Detection of circulating tumor cells in hepatocellular carcinoma: applications in diagnosis, prognosis prediction and personalized treatment

Abstract

Hepatocellular carcinoma (HCC) is one of the most common malignant tumors worldwide and is associated with poor clinical prognosis, which is mainly caused by tumor recurrence and metastasis. Circulating tumor cells (CTCs) are tumor cells shed into the bloodstream and regarded as the “seeds” of tumor recurrent or metastatic lesions. Over the past decade, the clinical value of CTC analysis has been extensively explored. CTC analysis is a representative form of liquid biopsy, offering a novel solution that can bypass the problems of invasive biopsy procedures, enabling comprehensive, non-invasive, and real-time disease monitoring. In HCC, CTC analysis has facilitated early detection and prognosis prediction, as well as treatment monitoring and therapeutic intervention guiding. In this review, we summarize available literature and provide an overview of CTC biology, detection technologies, and clinical applications in the diagnosis, prognosis prediction, and personalized treatment of HCC.

Keywords

Hepatocellular carcinoma / liquid biopsy / circulating tumor cells / biomarkers / personalized medicine

References

[1] Siegel RL,Jemal A.Cancer statistics, 2020..CA Cancer J Clin2020;70:7-30

[2] Klein CA.Parallel progression of primary tumours and metastases..Nat Rev Cancer2009;9:302-12

[3] European Association for the Study of the Liver. Electronic address eee, European Association for the Study of the LEASL Clinical Practice Guidelines: management of hepatocellular carcinoma..J Hepatol2018;69:182-236

[4] Villanueva A,Llovet JM.Medical therapies for hepatocellular carcinoma: a critical view of the evidence..Nat Rev Gastroenterol Hepatol2013;10:34-42

[5] Tabrizian P,Shrager B,Roayaie S.Recurrence of hepatocellular cancer after resection: patterns, treatments, and prognosis..Ann Surg2015;261:947-55

[6] Heimbach JK,Finn RS,Abecassis MM.AASLD guidelines for the treatment of hepatocellular carcinoma..Hepatology2018;67:358-80

[7] Agopian VG,Markovic D,Kaldas FM.Evaluation of Patients With Hepatocellular Carcinomas That Do Not Produce alpha-Fetoprotein..JAMA Surg2017;152:55-64

[8] Chen VL,Wicha MS,Parikh ND.Utility of liquid biopsy analysis in detection of hepatocellular carcinoma, determination of prognosis, and disease monitoring: a systematic review..Clin Gastroenterol Hepatol2020;doi: 10.1016/j.cgh.2020.04.019

[9] Mann J,Feldstein AE.Liquid biopsy for liver diseases..Gut2018;67:2204-12

[10] Diaz LA Jr.Liquid biopsies: genotyping circulating tumor DNA..J Clin Oncol2014;32:579-86 PMCID:PMC4820760

[11] Massague J.Metastatic colonization by circulating tumour cells..Nature2016;529:298-306 PMCID:PMC5029466

[12] Keller L.Unravelling tumour heterogeneity by single-cell profiling of circulating tumour cells..Nat Rev Cancer2019;19:553-67

[13] de Bono JS,Montgomery RB,Miller MC.Circulating tumor cells predict survival benefit from treatment in metastatic castration-resistant prostate cancer..Clin Cancer Res2008;14:6302-9

[14] Ahn JC,Chen PJ,Tseng HR.Detection of circulating tumor cells and their implications as a novel biomarker for diagnosis, prognostication, and therapeutic monitoring in hepatocellular carcinoma..Hepatology2020;doi: 10.1002/hep.31165

[15] Ye Q,Zheng S.Liquid biopsy in hepatocellular carcinoma: circulating tumor cells and circulating tumor DNA..Mol Cancer2019;18:114 PMCID:PMC6607541

[16] Allard WJ,Miller MC,Connelly MC.Tumor cells circulate in the peripheral blood of all major carcinomas but not in healthy subjects or patients with nonmalignant diseases..Clin Cancer Res2004;10:6897-904

