Phenotypic Profiling of Circulating Tumor Cells in Metastatic Prostate Cancer Patients Using Nanoparticle-Mediated Ranking
- Brenda J. GreenBrenda J. GreenInstitute of Biomaterials and Biomedical Engineering, University of Toronto, 144 College Street, Toronto, Ontario M5S 3M2, CanadaMore by Brenda J. Green
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- Vivian NguyenVivian NguyenDepartment of Pharmaceutical Sciences, University of Toronto, Toronto M5S 3M2, CanadaMore by Vivian Nguyen
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- Eshetu AtenafuEshetu AtenafuPrincess Margaret Cancer Centre, University Health Network, Toronto, Ontario M5G 2C1, CanadaMore by Eshetu Atenafu
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- Phillip WeeberPhillip WeeberDepartment of Pharmaceutical Sciences, University of Toronto, Toronto M5S 3M2, CanadaMore by Phillip Weeber
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- Bill T. V. DuongBill T. V. DuongDepartment of Chemistry, University of Toronto, Toronto, Ontario M5S 3H6, CanadaMore by Bill T. V. Duong
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- Punithan ThiagalingamPunithan ThiagalingamDepartment of Pharmaceutical Sciences, University of Toronto, Toronto M5S 3M2, CanadaMore by Punithan Thiagalingam
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- Mahmoud LabibMahmoud LabibDepartment of Pharmaceutical Sciences, University of Toronto, Toronto M5S 3M2, CanadaMore by Mahmoud Labib
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- Reza M. MohamadiReza M. MohamadiDepartment of Pharmaceutical Sciences, University of Toronto, Toronto M5S 3M2, CanadaMore by Reza M. Mohamadi
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- Aaron R. HansenAaron R. HansenPrincess Margaret Cancer Centre, University Health Network, Toronto, Ontario M5G 2C1, CanadaMore by Aaron R. Hansen
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- Anthony M. Joshua*Anthony M. Joshua*E-mail: [email protected]Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario M5G 2C1, CanadaKinghorn Cancer Centre, St. Vincent’s Hospital Sydney, Darlinghurst, New South Wales 2010, AustraliaMore by Anthony M. Joshua
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- Shana O. Kelley*Shana O. Kelley*E-mail: [email protected]Institute of Biomaterials and Biomedical Engineering, University of Toronto, 144 College Street, Toronto, Ontario M5S 3M2, CanadaDepartment of Pharmaceutical Sciences, University of Toronto, Toronto M5S 3M2, CanadaDepartment of Chemistry, University of Toronto, Toronto, Ontario M5S 3H6, CanadaDepartment of Biochemistry, Faculty of Medicine, University of Toronto, Toronto, Ontario M5S 1A8, CanadaMore by Shana O. Kelley
Copyright © 2019 American Chemical Society
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Abstract
The analysis of circulating tumor cells (CTCs) provides a means to collect information about the evolving properties of a tumor during cancer progression and treatment. For patients with metastatic prostate cancer, noninvasive serial measurements of bloodborne cells may provide a means to tailor therapeutic decisions based on an individual patient’s response. Here, we used a high-sensitivity profiling approach to monitor CTCs in patients with metastatic castrate-resistant prostate cancer (mCRPC) undergoing treatment with abiraterone and enzalutamide, two drugs used to treat advanced prostate cancer. The capture and profiling approach uses antibody-functionalized magnetic nanoparticles to sort cells according to protein expression levels. CTCs are tagged with magnetic nanoparticles conjugated to an antibody specific for the epithelial cell adhesion molecule (EpCAM) and sorted into four zones of a microfluidic device based on EpCAM expression levels. Our approach was compared to the FDA-cleared CellSearch method, and we demonstrate significantly higher capture efficiency of low-EpCAM cells compared to the commercial method. The nanoparticle-based approach detected CTCs from 86% of patients at baseline, compared to CellSearch which only detected CTCs from 60% of patients. Patients were stratified as prostate specific antigen (PSA) progressive versus responsive based on clinically acceptable definitions, and it was observed that patients with a limited response to therapy had elevated levels of androgen receptor variant 7 (ARV7) and the mesenchymal marker, N-cadherin, expressed on their CTCs. In addition, these CTCs exhibited lower EpCAM expression. The results highlight features of CTCs associated with disease progression on abiraterone or enzalutamide, including mesenchymal phenotypes and increased expression levels of ARV7. The use of a high-sensitivity method to capture and profile CTCs provides more informative data concerning the phenotypic properties of these cells as patients undergo treatment relative to an FDA-cleared method.
This publication is licensed for personal use by The American Chemical Society.
