Using circulating tumor cells to advance precision medicine in prostate cancer

Abstract

The field of circulating tumor cell (CTC) enrichment has seen many emerging technologies in recent years, which have resulted in the identification and monitoring of clinically relevant, CTC-based biomarkers that can be analyzed routinely without invasive procedures. Several molecular platforms have been used to investigate the molecular profile of the disease, from high throughput gene expression analyses down to single cell biological dissection. The established presence of CTC heterogeneity nevertheless constitutes a challenge for cell isolation as the several subpopulations can potentially display different molecular characteristics; in this scenario, careful consideration must be given to the isolation approach, whereas methods that discriminate against certain subpopulations may result in the exclusion of CTCs that carry biological relevance. In the context of prostate cancer, CTC molecular interrogation can enable longitudinal monitoring of key biological features during treatment with substantial clinical impact, as several biomarkers could predict tumor response to AR signaling inhibitors (abiraterone, enzalutamide) or standard chemotherapy (taxanes). Thus, CTCs represent a valuable opportunity to personalize medicine in current clinical practice.

Keywords

Circulating tumor cells / liquid biopsy / prostate cancer / precision medicine / predictive biomarkers / targeted therapy

References

[1] Parker C,Heinrich D,O'Sullivan JM,Chodacki A,Logue J,Widmark A,Hoskin P,James ND,Syndikus I,Wedel S,Dall'Oglio M,Coleman R,O'Bryan-Tear CG,Garcia-Vargas J,Bruland OS,Investigators A.Alpha emitter radium-223 and survival in metastatic prostate cancer..N Engl J Med2013;369:213-23

[2] Beer TM,Rathkopf DE,Sternberg CN,Iversen P,Carles J,Davis ID,Evans CP,Joshua AM,Kimura G,Mansbach H,Noonberg SB,Phung D,Scher HI,Venner PM,Investigators P.Enzalutamide in metastatic prostate cancer before chemotherapy..N Engl J Med2014;371:424-33 PMCID:PMC4418931

[3] Sweeney CJ,Carducci M,Jarrard DF,Wong YN,Kohli M,Dreicer R,Picus J,Hussain M,DiPaola RS.Chemohormonal therapy in metastatic hormone-sensitive prostate cancer..N Engl J Med2015;373:737-46 PMCID:PMC4562797

[4] James ND,Clarke NW,Dearnaley DP,Ritchie AW,Russell JM,de Bono J,Jones RJ,Amos C,Millman R,Beesley S,Brock S,Chakraborti P,Cook A,Gale J,Graham JD,Hughes R,McKinna F,O'Sullivan JM,Peedell C,Robinson AJ,Srinivasan R,Sundar S,Tsang D,Parmar MK.Addition of docetaxel, zoledronic acid, or both to first-line long-term hormone therapy in prostate cancer (STAMPEDE): survival results from an adaptive, multiarm, multistage, platform randomised controlled trial..Lancet2016;387:1163-77

[5] James ND,Spears MR,Mason MD,Ritchie AWS,Gilson C,Matheson D,Attard G,Cross WR,Parker CC,Berthold DR,Adab F,Birtle AJ,Brock S,Ferguson C,Gray E,Hoskin PJ,Malik ZI,McPhail N,O'Sullivan J,Protheroe A,Srihari NN,Wagstaff J,Zarkar A,Sydes MR.Abiraterone for prostate cancer not previously treated with hormone therapy..N Engl J Med2017;377:338-51

[6] Fizazi K,Fein L,Rodriguez-Antolin A,Ozguroglu M,Feyerabend S,De Porre P,Park YC,Chi KN.Abiraterone plus prednisone in metastatic, castration-sensitive prostate cancer..N Engl J Med2017;377:352-60

[7] Galletti G,Tagawa ST,Giannakakou P.Circulating tumor cells in prostate cancer diagnosis and monitoring: an appraisal of clinical potential..Mol Diagn Ther2014;18:389-402 PMCID:PMC4149177

[8] Alix-Panabieres C.Challenges in circulating tumour cell research..Nat Rev Cancer2014;14:623-31

[9] Pantel K.Detection methods of circulating tumor cells..J Thorac Dis2012;4:446-7 PMCID:PMC3461061