[17] Sun YF,Yang XR,Zhang X.Circulating stem cell-like epithelial cell adhesion molecule-positive tumor cells indicate poor prognosis of hepatocellular carcinoma after curative resection..Hepatology2013;57:1458-68

[18] Chang YS,McDonald DM,Jain RK.Mosaic blood vessels in tumors: frequency of cancer cells in contact with flowing blood..Proc Natl Acad Sci U S A2000;97:14608-13 PMCID:PMC18966

[19] Mohme M,Pantel K.Circulating and disseminated tumour cells - mechanisms of immune surveillance and escape..Nat Rev Clin Oncol2017;14:155-67

[20] Liberko M,Bobek V.Essentials of circulating tumor cells for clinical research and practice..Crit Rev Oncol Hematol2013;88:338-56

[21] Kim MY,Acharyya S,Zhang XH.Tumor self-seeding by circulating cancer cells..Cell2009;139:1315-26 PMCID:PMC2810531

[22] Coumans FA,Terstappen LW.Detection of cancer before distant metastasis..BMC Cancer2013;13:1-12 PMCID:PMC3684526

[23] Ebright RY,Wittner BS,Nicholson BT.Deregulation of ribosomal protein expression and translation promotes breast cancer metastasis..Science2020;367:1468-73 PMCID:PMC7307008

[24] Mani SA,Liao MJ,Ayyanan A.The epithelial-mesenchymal transition generates cells with properties of stem cells..Cell2008;133:704-15 PMCID:PMC2728032

[25] Kalluri R.EMT: when epithelial cells decide to become mesenchymal-like cells..J Clin Invest2009;119:1417-9 PMCID:PMC2689122

[26] Yu M,Wittner BS,Smas ME.Circulating breast tumor cells exhibit dynamic changes in epithelial and mesenchymal composition..Science2013;339:580-4 PMCID:PMC3760262

[27] Sun YF,Xu Y,Gong ZJ.Circulating tumor cells from different vascular sites exhibit spatial heterogeneity in epithelial and mesenchymal composition and distinct clinical significance in hepatocellular carcinoma..Clin Cancer Res2018;24:547-59

[28] Padmanaban V,Suhail Y,Aceto N.E-cadherin is required for metastasis in multiple models of breast cancer..Nature2019;573:439-44 PMCID:PMC7365572

[29] Thompson EW.The social aspects of EMT-MET plasticity..Nat Med2011;17:1048-9

[30] Aceto N.Bring along your friends: homotypic and heterotypic circulating tumor cell clustering to accelerate metastasis..Biomed J2020;43:18-23 PMCID:PMC7090281

[31] Aceto N,Miyamoto DT,Wittner BS.Circulating tumor cell clusters are oligoclonal precursors of breast cancer metastasis..Cell2014;158:1110-22 PMCID:PMC4149753

[32] Manjunath Y,Suvilesh KN,Kimchi ET.Circulating tumor cell clusters are a potential biomarker for detection of non-small cell lung cancer..Lung Cancer2019;134:147-50

[33] Wang C,Chervoneva I,Ye Z.Longitudinally collected CTCs and CTC-clusters and clinical outcomes of metastatic breast cancer..Breast Cancer Res Treat2017;161:83-94

[34] Wang L,Xu J,Wang X.Quantified postsurgical small cell size CTCs and EpCAM(+) circulating tumor stem cells with cytogenetic abnormalities in hepatocellular carcinoma patients determine cancer relapse..Cancer Lett2018;412:99-107

[35] Gkountela S,Szczerba BM,Landin J.Circulating tumor cell clustering shapes DNA methylation to enable metastasis seeding..Cell2019;176:98-112.e14 PMCID:PMC6363966

[36] Szczerba BM,Vetter M,Gkountela S.Neutrophils escort circulating tumour cells to enable cell cycle progression..Nature2019;566:553-7

[37] Labuschagne CF,Blagih J,Vousden KH.Cell clustering promotes a metabolic switch that supports metastatic colonization..Cell Metab2019;30:720-34.e5 PMCID:PMC6863392