Circulating tumor cells (CTCs) are released from a primary tumor into the bloodstream. (1) These rare cells are then responsible for formation of the metastatic lesions that lead to high rates of mortality. (2) CTCs can adopt resilient phenotypes in hostile nonadherent environments to avoid programmed cell death and immune attack. (1) During cancer progression, CTCs initially released into the bloodstream may possess an epithelial phenotype and express surface proteins such as cytokeratin and epithelial cell adhesion molecule (EpCAM). As the primary tumor progresses, CTCs can lose their epithelial markers, gain mesenchymal markers (N-cadherin, vimentin, fibronectin), and undergo an epithelial-to-mesenchymal transition (EMT). (3) EMT and its role in disease progression is not definitively understood, (3−6) but the plasticity exhibited by cells undergoing EMT may be linked to metastatic potential.
CTCs are obtained via a blood draw and can provide information relating to the molecular-level properties of a tumor without requiring invasive diagnostic techniques such as tissue biopsy. (7−9) Characterizing these rare cells in serial measurements, an approach referred to as liquid biopsy, (9) may therefore provide a noninvasive means to inform treatment options and monitor patient responsiveness. However, the capture and analysis of CTCs is very challenging due to their low concentration in blood. Effective capture requires a high level of specificity and the ability to handle very low numbers of target cells from a billion-fold excess of normal cells. Microfluidic sorting technologies have the potential to improve detection of heterogeneous CTC populations. Devices featuring EpCAM-functionalized-microposts and other nanomaterials, (10−12) basement-membrane coated chips, (13) discrete cell adhesion surfaces, (14) and lateral displacement microarrays have been developed. (15) Several recent studies have profiled heterogeneous CTC populations using antibody-functionalized microfluidic structures, including capture of epithelial and EMT-like CTCs by dual- targeting of both EpCAM/CD133 and EpCAM/stem-cell marker 63B6, or single- targeting of podoplanin. (16,51,52) The deformability of CTCs as they pass through microfluidic constrictions has also been used for the isolation of these cells (17,18) and for the isolation of clusters using bifurcating traps under low shear stress conditions. (19)
In our laboratory, we developed a nanoparticle-mediated capture and profiling approach that allows CTCs to be isolated from blood on the basis of markers including surface proteins (20) and mRNAs. (21) This approach exhibits high levels of sensitivity and can report on low levels of CTCs in patient samples, and the phenotypic and genotypic properties of these rare cells.
The analysis of CTCs in prostate cancer (PCa) patients is an important capability that may lead to new approaches for treatment monitoring. (8,22−26) PCa CTCs with an EMT phenotype have been identified in various studies, and the transition between epithelial-like and mesenchymal-like states has been linked to aggressive metastatic behavior. (27−31) CellSearch is the only FDA-cleared CTC assay presently and defines favorable or unfavorable CTC counts as <5 versus ≥5 CTC/7.5 mL of blood, respectively. (32)
Metastatic castrate resistant prostate cancer (mCRPC) is defined by biochemical, radiological, or symptomatic disease progression despite castrate testosterone level. (33) mCRPC usually retains continued androgen receptor (AR) expression and signaling, thus leading to continued dependence of AR. (34,35) Androgen deprivation therapy (ADT) is the main therapeutic approach for advanced prostate cancer, and it leads to prostate specific antigen (PSA) responses and clinical improvements in more than 90% of patients. (33) Two hormonal agents, abiraterone and enzalutamide, have demonstrated overall survival improvements in the setting of castration-resistant prostate cancer. (33,35−37) Abiraterone is a class of androgen-deprivation therapy that selectively and irreversibly inhibits the CYP17A1 enzyme, which is necessary for the synthesis of testosterone precursors in the adrenal gland. (24) Enzalutamide is an AR antagonist that binds AR with a high affinity and prevents AR translocation and DNA binding. (36) The de novo and acquired resistance to these agents may be associated with truncated AR-variants lacking the ligand binding domain (LBD). AR-variants have been detected in PCa cell lines as well as clinical samples, with the androgen receptor variant 7 (ARV7) splice variant representing the most commonly detected sequence with associated resistance. (38,39) Thus, tracking full-length androgen receptor and splice variants on mCRPC CTCs can provide relevant information to direct clinical decisions.