[10] Miyamoto DT.Cell-free and circulating tumor cell-based biomarkers in men with metastatic prostate cancer: tools for real-time precision medicine?.Urol Oncol2016;34:490-501

[11] Heitzer E,Gasch C,Ulz P,Lax S,Mauermann O,Hofler G,Sill H,Pantel K,Bauernhofer T,Speicher MR.Complex tumor genomes inferred from single circulating tumor cells by array-CGH and next-generation sequencing..Cancer Res2013;73:2965-75

[12] Lohr JG,Cibulskis K,Rosenberg M,Francis JM,Shalek AK,Trombetta JJ,Tallapragada N,Kim S,Sougnez C,Wong B,Van Allen EM,Lis RT,Li T,Ly A,Clancy TE,Regev A,Hahn WC,Golub TR,Boehm JS.Whole-exome sequencing of circulating tumor cells provides a window into metastatic prostate cancer..Nat Biotechnol2014;32:479-84 PMCID:PMC4034575

[13] Millner LM,Valdes R, Jr..Circulating tumor cells: a review of present methods and the need to identify heterogeneous phenotypes..Ann Clin Lab Sci2013;43:295-304 PMCID:PMC5060940

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

[15] Kalluri R.The basics of epithelial-mesenchymal transition..J Clin Invest2009;119:1420-8 PMCID:PMC2689101

[16] Harouaka R,Zheng SY.Circulating tumor cells: advances in isolation and analysis, and challenges for clinical applications..Pharmacol Ther2014;141:209-21 PMCID:PMC3947247

[17] Marrinucci D,Kolatkar A,Malchiodi M,Voigt K,Nieva J,Ko AH,Schram E,Yang X,Lamy R,Yoshioka C,Petrova Y,Nelson D.Fluid biopsy in patients with metastatic prostate, pancreatic and breast cancers..Phys Biol2012;9:016003 PMCID:PMC3387996

[18] Nieva J,Luttgen MS,Bazhenova L,Santala R,Burke J,Bethel K.High-definition imaging of circulating tumor cells and associated cellular events in non-small cell lung cancer patients: a longitudinal analysis..Phys Biol2012;9:016004 PMCID:PMC3388002

[19] Werner SL,Landers M,Schroeder M,Bales N,Marrinucci D.Analytical validation and capabilities of the epic CTC platform: enrichment-free circulating tumour cell detection and characterization..J Circulating Biomarkers2015;4:3

[20] Anantharaman A,Lu D,Premasekharan G,Edwards M,Lindquist K,Jendrisak A,Dugan L,Wang Y,Paris PL.Programmed death-ligand 1 (PD-L1) characterization of circulating tumor cells (CTCs) in muscle invasive and metastatic bladder cancer patients..BMC Cancer2016;16:744 PMCID:PMC5034508

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

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

[23] Vona G,Louha M,Romana S,Capron F,Pazzagli M,Lacour B,Paterlini-Brechot P.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

[24] Farace F,Vimond N,Jacques N,Laplanche A,Lacroix L,Le Moulec S,Fizazi K,Vielh P.A direct comparison of CellSearch and ISET for circulating tumour-cell detection in patients with metastatic carcinomas..Br J Cancer2011;105:847-53 PMCID:PMC3171010

[25] Krebs MG,Carter L,Blackhall FH.Molecular analysis of circulating tumour cells-biology and biomarkers..Nat Rev Clin Oncol2014;11:129-44

[26] Hou JM,Ward T,Priest L,Clack G,Blackhall F.Circulating tumor cells as a window on metastasis biology in lung cancer..Am J Pathol2011;178:989-96 PMCID:PMC3069884

[27] Aceto N,Miyamoto DT,Wittner BS,Yu M,Engstrom A,Brannigan BW,Stott SL,Ramaswamy S,Lin CP,Haber DA.Circulating tumor cell clusters are oligoclonal precursors of breast cancer metastasis..Cell2014;158:1110-22 PMCID:PMC4149753

[28] Lazar DC,Luttgen MS,Uson ML,Gross ME.Cytometric comparisons between circulating tumor cells from prostate cancer patients and the prostate-tumor-derived LNCaP cell line..Phys Biol2012;9:016002 PMCID:PMC3387997