[38] Liu X,Kawaguchi M,Chen W.Homophilic CD44 interactions mediate tumor cell aggregation and polyclonal metastasis in patient-derived breast cancer models..Cancer Discov2019;9:96-113 PMCID:PMC6328322

[39] Guo J,Lou Y,Xiao B.Detecting carcinoma cells in peripheral blood of patients with hepatocellular carcinoma by immunomagnetic beads and rt-PCR..J Clin Gastroenterol2007;41:783-8

[40] Shen Z,Chen X.Current detection technologies for circulating tumor cells..Chem Soc Rev2017;46:2038-56 PMCID:PMC5598784

[41] Joosse SA,Pantel K.Biology, detection, and clinical implications of circulating tumor cells..EMBO Mol Med2015;7:1-11 PMCID:PMC4309663

[42] Deng G,Burgess D,Krag D.Enrichment with anti-cytokeratin alone or combined with anti-EpCAM antibodies significantly increases the sensitivity for circulating tumor cell detection in metastatic breast cancer patients..Breast Cancer Res2008;10:R69 PMCID:PMC2575542

[43] Andree KC,Terstappen LW.Challenges in circulating tumor cell detection by the CellSearch system..Mol Oncol2016;10:395-407 PMCID:PMC5528971

[44] Yu D,Qiu Y,Wu Y.Identification and clinical significance of mobilized endothelial progenitor cells in tumor vasculogenesis of hepatocellular carcinoma..Clin Cancer Res2007;13:3814-24

[45] Yang ZF,Ho DW,Ng MN.Identification of local and circulating cancer stem cells in human liver cancer..Hepatology2008;47:919-28

[46] Liu S,Yu X,Cheng K.Expression of intercellular adhesion molecule 1 by hepatocellular carcinoma stem cells and circulating tumor cells..Gastroenterology2013;144:1031-41.e10

[47] Harouaka RA,Zheng SY.Circulating tumor cell enrichment based on physical properties..J Lab Autom2013;18:455-68 PMCID:PMC3830628

[48] Song Y,Shi Y,Zou Y.Enrichment and single-cell analysis of circulating tumor cells..Chem Sci2017;8:1736-51 PMCID:PMC5396552

[49] Gertler R,Fuehrer K,Nekarda H.Detection of circulating tumor cells in blood using an optimized density gradient centrifugation.2003;162:149

[50] Weitz J,Lacroix J,Benner A.Dissemination of tumor cells in patients undergoing surgery for colorectal cancer..Clin Cancer Res1998;4:343-8

[51] Rosenberg R,Friederichs J,Dahm M.Comparison of two density gradient centrifugation systems for the enrichment of disseminated tumor cells in blood..Cytometry2002;49:150-8

[52] Zuccolo J,Deans JP.Efficient isolation of highly purified tonsil B lymphocytes using RosetteSep with allogeneic human red blood cells..BMC Immunol2009;10:30 PMCID:PMC2695429

[53] Hayes GM,Voogt J,Gee TA.Isolation of malignant B cells from patients with chronic lymphocytic leukemia (CLL) for analysis of cell proliferation: validation of a simplified method suitable for multi-center clinical studies..Leuk Res2010;34:809-15 PMCID:PMC5253225

[54] Vona G,Louha M,Romana S.Isolation by size of epithelial tumor cells: a new method for the immunomorphological and molecular characterization of circulatingtumor cells..Am J Pathol2000;156:57-63 PMCID:PMC1868645

[55] Vona G,Beroud C,Capron F.Impact of cytomorphological detection of circulating tumor cells in patients with liver cancer..Hepatology2004;39:792-7

[56] Desitter I,Benali-Furet N,Janne PA.A new device for rapid isolation by size and characterization of rare circulating tumor cells..Anticancer Res2011;31:427-41

[57] Dong Y,Merdek KD,Jiang C.Microfluidics and circulating tumor cells..J Mol Diagn2013;15:149-57

[58] Nagrath S,Maheswaran S,Irimia D.Isolation of rare circulating tumour cells in cancer patients by microchip technology..Nature2007;450:1235-9 PMCID:PMC3090667