In this study, we captured and profiled CTCs from a cohort of mCRPC patients using a microfluidic device which stratifies cells into zones based on their expression levels of EpCAM. (40−42) Patients received either abiraterone or enzalutamide, and blood samples were analyzed from patients before treatment began (week 0 or baseline) and at a time point during treatment ranging from 9 to 22 weeks after the baseline sampling. We demonstrated that the microfluidic device captures low-EpCAM mCRPC CTCs with high efficiency relative to the commercially available CellSearch device. Over the course of treatment, patients’ CTCs exhibit decreasing EpCAM levels. These low-EpCAM CTCs display reduced epithelial properties and increased expression levels of ARV7. Progressive patients had elevated ARV7 and N-cadherin CTC protein levels relative to responsive patients, highlighting the distinct properties of resistant cells. Importantly, this study identifies subpopulations of CTCs detected from late-stage mCRPC patients which undergo phenotypic changes during enzalutamide or abiraterone treatment.
Results and Discussion
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Confirmation of High-Efficiency Capture of Low-EpCAM CTCs
CTCs are highly heterogeneous and can display variable levels of surface proteins. Our approach to capture and analyze these cells employs a microfluidic device which stratifies cells into zones based on their expression levels of EpCAM. (40) We previously demonstrate that this device is capable of detecting low-EpCAM breast and prostate cancer cells. (42,20) Prior to using the approach to test clinical samples, the performance of the device was characterized with cultured prostate cancer cells.
To analyze spiked cells or CTCs, blood is incubated with magnetic nanoparticles conjugated to EpCAM antibodies, and target cells are stoichiometrically labeled with nanoparticles according to the number of protein molecules on the surface of the cells. X-shaped microstructures patterned within a microfluidic channel are used to increase the CTC capture efficiency in the presence of a magnetic field by creating low-flow regions (Figure 1A,B). As the cells move through the device, they experience a decrease in velocity as the channel cross-section area expands. (40) The magnetic force applied to a cell is proportional to the number of magnetic nanoparticles bound and to the number of EpCAM molecules on the cell surface. Cells with higher EpCAM expression experience a higher magnetic force and are trapped in early zones, while cells with lower EpCAM experience a lower magnetic force and are trapped in later zones. This allows for the spatial sorting of cells on the basis of EpCAM expression, which is crucial for monitoring CTCs as they progress through EMT. (34,42)
Figure 1
Figure 1. High-efficiency capture of low-EpCAM CTCs from prostate cancer patients. (A,B) Schematic of patient sample collection and processing through the microfluidic device. CTCs are captured from metastatic castrate resistant prostate cancer (mCRPC) patients (n = 36) receiving enzalutamide or abiraterone. Briefly, whole blood is incubated with magnetic nanoparticles (MNPs) conjugated to EpCAM and introduced into the device at a flow rate of 600 μL/h in the presence of an external magnetic field. CTCs are trapped in different zones based on their expression level of EpCAM. (C) LnCaP and PC3 cells were profiled with the microfluidic device. (n = 292 LnCaP cells and n = 343 PC3 cells). High-EpCAM LnCaP cells are trapped primarily in zone 1, whereas low-EpCAM expressing PC3 cells are trapped in zones 3 and 4. (D) Capture efficiency of LnCaP and PC3 cells in the microfluidic device compared to CellSearch (n = 138 LnCaP cells and n = 96 PC3 cells). Cells were spiked in 1 mL of whole blood. Statistics are performed with two-tailed t test, *p < 0.05.
To verify the performance of this approach, the capture and profiling of two prostate cancer cells lines, LnCaP and PC3, were pursued. LnCaP cells have a high level of EpCAM protein expression, while PC3 cells have a lower level. (43) The level of expression for these cell lines was reflected in the capture profile observed, with LnCaP cells trapped in the earlier zones of the device and PC3 cells trapped in the later zones of the device (Figure 1C). Despite the difference in capture profiles and EpCAM expression levels, the overall capture efficiency for both cell lines was high (∼90%) and within error (Figure 1D).
The capture efficiency and sensitivity of the nanoparticle-mediated approach was compared to that achieved with the CellSearch technology (Figure 1D). While the capture efficiency of the LnCaP cells was only slightly lower using CellSearch versus the nanoparticle-based method, a marked decrease in capture efficiency was observed for the PC3 cells. This trend indicates that CTC recovery using CellSearch is sensitive to EpCAM levels. It is noteworthy that the ∼35% of PC3 cells captured using CellSearch likely correspond to the ∼30% of cells captured in zones 1 and 2 of the microfluidic device, indicating that the low-EpCAM cells in zones 3 and 4 are those not detectable using CellSearch.
Profiling CTCs for Patients with mCRPC
To test this approach on clinical specimens, CTCs were tracked from 36 mCRPC patients over a median treatment time of 13 weeks (Table 1). Primary treatment for the patients studied included prostatectomy (39%), radiation therapy (53%), brachytherapy (6%), or focal therapy (3%). All patients received prior androgen therapy, including Luteinizing Hormone Releasing Hormone (LHRH) agonists, antiandrogens, steroid, or immune therapy (Table S1). Patients did not receive prior treatment of abiraterone, enzalutamide, nor docetaxel. At the onset of the study, patients exhibited metastatic disease in the bone (50%) or lymph nodes (22%) or both (17%) (Tables S2 and S3).
