigate the biological drivers of the various phases of the metastatic process in order to improve the selection of molecularly driven therapies. The detection, enumeration and molecular analysis of circulating tumor cells （CTCs） provide an intriguing opportunity to advance this knowledge. CTCs enumerated by the Food and Drugs Administration-cleared CellSearchTM system are an independent prognostic factor of progression- free survival （PFS） and overall survival （OS） in MBC patients. Several published papers demonstrated the poor prognosis for MBC patients that presented basal CTC count _〉5 in 7.5 mL of blood. Therefore, the enumeration of CTCs during treatment for MBC provides a tool with the ability to predict progression of disease earlier than standard timing of anatomical assessment using conventional radiological tests. During the metastatic process cancer cells exhibit morphological and phenotypic plasticity undergoing epithelial- mesenchymal transition （EMT）. This important phenomenon is associated with down regulation of epithelial marker （e.g., EpCAM） with potential limitations in the applicability of current CTCs enrichment methods. Such observations translated in a number of investigations aimed at improving our capabilities to enumerate and perform molecular characterization of CTCs. Theoretically, the phenotypic analysis of CTCs can represent a ＂liquid＂ biopsy of breast tumor that is able to identify a new potential target against the metastatic disease and advance the development and monitoring of personalized therapies.

甲状腺癌患者131I治疗前后CTC水平变化及临床意义

目的:探究甲状腺癌患者放射性碘同位素(131 I)治疗前后外周血循环肿瘤细胞(CTC)水平变化,分析其临床意义。方法:研究对象选取自本院2022年1月至2022年12月收治的108例行131 I治疗的甲状腺癌患者,根据131 I治疗12个月后预后情况将患者分为转移组(19例)和未转移组(89例),与131 I治疗前后比较两组CTC水平,并对两组患者的一般资料进行比较,应用Logistic回归分析筛选影响患者131 I治疗预后的因素。结果:131 I治疗前,CTC水平为15.51±1.67(FR/mL)显著高于治疗后CTC水平8.50±2.30(FR/mL)(P<0.05);未转移组治疗前后CTC水平差值显著高于转移组(P<0.05);两组患者的年龄、性别、住院天数、肿瘤类型、肿瘤位置、包膜侵犯情况、肿瘤T分期、残余甲状腺质量、手术时间至131 I治疗时间、131 I治疗的次数、颈部侧区淋巴结阳性数、是否合并桥本比较差异无统计学意义(P>0.05),颈部中央区淋巴结阳性数比较差异具有统计学意义(P<0.05);Logistic回归分析显示颈部中央区淋巴结阳性数是甲状腺癌患者甲状腺切除术后131 I治疗后复发转移的独立影响因素。结论:甲状腺切除术后进行131 I治疗能有效改善患者CTC水平,出现复发转移患者的治疗前后CTC水平改善程度更低,CTC对预测甲状腺切除术联合131 I治疗后复发转移具有较好的价值,颈部中央区淋巴结阳性数也是甲状腺癌患者131 I治疗后复发转移的影响因素。

Evaluation of epithelial-mesenchymal transitioned circulating tumor cells in patients with resectable gastric cancer: Relevance to therapy response