[59] Stott SL,Tsukrov DI,Miyamoto DT.Isolation of circulating tumor cells using a microvortex-generating herringbone-chip..Proc Natl Acad Sci U S A2010;107:18392-7 PMCID:PMC2972993

[60] Karabacak NM,Fachin F,Pai V.Microfluidic, marker-free isolation of circulating tumor cells from blood samples..Nat Protoc2014;9:694-710 PMCID:PMC4179254

[61] Sarioglu AF,Kojic N,Zeinali M.A microfluidic device for label-free, physical capture of circulating tumor cell clusters..Nat Methods2015;12:685-91 PMCID:PMC4490017

[62] Qian W,Chen W.Capturing cancer: emerging microfluidic technologies for the capture and characterization of circulating tumor cells..Small2015;11:3850-72

[63] Guo W,Shen MN,Wu J.Circulating tumor cells with stem-like phenotypes for diagnosis, prognosis, and therapeutic response evaluation in hepatocellular carcinoma..Clin Cancer Res2018;24:2203-13

[64] de Boer CJ,Janssen-van Rhijn CM.Expression of Ep-CAM in normal, regenerating, metaplastic, and neoplastic liver..J Pathol1999;188:201-6

[65] Yamashita T,Wang W,Ye Q.EpCAM and alpha-fetoprotein expression defines novel prognostic subtypes of hepatocellular carcinoma..Cancer Res2008;68:1451-61

[66] Nicolazzo C,Loreni F,Gazzaniga P.EpCAM(low) circulating tumor cells: gold in the waste..Dis Markers2019;2019:1718920 PMCID:PMC6766153

[67] Xu W,Chen L,Zhang XF.Isolation of circulating tumor cells in patients with hepatocellular carcinoma using a novel cell separation strategy..Clin Cancer Res2011;17:3783-93

[68] Li J,Zhang X,Liu H.Detection of circulating tumor cells in hepatocellular carcinoma using antibodies against asialoglycoprotein receptor, carbamoyl phosphate synthetase 1 and pan-cytokeratin..PLoS One2014;9:e96185 PMCID:PMC3999270

[69] Hamaoka M,Tanaka Y,Ohdan H.Clinical significance of glypican-3-positive circulating tumor cells of hepatocellular carcinoma patients: a prospective study..PLoS One2019;14:e0217586 PMCID:PMC6541303

[70] Court CM,Winograd P,Li QW.A novel multimarker assay for the phenotypic profiling of circulating tumor cells in hepatocellular carcinoma..Liver Transpl2018;24:946-60 PMCID:PMC6097911

[71] Guo W,Sun YF,Ma XL.Clinical significance of EpCAM mRNA-positive circulating tumor cells in hepatocellular carcinoma by an optimized negative enrichment and qRT-PCR-based platform..Clin Cancer Res2014;20:4794-805

[72] Kalinich M,Kwan TT,Javaid S.An RNA-based signature enables high specificity detection of circulating tumor cells in hepatocellular carcinoma..Proc Natl Acad Sci U S A2017;114:1123-8 PMCID:PMC5293050

[73] Liu YK,Li ZL,Li Y.An improved strategy to detect the epithelial-mesenchymal transition process in circulating tumor cells in hepatocellular carcinoma patients..Hepatol Int2016;10:640-6

[74] Wan S,Smith KJ,Park GS.New labyrinth microfluidic device detects circulating tumor cells expressing cancer stem cell marker and circulating tumor microemboli in hepatocellular carcinoma..Sci Rep2019;9:18575 PMCID:PMC6901480

[75] Hu B,Xu Y,Sun C.Systemic immune-inflammation index predicts prognosis of patients after curative resection for hepatocellular carcinoma..Clin Cancer Res2014;20:6212-22

[76] Wang BL,Gao XH,Wu J.Dynamic change of the systemic immune inflammation index predicts the prognosis of patients with hepatocellular carcinoma after curative resection..Clin Chem Lab Med2016;54:1963-9