Table 1. Patient Demographicsa
| patient characteristics | all patients |
|---|---|
| unique patients, no. | 36 |
| age, median (range), years | 72 (55–93) |
| Gleason score, median (range) | 7 (6–9) |
| median (range) on-treatment, weeks | 13 (9–22) |
| Primary Treatment, No. (%) | |
| prostatectomy | 6 (17) |
| radiation | 9 (25) |
| prostatectomy and radiation | 8 (22) |
| radiation and focal therapy | 1 (3) |
| radiation and brachytherapy | 1 (3) |
| brachytherapy | 1 (3) |
| none | 10 (27) |
| Prior Exposure to Life-Prolonging Therapies, No. (%) | |
| LHRH agonists | 1 (3) |
| LHRH agonists and antiandrogens | 26 (72) |
| LHRH agonists and steroid | 1 (3) |
| LHRH agonists and antiandrogens and steroid and/or immune therapy | 8 (22) |
| AR Therapy, No. (%) | |
| enzalutamide | 25 (69) |
| abiraterone acetate | 11 (31) |
| Metastatic Disease, No. (%) | |
| bone only | 18 (50) |
| LN only | 8 (22) |
| visceral only | 1 (3) |
| bone and LN | 6 (17) |
| bone and visceral and/or LN | 3 (8) |
| Laboratory Measures Pretherapy, Median (Range) | |
| PSA, μg/L | 16.46 (0.16–305.23) |
| Hgb, g/L | 132 (107–155) |
| ALP, U/L | 80 (50–381) |
| LDH, U/L | 227 (131–317) |
a
Abbreviations: LHRH agonists, luteinizing hormone releasing hormone, includes triptorelin, leuprolide, goserelin, or degarelix. AA, antiandrogens, includes bicalutamide, nilutamide, or apalutamide. Steroid treatment includes prednisone. Immune therapy includes prostvac. LN, lymph node; PSA, prostate specific antigen; Hgb, hemoglobin; ALP, alkaline phosphatase; and LDH, lactic dehydrogenase.
CTCs isolated from mCRPC patients were stratified as high-EpCAM (zone 1 and zone 2) and low-EpCAM (zone 3 and zone 4) (Figure 1B). Post-capture, CTCs were identified using immunostaining with anti-cytokeratin (Ck), anti-N-cadherin (NCad), anti-AR, anti-ARV7, and anti-CD45 fluorescently tagged antibodies in separate microfluidic devices.
mCRPC CTCs were primarily (88%) trapped in low-EpCAM zones 3 and 4 of the microfluidic device (Figure 2 and Figure 3). The capture profile corresponds with zone profiles of tumorigenic PC3 cells (Figure 1C, Figure S9). The microfluidic device detected CTCs from 86% of patients at baseline (week 0) and 77% of patients undergoing treatment (9–22 weeks). In comparison, CellSearch only detected CTCs from 60% of patients at baseline and in 18% of patients during treatment.
Figure 2
Figure 2. CTC profiles from prostate cancer patients. CTC counts were profiled with the microfluidic device (CTCs/mL) and compared with CellSearch (CTCs/mL) over (A) 0 weeks and (B) 9–22 weeks on-treatment. CTCs were identified as DAPI+/CK+/CD45–. Microfluidic device CTC counts were divided into CTCs captured in low-EpCAM zones 1 and 2 (light gray) and high-EpCAM zones 3 and 4 (black). CTC profiles were obtained from 36 mCRPC patients receiving enzalutamide or abiraterone. The horizontal dashed line indicates data is not available for a particular patient. No bar indicates 0 CTC count. The dotted line separates PSA responsive vs progressive patients, as defined according to PCWG3 criteria. (32) PSA response is defined as a >50% decline from baseline measured twice 3 to 4 weeks apart. PSA progression is defined as the first PSA rise that is ≥25% and ≥2 ng/mL above the nadir, confirmed 3 or more weeks later. If there is no initial decline from baseline, PSA progression is defined as ≥25% and ≥2 ng/mL after 12 weeks.