[29] Meunier A,Turner K,Veres T.Combination of mechanical and molecular filtration for enhanced enrichment of circulating tumor cells..Anal Chem2016;88:8510-7

[30] Allard WJ,Miller MC,Connelly MC,Tibbe AG,Terstappen LW.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

[31] Cristofanilli M,Ellis MJ,Matera J,Reuben JM,Allard WJ,Hayes DF.Circulating tumor cells, disease progression, and survival in metastatic breast cancer..N Engl J Med2004;351:781-91

[32] Cohen SJ,Iannotti N,Sabbath KD,Picus J,Mitchell E,Doyle GV,Terstappen LW.Relationship of circulating tumor cells to tumor response, progression-free survival, and overall survival in patients with metastatic colorectal cancer..J Clin Oncol2008;26:3213-21

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

[34] Talasaz AH,Huber DE,Roh KH,Xiao W,Pease RF,Jeffrey SS.Isolating highly enriched populations of circulating epithelial cells and other rare cells from blood using a magnetic sweeper device..Proc Natl Acad Sci U S A2009;106:3970-5 PMCID:PMC2645911

[35] Cann GM,Cooper S,Luo S,Stuart S,Srinivas S,Brooks JD.mRNA-Seq of single prostate cancer circulating tumor cells reveals recapitulation of gene expression and pathways found in prostate cancer..PLoS One2012;7:e49144 PMCID:PMC3492322

[36] Yu M,Wittner BS,Smas ME,Isakoff SJ,Wells MN,Concannon KF,Sequist LV,Sgroi D,Ramaswamy S,Haber DA.Circulating breast tumor cells exhibit dynamic changes in epithelial and mesenchymal composition..Science2013;339:580-4 PMCID:PMC3760262

[37] Capoun O,Jancikova M,Kolostova K,Michael P,Hanus T.Prognosis of castration-resistant prostate cancer patients - use of the AdnaTest® system for detection of circulating tumor cells..Anticancer Res2016;36:2019-26

[38] Todenhofer T,Feyerabend S,Mischinger J,Gerber V,Schilling D,Stenzl A.Preliminary experience on the use of the AdnaTest® system for detection of circulating tumor cells in prostate cancer patients..Anticancer Res2012;32:3507-13

[39] Signoretti S,Manola J,Tam C,Balk S,Kaplan I,Hahnfeldt P,Loda M.Her-2-neu expression and progression toward androgen independence in human prostate cancer..J Natl Cancer Inst2000;92:1918-25

[40] Scher HI.HER2 in prostate cancer -- a viable target or innocent bystander?.J Natl Cancer Inst2000;92:1866-8

[41] Hanssen A,Gorges TM,Uzunoglu FG,Stoecklein NH,Angenendt S,Atanackovic D,Riethdorf S,Wikman H.Characterization of different CTC subpopulations in non-small cell lung cancer..Sci Rep2016;6:28010 PMCID:PMC4908396

[42] Antonarakis ES,Wang H,Nakazawa M,Chen Y,Chen Y,Lotan TL,De Marzo AM,Isaacs WB,Paller CJ,Carducci MA,Luo J.AR-V7 and resistance to enzalutamide and abiraterone in prostate cancer..N Engl J Med2014;371:1028-38 PMCID:PMC4201502

[43] Nagrath S,Maheswaran S,Irimia D,Smith MR,Digumarthy S,Ryan P,Tompkins RG,Toner M.Isolation of rare circulating tumour cells in cancer patients by microchip technology..Nature2007;450:1235-9 PMCID:PMC3090667

[44] Stott SL,Tsukrov DI,Miyamoto DT,Rothenberg SM,Smas ME,Floyd FPJr,Lord JB,Springer S,Nagrath S,Lee RJ,Maheswaran S,Toner M.Isolation of circulating tumor cells using a microvortex-generating herringbone-chip..Proc Natl Acad Sci U S A2010;107:18392-7 PMCID:PMC2972993

[45] Gleghorn JP,Denning D,Bander NH,Nanus DM,Kirby BJ.Capture of circulating tumor cells from whole blood of prostate cancer patients using geometrically enhanced differential immunocapture (GEDI) and a prostate-specific antibody..Lab Chip2010;10:27-9 PMCID:PMC3031459