[77] Schulze K,Staufer K,Lohse AW.Presence of EpCAM-positive circulating tumor cells as biomarker for systemic disease strongly correlates to survival in patients with hepatocellular carcinoma..Int J Cancer2013;133:2165-71

[78] Kelley RK,Butler TM,Hwang J.Circulating tumor cells in hepatocellular carcinoma: a pilot study of detection, enumeration, and next-generation sequencing in cases and controls..BMC Cancer2015;15:206 PMCID:PMC4399150

[79] Yu JJ,Dong SL,Zhang ZW.Effect of surgical liver resection on circulating tumor cells in patients with hepatocellular carcinoma..BMC Cancer2018;18:835 PMCID:PMC6102841

[80] von Felden J,Krech T,Nashan B.Circulating tumor cells as liquid biomarker for high HCC recurrence risk after curative liver resection..Oncotarget2017;8:89978-87 PMCID:PMC5685725

[81] Shen J,Zhu XL.High epithelial cell adhesion molecule-positive circulating tumor cell count predicts poor survival of patients with unresectable hepatocellular carcinoma treated with transcatheter arterial chemoembolization..J Vasc Interv Radiol2018;29:1678-84

[82] Yao F,Xu CF,Xiao BX.Detecting AFP mRNA in peripheral blood of the patients with hepatocellular carcinoma, liver cirrhosis and hepatitis..Clin Chim Acta2005;361:119-27

[83] Zhou Y,Wu J,Zhou Y.Association of preoperative EpCAM circulating tumor cells and peripheral Treg cell levels with early recurrence of hepatocellular carcinoma following radical hepatic resection..BMC Cancer2016;16:506 PMCID:PMC4955266

[84] Taketa K.Alpha-fetoprotein: reevaluation in hepatology..Hepatology1990;12:1420-32

[85] Trevisani F,Morselli-Labate AM,Accogli E.Serum alpha-fetoprotein for diagnosis of hepatocellular carcinoma in patients with chronic liver disease: influence of HBsAg and anti-HCV status..J Hepatol2001;34:570-5

[86] Fan ST,Ho DW,Yu WC.Prediction of posthepatectomy recurrence of hepatocellular carcinoma by circulating cancer stem cells: a prospective study..Ann Surg2011;254:569-76

[87] Li YM,Li J,Pi HF.Epithelial-mesenchymal transition markers expressed in circulating tumor cells in hepatocellular carcinoma patients with different stages of disease..Cell Death Dis2013;4:e831 PMCID:PMC3824657

[88] Chen J,Cai Z,Wang Q.Epithelial-mesenchymal transition phenotypes of circulating tumor cells correlate with the clinical stages and cancer metastasis in hepatocellular carcinoma patients..Cancer Biomark2017;20:487-98

[89] Ou H,Xiang L,Fang Y.Circulating tumor cell phenotype indicates poor survival and recurrence after surgery for hepatocellular carcinoma..Dig Dis Sci2018;63:2373-80

[90] Yin LC,Gao YX,Peng Q.Twist expression in circulating hepatocellular carcinoma cells predicts metastasis and prognoses..Biomed Res Int2018;2018:3789613 PMCID:PMC6038670

[91] Ye X,Han C,Shang L.Circulating tumor cells as a potential biomarker for postoperative clinical outcome in HBV-related hepatocellular carcinoma..Cancer Manag Res2018;10:5639-47 PMCID:PMC6245351

[92] Cheng Y,Zhang J,Tang Y.Diagnostic value of different phenotype circulating tumor cells in hepatocellular carcinoma..J Gastrointest Surg2019;23:2354-61

[93] Wang Z,Cheng Y,Peng B.Correlation between postoperative early recurrence of hepatocellular carcinoma and mesenchymal circulating tumor cells in peripheral blood..J Gastrointest Surg2018;22:633-9 PMCID:PMC5869875

[94] Qi LN,Wu FX,Zhong JH.Circulating tumor cells undergoing EMT provide a metric for diagnosis and prognosis of patients with hepatocellular carcinoma..Cancer Res2018;78:4731-44