Figure 3
Figure 3. mCRPC CTCs exhibit a low-EpCAM phenotype. CTCs were profiled with the microfluidic device (CTCs/mL) and separated into (A) high-EpCAM (zone 1 and zone 2) and (B) low-EpCAM (zone 3 and zone 4) counts. (C) CellSearch CTC counts (CTCs/mL). Patients were profiled at 0 weeks and 9–22 weeks on-treatment. The dotted line separates PSA responsive vs progressive patients (refer to definition in the Materials and Methods). CTC counts were obtained from 36 mCRPC patients receiving abiraterone or enzalutamide. CTCs were identified as DAPI+/CK+/CD45–. Each dot represents a patient CTC count. Box plots represent standard error of the mean. The central square represents the mean, and the line represents the median.
Patients were stratified as PSA responsive or progressive based on the Prostate Cancer Clinical Trials Working Group criteria (PCWG3), (32) (Materials and Methods, Figures S10 and S11). The dynamic change of CTC counts over disease progression is associated with a significant prognostic effect and previously correlated with PSA response. (35,29,44) In our study, we observed a detectable increase in low-EpCAM CTCs in progressive patients between the baseline measurement and treatment time point (6 ± 2 and 11 ± 3 CTCs/mL, respectively; Figure 3B). The CellSearch analysis conducted with these same samples did not exhibit the same trend, and the overall counts were very low (≤1 CTCs/mL; Figure 3C, Figure S12). These results are consistent with prior demonstration that CellSearch does not effectively detect mesenchymal CTCs with an enhanced capacity for metastasis. (25)
Low-EpCAM CTCs Exhibit Increased ARV7 Levels and Reduced Epithelial Properties Associated with Progressive Patients
To complement the EpCAM-based profiling described above, we used our microfluidic approach to study other phenotypic markers. The presence of the ARV7 androgen receptor splice variant was monitored as a function of the microfluidic capture zone for patient CTCs as shown in Figure 4. While levels of cytokeratin, an epithelial marker, decreased in later zones, the levels of ARV7 increased (Figure 4A,B). Interestingly, this observation indicates that less epithelial CTCs may harbor truncated AR.
Figure 4
Figure 4. Low-EpCAM CTCs exhibit increased ARV7 levels and reduced epithelial properties. (A) Representative immunofluorescent images obtained from patient CTCs isolated from the four zones of the microfluidic device. CTCs are identified as DAPI+/CK+/ARV7+/CD45– and obtained with a 50× objective (CD45 image is not shown in the panel). (B,C) Relative fluorescent intensity of androgen receptor variant 7 (ARV7) and cytokeratin CTCs captured in the four zones of the microfluidic device (n = 76 CTCs, n = 104 CTCs, respectively). Fluorescent intensities are obtained using immunofluorescence image analysis. (D) Percentage of androgen receptor variant 7 (ARV7) CTCs: cytokeratin positive CTCs for responsive versus progressive patients. (E) Percentage of N-cadherin positive CTCs: cytokeratin positive CTCs for responsive versus progressive patients. Zone 3 and zone 4 counts were considered for CTC ratio analysis. CTCs are captured with EpCAM-MNPs in separate microfluidic devices: in the first device, CTCs are identified as DAPI+/CK+/CD45–; in the second device, CTCs are identified as DAPI+/CK+/ARV7+/CD45–; and in the third device, CTCs are identified as DAPI+/NCad+/CD45–. CTC counts are obtained at 0- and 9–22-weeks on-treatment. Statistics were performed with the Mann–Whitney test. *p < 0.05 is significant.
The percentage of CTCs captured in low-EpCAM zones significantly increased from 57% at baseline to 70% during treatment (Figure S4). The median zone values for baseline and 9–22 weeks of treatment are 3.3 CTCs/mL and 3.5 CTCs/mL, respectively, indicating a shift toward reduced EpCAM phenotype over the treatment period. Expression of EMT-related genes in CTCs may be associated with mCRPC, such as reduced cytokeratin levels and increased N-cadherin expression. (29) Using the microfluidic device, we have previously demonstrated that CTCs with low epithelial phenotypes exhibited invasive features, such as enhanced collagen uptake and folate-induced NAD(P)H metabolism. (41) In addition, we previously observed that CTCs in vivo were trapped in low-EpCAM zones, and mice receiving docetaxel chemotherapy exhibited a phenotypic shift in CTC profiles toward lower-EpCAM zones after 6 weeks of treatment. (45,46) Thus, the shift toward lower-EpCAM zones in mCRPC patients may suggest increased invasiveness of the residual tumor cells.
The proportion of ARV7-positive and N-cadherin-positive CTCs were assessed in progressive patients relative to responsive patients (Figure 4D,E). At baseline, the percentage of ARV7-positive CTCs relative to cytokeratin-positive CTCs was 52% and 26% for progressive vs responsive patients, respectively. In comparison, the baseline percentage of N-cadherin-positive CTCs relative to cytokeratin-positive CTCs was 213% and 66% for progressive vs responsive patients, respectively. AR-positive CTCs did not vary between progressive and responsive patients (80% and 79%, respectively) (Figure S7).