AIM: To evaluate the epithelial-to-mesenchymal transition(EMT) of circulating tumor cells(CTCs) in gastric cancer patients.METHODS: We detected tumor cells for expression of four epithelial(E^+) transcripts(keratins 8, 18, and 19 and epithelial cell adhesion molecule) and two mesenchymal(M^+) transcripts(Vimentin and Twist) by a quantifiable, dual-colorimetric RNA-in situ hybridization assay. Between July 2014 and October 2014, 44 patients with gastric cancer were recruited for CTC evaluation. Blood samples were obtained from selected patients during the treatment course [before surgery, after surgery and at the 6^(th) cycle of XELOX based chemotherapy(about 6 mo postoperatively)].RESULTS: We found the EMT phenomenon in which there were a few biphenotypic E^+/M^+ cells in primary human gastric cancer specimens. Of the 44 patients, the presence of CTCs was reported in 35(79.5%) patients at baseline. Five types of cells including from exclusively E^+ CTCs to intermediate CTCs and exclusively M^+ CTCs were identified(4 patients with M^+ CTCs and 10 patients with M^+ or M^+ > E^+ CTCs). Further, a chemotherapy patient having progressive disease showed a proportional increase of mesenchymal CTCs in the post-treatment blood specimens. We used NCI-N87 cells to analyze the linearity and sensitivity of Can Patrol^(TM) system and the correlation coefficient(R^2) was 0.999.CONCLUSION: The findings suggest that the EMT phenomenon was both in a few cells of primary tumors and abundantly in CTCs from the blood of gastric cancer patients, which might be used to monitor therapy response.

Prognostic and predictive blood biomarkers in gastric cancer and the potential application of circulating tumor cells

Gastric cancer(GC), with its high incidence and mortality rates, is a highly fatal cancer that is common in East Asia particularly in China. Its recurrence and metastasis are the main causes of its poor prognosis. Circulating tumor cells(CTCs) or other blood biomarkers that are released into the circulating blood stream by tumors are thought to play a crucial role in the recurrence and metastasis of gastric cancer. Therefore, the detection of CTCs and other blood biomarkers has an important clinical significance; in fact, they can help predict the prognosis, assess the staging, monitor the therapeutic effects and determine the drug susceptibility. Recent research has identified many blood biomarkers in GC, such as various serum proteins, autoantibodies against tumor associated antigens, and cell-free DNAs. The analysis of CTCs and circulating cell-free tumor DNA(ctDNA) in the peripheral blood of patients with gastric cancer is called as liquid biopsy. These blood biomarkers provide the disease status for individuals and have clinical meaning. In this review, we focus on the recent scientific advances regarding CTCs and other blood biomarkers, and discuss their origins and clinical meaning.

Molecular detection of epithelial-mesenchymal transition markers in circulating tumor cells from pancreatic cancer patients:Potential role in clinical practice

AIM To evaluate the clinical properties of three subpopulations of circulating tumor cells(CTCs) undergoing epithelial-mesenchymal transition(EMT) in pancreatic ductal adenocarcinoma(PDAC) patients.METHODS We identified CTCs for expression of the epithelial cell marker cytokeratin or epithelial cell adhesion molecule(EpCAM)(E-CTC), the mesenchymal cell markers vimentin and twist(M-CTC), or both(E/M-CTC) using the CanPatrol system. Between July 2014 and July 2016, 107 patients with PDAC were enrolled for CTC evaluation. CTC enumeration and classification were correlated with patient clinicopathological features and outcomes.RESULTS CTCs were detected in 78.5% of PDAC patients. The number of total CTCs ranged from 0 to 26 across all 107 patients, with a median value of six. CTC status correlated with lymph node metastasis, TNM stage, distant metastasis, blood lymphocyte counts, and neutrophil-to-lymphocyte ratio(NLR). Kaplan-Meier survival analysis showed that patients with ≥ 6 total CTCs had significantly decreased overall survival and progression-free survival compared with patients with < 6 total CTCs. The presence of M-CTCs was positively correlated with TNM stage(P < 0.01) and distant metastasis(P < 0.01). Additionally, lymphocyte counts and NLR in patients without CTCs were significantly different from those in patients testing positive for each CTC subpopulation(P < 0.01).CONCLUSION Classifying CTCs by EMT markers helps to identify the more aggressive CTC subpopulations and provides useful evidence for determining a suitable clinical approach.

Circulating tumor cells in pancreatic cancer patients:Enrichment and cultivation

AIM: To investigate the feasibility of separation and cultivation of circulating tumor cells (CTCs) in pancreatic cancer (PaC) using a filtration device.