[95] Chen Y,Li W,Zhang X.Circulating tumor cells undergoing EMT are poorly correlated with clinical stages or predictive of recurrence in hepatocellular carcinoma..Sci Rep2019;9:7084 PMCID:PMC6506548

[96] Wang S,Liu J,Zhou J.Analysis of circulating tumor cells in patients with hepatocellular carcinoma recurrence following liver transplantation..J Investig Med2018;66:1-6

[97] Xue F,Zhang Z,Zheng J.Application of a novel liquid biopsy in patients with hepatocellular carcinoma undergoing liver transplantation..Oncol Lett2018;15:5481-8 PMCID:PMC5840532

[98] Qi LN,Chen YY,Zhong JH.Outcomes of anatomical versus non-anatomical resection for hepatocellular carcinoma according to circulating tumour-cell status..Ann Med2020;52:21-31

[99] Fan ZC,Liu GD,Weng XF.Real-time monitoring of rare circulating hepatocellular carcinoma cells in an orthotopic model by in vivo flow cytometry assesses resection on metastasis..Cancer Res2012;72:2683-91

[100] Ha Y,Shim JE,Jun MJ.Circulating tumor cells are associated with poor outcomes in early-stage hepatocellular carcinoma: a prospective study..Hepatol Int2019;13:726-35

[101] Jin J,Zou L,Li S.AFP mRNA level in enriched circulating tumor cells from hepatocellular carcinoma patient blood samples is a pivotal predictive marker for metastasis..Cancer Lett2016;378:33-7

[102] Wu X,Yu H,Shan Y.The predictive values of serum dickkopf-1 and circulating tumor cells in evaluating the efficacy of transcatheter arterial chemoembolization treatment on hepatocellular carcinoma..Medicine (Baltimore)2019;98:e16579 PMCID:PMC6709003

[103] Li Y,Wang C,Zhou M.Impact of liver tumor percutaneous radiofrequency ablation on circulating tumor cells..Oncol Lett2018;16:2839-50 PMCID:PMC6096180

[104] Rau KM,Hsiao YC,Wang TM.Sequential circulating tumor cell counts in patients with locally advanced or metastatic hepatocellular carcinoma: monitoring the treatment response..J Clin Med2020;9: PMCID:PMC7019972

[105] Tayoun T,Oulhen M,Pawlikowska P.CTC-derived models: a window into the seeding capacity of circulating tumor cells (CTCs)..Cells2019;8: PMCID:PMC6829286

[106] Baccelli I,Riethdorf S,Schillert A.Identification of a population of blood circulating tumor cells from breast cancer patients that initiates metastasis in a xenograft assay..Nat Biotechnol2013;31:539-44

[107] Yan J,Wu X,Jiang J.Circulating tumor cells are correlated with disease progression and treatment response in an orthotopic hepatocellular carcinoma model..Cytometry A2015;87:1020-8

[108] Zhu L,Zhang W,Wang J.Application of EGFR inhibitor reduces circulating tumor cells during transcatheter arterial embolization..Clin Transl Oncol2018;20:639-46

[109] Li J,Zhang X,Yang Y.pERK/pAkt phenotyping in circulating tumor cells as a biomarker for sorafenib efficacy in patients with advanced hepatocellular carcinoma..Oncotarget2016;7:2646-59 PMCID:PMC4823061

[110] Zhang Y,Zhang J,Zheng L.Microfluidic chip for isolation of viable circulating tumor cells of hepatocellular carcinoma for their culture and drug sensitivity assay..Cancer Biol Ther2016;17:1177-87 PMCID:PMC5137491

[111] Aoki M,Kashiro A,Shimizu Y.Prospects for comprehensive analyses of circulating tumor cells in tumor biology..Cancers (Basel)2020;12: PMCID:PMC7281475

[112] D’Avola D,Martins-Filho SN,Labgaa I.High-density single cell mRNA sequencing to characterize circulating tumor cells in hepatocellular carcinoma..Sci Rep2018;8:1157 PMCID:PMC6070499