Progressive patients exhibit a higher incidence of bone metastases relative to responsive patients prior to the onset of abiraterone or enzalutamide treatment (67% vs 55%, respectively, Figure S1). The increased incidence of bone metastases prior to the administrative of abiraterone/enzalutamide may correlate with higher proportions of ARV7-positive and N-cadherin-positive CTCs detected in these patients. Increased incidence of ARV7 are considered an adaptive response to androgen deprivation therapy. (47) The two most accepted clinical assays for evaluating the presence of ARV7 in CTCs include the label-free immunofluorescent assay from Epic Sciences (47) and the mRNA-based AdnaTest platform. (48,49) Elevated ARV7 expression levels have been associated with resistance to treatment with enzalutamide or abiraterone, lower PSA response rates, and reduced overall survival. (50) Thus, tracking heterogeneous populations of CTCs at different stages of disease progression may significantly influence clinical decisions.
Conclusion
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We profiled CTCs from 36 mCRPC patients receiving abiraterone or enzalutamide over a median treatment time of 13 weeks. CTCs were captured in a microfluidic device which trapped low-EpCAM cells with high efficiency and sorted these cells into zones based on the expression level of EpCAM. Overall, the microfluidic device was capable of tracking low-EpCAM CTCs over the treatment period, which were largely missed by CellSearch. CTCs correlate with the responsiveness to treatment and shift toward lower EpCAM levels during patient treatment with abiraterone or enzalutamide. Progressive patients had increased percentages of N-cadherin-positive and ARV7-positive CTCs compared to responsive patients. Overall, this study identifies CTC biomarkers associated with disease-progressive patients and suggests that mesenchymal properties and increased ARV7 expression levels may be linked to sustained CTC presence in circulation. The ability to profile low-EpCAM CTC subtypes provides an important capability for monitoring patients with advanced cancers.
Supporting Information
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The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.analchem.9b01697.
Materials and Methods section along with supporting data and tables (PDF)
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Author Information
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- Shana O. Kelley - Institute of Biomaterials and Biomedical Engineering, University of Toronto, 144 College Street, Toronto, Ontario M5S 3M2, Canada; Department of Pharmaceutical Sciences, University of Toronto, Toronto M5S 3M2, Canada; Department of Chemistry, University of Toronto, Toronto, Ontario M5S 3H6, Canada; Department of Biochemistry, Faculty of Medicine, University of Toronto, Toronto, Ontario M5S 1A8, Canada;
http://orcid.org/0000-0003-3360-5359;
- Mahmoud Labib - Department of Pharmaceutical Sciences, University of Toronto, Toronto M5S 3M2, Canada;http://orcid.org/0000-0003-4565-2056
Acknowledgments
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We thank the ECTI facility at the University of Toronto for their clean room facilities. We also thank the Province of Ontario (Ontario Research Fund-Research Excellence), the Natural Sciences and Engineering Council, and the Canadian Institutes of Health Research for grants that supported this work.
References
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This article references 52 other publications.
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- 2Racila, E.; Euhus, D.; Weiss, A. J.; Rao, C.; McConnell, J.; Terstappen, L. W.; Uhr, J. W. Detection and characterization of carcinoma cells in the blood. Proc. Natl. Acad. Sci. U. S. A. 1998, 95 (8), 4589– 94, DOI: 10.1073/pnas.95.8.4589Google Scholar2https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADyaK1cXis1OgsLg%253D&md5=0f9c56c3060e225fdf39e0920e8aebc0Detection and characterization of carcinoma cells in the bloodRacila, Emilian; Euhus, David; Weiss, Arthur J.; Rao, Chandra; Mcconnell, John; Terstappen, Leon W. M. M.; Uhr, Jonathan W.Proceedings of the National Academy of Sciences of the United States of America (1998), 95 (8), 4589-4594CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)A highly sensitive assay combining immunomagnetic enrichment with multiparameter flow cytometric and immunocytochem. anal. has been developed to detect, enumerate, and characterize carcinoma cells in the blood. The assay can detect one epithelial cell or less in 1 mL of blood. Peripheral blood (10-20 mL) from 30 patients with carcinoma of the breast, from 3 patients with prostate cancer, and from 13 controls was examd. by flow cytometry for the presence of circulating epithelial cells defined as nucleic acid+, CD45-, and cytokeratin+. Highly significant differences in the no. of circulating epithelial cells were found between normal controls and patients with cancer including 17 with organ-confined disease. To det. whether the circulating epithelial cells in the cancer patients were neoplastic cells, cytospin prepns. were made after immunomagnetic enrichment and were analyzed. Epithelial cells from patients with breast cancer generally stained with mAbs against cytokeratin and 3 of 5 for mucin-1. In contrast, no cells that stained for these antigens were obsd. in the blood from normal controls. The morphol. of the stained cells was consistent with that of neoplastic cells. Of 8 patients with breast cancer followed for 1-10 mo, there was a good correlation between changes in the level of tumor cells in the blood with both treatment with chemotherapy and clin. status. The present assay may be helpful in early detection, in monitoring disease, and in prognostication.