[46] Kirby BJ,Loftus MS,Pratt ED,Gleghorn JP,Liu H,Navarro VN,Bander NH,Giannakakou P.Functional characterization of circulating tumor cells with a prostate-cancer-specific microfluidic device..PLoS One2012;7:e35976 PMCID:PMC3338784

[47] Sperger JM,Welsh A,Chalmers Z,Heninger E,Tokar J,Guckenberger DJ,Dehm SM,Beebe DJ,Lang JM.Integrated analysis of multiple biomarkers from circulating tumor cells enabled by exclusion-based analyte isolation..Clin Cancer Res2016;

[48] Gerges N,Jabado N.New technologies for the detection of circulating tumour cells..Br Med Bull2010;94:49-64

[49] Hodgkinson CL,Li Y,Rothwell DG,Polanski R,Simpson KL,Pepper SD,Greystoke A,Bola B,Antonello J,Faulkner S,Carter L,Miller CJ,Brady G.Tumorigenicity and genetic profiling of circulating tumor cells in small-cell lung cancer..Nat Med2014;20:897-903

[50] Karabacak NM,Fachin F,Pai V,Martel JM,Smith K,Yang J,Morgan B,Barber TA,Maheswaran S,Haber DA.Microfluidic, marker-free isolation of circulating tumor cells from blood samples..Nat Protoc2014;9:694-710 PMCID:PMC4179254

[51] Saucedo-Zeni N,Niestroj R,Murawa D,Tomasi T,Dworacki G,Jansen H,Sterzynska K,Zabel M,Reuning U,Lucke K.A novel method for the in vivo isolation of circulating tumor cells from peripheral blood of cancer patients using a functionalized and structured medical wire..Int J Oncol2012;41:1241-50 PMCID:PMC3583719

[52] Theil G,Weber E,Hoda R,Fornara P.The use of a new cellcollector to isolate circulating tumor cells from the blood of patients with different stages of prostate cancer and clinical outcomes - a proof-of-concept study..PLoS One2016;11:e0158354 PMCID:PMC4968821

[53] Kuske A,Tennstedt P,Pompe R,Prues S,Markou A,Peine S,Riethdorf S,Schlomm T.Improved detection of circulating tumor cells in non-metastatic high-risk prostate cancer patients..Sci Rep2016;6:39736 PMCID:PMC5175156

[54] Drost J,Gao D,Sawyers CL,Clevers H.Organoid culture systems for prostate epithelial and cancer tissue..Nat Protoc2016;11:347-58 PMCID:PMC4793718

[55] Gao D,Sboner A,Karthaus WR,Dowling C,Undvall EA,Wongvipat J,Ramazanoglu S,Di W,Zhang QF,Ran L,Beltran H,Touijer KA,Laudone VP,Rathkopf DE,Danila DC,Solomon SB,Chi P,Rubin MA,Clevers H,Chen Y.Organoid cultures derived from patients with advanced prostate cancer..Cell2014;159:176-87 PMCID:PMC4237931

[56] Gupta S,Kemeny G,Beaver J,Ware KE,Armstrong AJ.Whole genomic copy number alterations in circulating tumor cells from men with abiraterone or enzalutamide resistant metastatic castration-resistant prostate cancer..Clin Cancer Res2017;23:1346-57

[57] Miyamoto DT,Wittner BS,Zhu H,Desai R,Brannigan BW,Arora KS,Dahl DM,Smith MR,Wu CL,Ramaswamy S,Toner M,Haber DA.RNA-Seq of single prostate CTCs implicates noncanonical Wnt signaling in antiandrogen resistance..Science2015;349:1351-6 PMCID:PMC4872391

[58] Watson PA,Sawyers CL.Emerging mechanisms of resistance to androgen receptor inhibitors in prostate cancer..Nat Rev Cancer2015;15:701-11 PMCID:PMC4771416

[59] Reyes EE,Isikbay M,Campanile A,Vander Griend DJ.Quantitative characterization of androgen receptor protein expression and cellular localization in circulating tumor cells from patients with metastatic castration-resistant prostate cancer..J Transl Med2014;12:313 PMCID:PMC4252013