Liquid biopsy in patients with pancreatic cancer: Circulating tumor cells and cell-free nucleic acids

Despite recent advances in surgical techniques and perioperative management, the prognosis of pancreatic cancer(PCa) remains extremely poor. To provide optimal treatment for each patient with Pca, superior biomarkers are urgently needed in all phases of management from early detection to staging, treatment monitoring, and prognosis. In the blood of patients with cancer, circulating tumor cells(CTCs) and cell-free nucleic acids(cf NAs), such as DNA, m RNA, and noncoding RNA have been recognized. In the recent years, their presence in the blood has encouraged researchers to investigate their potential use as novel blood biomarkers, and numerous studies have demonstrated their potential clinical utility as a biomarker for certain types of cancer. This concept, called "liquid biopsy" has been focused on as a less invasive, alternative approach to cancer tissue biopsy for obtaining genetic and epigenetic aberrations that contribute to oncogenesis and cancer progression. In this article, we review the available literature on CTCs and cfN As in patients with cancer, particularly focusing on PCa, and discuss future perspectives in this field.

High circulating tumor cell concentrations in a specific subtype of gastric cancer with diffuse bone metastasis at diagnosis

AIM: To clarify the biological feature contributing to gastric cancer with diffuse bone metastases at diagnosis.METHODS: The participants visited the Department of Clinical Oncology, Akita University Hospital, from January 2014 to August 2015. The selection criterion for gastric cancer with diffuse bone metastases at diagnosis includes over 29 hot spots of bone scintigraphy. Circulating tumor cell were collected from 20 m L of peripheral venous blood drawn using a Cell Search kit and a Cell Tracks Auto Prep system by SRL, a clinical laboratory. The endpoints of this study were correlations between circulating tumor cells(CTC) count and therapeutic outcomes. RESULTS: Among 39 patients with gastric cancer, 5 patients met the criterion. The incidence of this subtype was 12.8%. CTC counts ranged from 235 to 6440 cells/7.5 m L of peripheral blood(median of 1724). These values were much higher than common gastric cancers(2 cells). In chemo-sensitive cases, CTC counts decreased within 14 d(median) from 275, 235 and 1724 to 2, 7 and 66, respectively. On the other hand, CTC counts increased after treatment failure or insensitive case from 2, 7 and 6440 to 787, 513 and 7885, respectively. The correlation between CTC count and survival time showed a trend, but did not reach significance(Y = 234.6- 0.03 X, P = 0.085).CONCLUSION: High CTC count is a biological hallmark of this subtype, and can be used as a direct and definitive indicator of therapeutic outcome.

Circulating tumor and cancer stem cells in hepatitis C virusassociated liver disease

AIM: To assess the role of circulating tumor cells (CTCs) and cancer stem cells (CSCs) in hepatitis C virus (HCV)-associated liver disease.

Liquid biopsy in patients with hepatocellular carcinoma: Circulating tumor cells and cell-free nucleic acids

Hepatocellular carcinoma(HCC), with its high incidence and mortality rate, is one of the most common malignant tumors. Despite recent development of a diagnostic and treatment method, the prognosis of HCC remains poor. Therefore, to provide optimal treatment for each patient with HCC, more precise and effective biomarkers are urgently needed which could facilitate a more detailed individualized decision-making during HCC treatment, including the following; risk assessment, early cancer detection, prediction of treatment or prognostic outcome. In the blood of cancer patients, accumulating evidence about circulating tumor cells and cell-free nucleic acids has suggested their potent clinical utilities as novel biomarker. This concept, so-called "liquid biopsy" is widely known as an alternative approach to cancer tissue biopsy. This method might facilitate a more sensitive diagnosis and better decision-making by obtaining genetic and epigenetic aberrations that are closely associated with cancer initiation and progression. In this article, we review recent developments based on the available literature on both circulating tumor cells and cell-free nucleic acids in cancer patients, especially focusing on Hepatocellular carcinoma.