- 3Mehlen, P.; Puisieux, A. Metastasis: a question of life or death. Nat. Rev. Cancer 2006, 6 (6), 449– 58, DOI: 10.1038/nrc1886Google Scholar3https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD28XkvVyltrY%253D&md5=394f27e4ce2c4643e306aeb92cddc9f8Metastasis: a question of life or deathMehlen, Patrick; Puisieux, AlainNature Reviews Cancer (2006), 6 (6), 449-458CODEN: NRCAC4; ISSN:1474-175X. (Nature Publishing Group)A review. The metastatic process is highly inefficient - very few of the many cells that migrate from the primary tumor successfully colonize distant sites. One proposed mechanism to explain this inefficiency is provided by the cancer stem cell model, which hypothesizes that micro-metastases can only be established by tumor stem cells, which are few in no. However, recent in vitro and in vivo observations indicate that apoptosis is an important process regulating metastasis. Here we stress that the inhibition of cell death, apart from its extensively described function in primary tumor development, is a crucial characteristic of metastatic cancer cells.
- 4Yu, M.; Bardia, A.; Wittner, B. S.; Stott, S. L.; Smas, M. E.; Ting, D. T.; Isakoff, S. J.; Ciciliano, J. C.; Wells, M. N.; Shah, A. M.; Concannon, K. F.; Donaldson, M. C.; Sequist, L. V.; Brachtel, E.; Sgroi, D.; Baselga, J.; Ramaswamy, S.; Toner, M.; Haber, D. A.; Maheswaran, S. Circulating breast tumor cells exhibit dynamic changes in epithelial and mesenchymal composition. Science 2013, 339 (6119), 580– 4, DOI: 10.1126/science.1228522Google Scholar4https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhsFGku7o%253D&md5=e9624550d2af76db63e5a62edb19d65dCirculating breast tumor cells exhibit dynamic changes in epithelial and mesenchymal compositionYu, Min; Bardia, Aditya; Wittner, Ben S.; Stott, Shannon L.; Smas, Malgorzata E.; Ting, David T.; Isakoff, Steven J.; Ciciliano, Jordan C.; Wells, Marissa N.; Shah, Ajay M.; Concannon, Kyle F.; Donaldson, Maria C.; Sequist, Lecia V.; Brachtel, Elena; Sgroi, Dennis; Baselga, Jose; Ramaswamy, Sridhar; Toner, Mehmet; Haber, Daniel A.; Maheswaran, ShyamalaScience (Washington, DC, United States) (2013), 339 (6119), 580-584CODEN: SCIEAS; ISSN:0036-8075. (American Association for the Advancement of Science)Epithelial-mesenchymal transition (EMT) of adherent epithelial cells to a migratory mesenchymal state has been implicated in tumor metastasis in preclin. models. To investigate its role in human cancer, we characterized EMT in circulating tumor cells (CTCs) from breast cancer patients. Rare primary tumor cells simultaneously expressed mesenchymal and epithelial markers, but mesenchymal cells were highly enriched in CTCs. Serial CTC monitoring in 11 patients suggested an assocn. of mesenchymal CTCs with disease progression. In an index patient, reversible shifts between these cell fates accompanied each cycle of response to therapy and disease progression. Mesenchymal CTCs occurred as both single cells and multicellular clusters, expressing known EMT regulators, including transforming growth factor (TGF)-β pathway components and the FOXC1 transcription factor. These data support a role for EMT in the blood-borne dissemination of human breast cancer.
- 5Jin, X.; Mu, P. Targeting Breast Cancer Metastasis. Breast Cancer: Basic Clin. Res. 2015, 9s1, 23– 34, DOI: 10.4137/BCBCR.S25460Google ScholarThere is no corresponding record for this reference.