[60] Crespo M,Ferraldeschi R,Sideris S,Bianchini D,Riisnaes R,Figueiredo I,Nowakowska K,Terstappen LW.Androgen receptor expression in circulating tumour cells from castration-resistant prostate cancer patients treated with novel endocrine agents..Br J Cancer2015;112:1166-74 PMCID:PMC4385957

[61] Miyamoto DT,Stott SL,Wittner BS,Smas ME,Brannigan BW,Bander NH,Sequist LV,Ramaswamy S,Maheswaran S.Androgen receptor signaling in circulating tumor cells as a marker of hormonally responsive prostate cancer..Cancer Discov2012;2:995-1003 PMCID:PMC3508523

[62] Darshan MS,Thadani-Mulero M,Escuin D,Gjyrezi A,Shen R,Bander NH,Giannakakou P.Taxane-induced blockade to nuclear accumulation of the androgen receptor predicts clinical responses in metastatic prostate cancer..Cancer Res2011;71:6019-29 PMCID:PMC3354631

[63] Antonarakis ES,Galletti G,Ballman K,Sonpavde G,Albany C,Stewart J,Szatrowski T,Gjyrezi A,Portella L,Lannin TB,Gruber CN,Kirby BJ,Nanus DM,Giannakakou P.Randomized, noncomparative, Phase II trial of early switch from docetaxel to cabazitaxel or vice versa, with integrated biomarker analysis, in men with chemotherapy-naive, metastatic, castration-resistant prostate cancer..J Clin Oncol2017;

[64] Guo Z,Sun F,Linn DE,Chen H,Melamed J,Kung HJ,Edwards J.A novel androgen receptor splice variant is up-regulated during prostate cancer progression and promotes androgen depletion-resistant growth..Cancer Res2009;69:2305-13 PMCID:PMC2672822

[65] Sun S,Vessella RL,Soriano K,Page ST,Nguyen HM,Nelson PS.Castration resistance in human prostate cancer is conferred by a frequently occurring androgen receptor splice variant..J Clin Invest2010;120:2715-30 PMCID:PMC2912187

[66] Li Y,Brand LJ,Silverstein KA.Androgen receptor splice variants mediate enzalutamide resistance in castration-resistant prostate cancer cell lines..Cancer Res2013;73:483-9 PMCID:PMC3549016

[67] Thadani-Mulero M,Sun S,Matov A,Corey E,Plymate SR.Androgen receptor splice variants determine taxane sensitivity in prostate cancer..Cancer Res2014;74:2270-82 PMCID:PMC4012562

[68] Antonarakis ES,Luber B,Chen Y,Nadal R,Denmeade SR,Eisenberger MA.Androgen receptor splice variant 7 and efficacy of taxane chemotherapy in patients with metastatic castration-resistant prostate cancer..JAMA Oncol2015;1:582-91 PMCID:PMC4537351

[69] Scher HI,Schreiber NA,Graf RP,Johnson A,Bambury R,McLaughlin B,Greene SB,Marrinucci D,Dittamore R.Association of AR-V7 on circulating tumor cells as a treatment-specific biomarker with outcomes and survival in castration-resistant prostate cancer..JAMA Oncol2016;2:1441-9 PMCID:PMC5206761

[70] Liu X,Li D,Steinberger A,Li J,Silberstein J,Dong Y,Zhang H.A whole blood assay for AR-V7 and ARv567es in patients with prostate cancer..J Urol2016;196:1758-63

[71] Qu F,Nakabayashi M,Jeong SH,Komura K,Sartor O,Kantoff PW.Association of AR-V7 and prostate specific antigen RNA levels in blood with efficacy of abiraterone acetate and enzalutamide treatment in men with prostate cancer..Clin Cancer Res2017;23:726-34

[72] Tomlins SA,Perner S,Mehra R,Varambally S,Tchinda J,Lee C,Shah RB,Rubin MA.Recurrent fusion of TMPRSS2 and ETS transcription factor genes in prostate cancer..Science2005;310:644-8

[73] Cancer Genome Atlas Research Network..The molecular taxonomy of primary prostate cancer..Cell2015;163:1011-25 PMCID:PMC4695400

[74] Kulda V,Kucera R,Srbecka K,Hes O,Babuska V,Benson V.Prognostic significance of TMPRSS2-ERG fusion gene in prostate cancer..Anticancer Res2016;36:4787-93

[75] McDaniel AS,Krupa R,Graf R,Jendrisak A,Marrinucci D,Flohr P,Crespo M,Mateo J,Dittamore R,Attard G.Phenotypic diversity of circulating tumour cells in patients with metastatic castration-resistant prostate cancer..BJU Int2016;

[76] Attard G,Olmos D,Vickers E,Levink R,Moreira J,Oommen NB,Jameson C,Sipkema R,Parker C,Kaye SB,Molina A,Terstappen LW.Characterization of ERG, AR and PTEN gene status in circulating tumor cells from patients with castration-resistant prostate cancer..Cancer Res2009;69:2912-8

[77] Attard G,Logothetis CJ,Mukherjee SD,Schrijvers D,Li W,Griffin TW,Ricci DS,Rathkopf DE,Ryan CJ.Improvements in radiographic progression-free survival stratified by erg gene status in metastatic castration-resistant prostate cancer patients treated with abiraterone acetate..Clin Cancer Res2015;21:1621-7 PMCID:PMC4384987

[78] Danila DC,Sung CC,Leversha MA,Lilja H,Sawyers CL,Scher HI.TMPRSS2-ERG status in circulating tumor cells as a predictive biomarker of sensitivity in castration-resistant prostate cancer patients treated with abiraterone acetate..Eur Urol2011;60:897-904 PMCID:PMC3185163

[79] Gaudreau PO,Soulieres D.The present and future of biomarkers in prostate cancer: proteomics, genomics, and immunology advancements..Biomark Cancer2016;8:15-33 PMCID:PMC4859450

[80] Galletti G,Beltran H,Miguel Mosquera J,MacDonald TY,O'Toole S,Suk Chae S,Tagawa ST,Rubin MA,Giannakakou P.ERG induces taxane resistance in castration-resistant prostate cancer..Nat Commun2014;5:5548 PMCID:PMC4244604

[81] Reig O,Carrera G,Pare L,Gaba L,Fernandez P,Mellado B.TMPRSS2-ERG in blood and docetaxel resistance in metastatic castration-resistant prostate cancer..Eur Urol2016;70:709-13

[82] Mitsiades N,Schultz N,He B,Fleisher M,Sawyers CL.Distinct patterns of dysregulated expression of enzymes involved in androgen synthesis and metabolism in metastatic prostate cancer tumors..Cancer Res2012;72:6142-52 PMCID:PMC3685485

[83] Wang Y,Chai K,Zhou BP.The role of snail in EMT and tumorigenesis..Curr Cancer Drug Targets2013;13:963-72 PMCID:PMC4004763

[84] Ware KE,Schaeffer D,Zhang T,Patierno SR,Garcia-Blanco MA.Snail promotes resistance to enzalutamide through regulation of androgen receptor activity in prostate cancer..Oncotarget2016;7:50507-21 PMCID:PMC5226599

[85] Li YM,Li J,Pi HF,Zuo GH,Li HY,Yu ZP,Liang P.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

[86] Chen CL,Osmulski P,Wang CM,Chang TC,Kumar AP,Gaczynska ME,Huang TH.Single-cell analysis of circulating tumor cells identifies cumulative expression patterns of EMT-related genes in metastatic prostate cancer..Prostate2013;73:813-26 PMCID:PMC4882087

[87] Beltran H,Landers M,Kossai M,Krupa R,Schreiber NA,Tagawa ST,Dittamore R.The initial detection and partial characterization of circulating tumor cells in neuroendocrine prostate cancer..Clin Cancer Res2016;22:1510-9 PMCID:PMC4990782

[88] Shaffer DR,Danila DC,Gonzalez-Espinoza R,Anand A,Maslak P,Terstappen LW,Heller G,Scher HI.Circulating tumor cell analysis in patients with progressive castration-resistant prostate cancer..Clin Cancer Res2007;13:2023-9

[89] Paris PL,Zhao Q,Sridharan S,Adler HL,Zarrabi MH,Simko J,Rosenberg J.Functional phenotyping and genotyping of circulating tumor cells from patients with castration resistant prostate cancer..Cancer Lett2009;277:164-73