- 6Green, B. J.; Panagiotakopoulou, M.; Pramotton, F. M.; Stefopoulos, G.; Kelley, S. O.; Poulikakos, D.; Ferrari, A. Pore shape defines paths of metastatic cell migration. Nano Lett. 2018, 18, 2140, DOI: 10.1021/acs.nanolett.8b00431Google Scholar6https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXjtlOnur8%253D&md5=c2bc204cfa33e9f955ff9ddf7ac9f537Pore shape defines paths of metastatic cell migrationGreen, Brenda J.; Panagiotakopoulou, Magdalini; Pramotton, Francesca Michela; Stefopoulos, Georgios; Kelley, Shana O.; Poulikakos, Dimos; Ferrari, AldoNano Letters (2018), 18 (3), 2140-2147CODEN: NALEFD; ISSN:1530-6984. (American Chemical Society)Invasion of dense tissues by cancer cells involves the interplay between the penetration resistance offered by interstitial pores and the deformability of cells. Metastatic cancer cells find optimal paths of minimal resistance through an adaptive path-finding process, which leads to successful dissemination. The phys. limit of nuclear deformation is related to the minimal cross section of pores that can be successfully penetrated. However, this single biophys. parameter does not fully describe the architectural complexity of tissues featuring pores of variable area and shape. Here, employing laser nanolithog., we fabricate pore microenvironment models with well-controlled pore shapes, through which human breast cells (MCF10A) and their metastatic offspring (MCF10CA1a.cl1) could pervade. In these exptl. settings, we demonstrate that the actual pore shape, and not only the cross section, is a major and independent determinant of cancer penetration efficiency. In complex architectures contg. pores demanding large deformations from invading cells, tall and narrow rectangular openings facilitate cancer migration. In addn., we highlight the characteristic traits of the explorative behavior enabling metastatic cells to identify and select such pore shapes in a complex multishape pore environment, pinpointing paths of least resistance to invasion.
- 7Green, B. J.; Saberi Safaei, T.; Mepham, A.; Labib, M.; Mohamadi, R. M.; Kelley, S. O. Beyond the Capture of Circulating Tumor Cells: Next-Generation Devices and Materials. Angew. Chem., Int. Ed. 2016, 55 (4), 1252– 65, DOI: 10.1002/anie.201505100Google Scholar7https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhvFKiurvM&md5=44ad3d40a3090e30b791c01cec97beefBeyond the Capture of Circulating Tumor Cells: Next-Generation Devices and MaterialsGreen, Brenda J.; Saberi Safaei, Tina; Mepham, Adam; Labib, Mahmoud; Mohamadi, Reza M.; Kelley, Shana O.Angewandte Chemie, International Edition (2016), 55 (4), 1252-1265CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)A review. Over the last decade, significant progress has been made towards the development of approaches that enable the capture of rare circulating tumor cells (CTCs) from the blood of cancer patients, a crit. capability for noninvasive tumor profiling. These advances have leveraged new insights in materials chem. and microfluidics and allowed the capture and enumeration of CTCs with unprecedented sensitivity. However, it has become increasingly clear that simply capturing and counting tumor cells launched into the bloodstream may not provide the information needed to advance our understanding of the biol. of these rare cells, or to allow us to better exploit them in medicine. A variety of advances have now emerged demonstrating that more information can be extd. from CTCs with next-generation devices and materials featuring tailored phys. and chem. properties. In this Minireview, the last ten years of work in this area will be discussed, with an emphasis on the groundbreaking work of the last five years, during which the focus has moved beyond the simple capture of CTCs and gravitated towards approaches that enable in-depth anal.
- 8Poudineh, M.; Sargent, E. H.; Pantel, K.; Kelley, S. O. Profiling circulating tumour cells and other biomarkers of invasive cancers. Nat. Biomed. Eng. 2018, 2 (2), 72– 84, DOI: 10.1038/s41551-018-0190-5Google Scholar8https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1MXhtFegu7fN&md5=2dff83b64b0f2b1feaeeed716a21eb27Profiling circulating tumour cells and other biomarkers of invasive cancersPoudineh, Mahla; Sargent, Edward H.; Pantel, Klaus; Kelley, Shana O.Nature Biomedical Engineering (2018), 2 (2), 72-84CODEN: NBEAB3; ISSN:2157-846X. (Nature Research)A review. During cancer progression, many tumors shed circulating tumor cells (CTCs) and other biomarkers into the bloodstream. The anal. of CTCs offers the prospect of collecting a liq. biopsy from a patients blood to predict and monitor therapeutic responses and tumor recurrence. In this Review, we discuss progress towards the isolation and recovery of bulk CTCs from whole blood samples for the identification of cells with high metastatic potential. We provide an overview of the techniques that initially pointed to the clin. significance of CTCs and describe the key requirements for clin. applications of CTC anal. We also summarize recent advances that permit the functional and biochem. phenotypes of CTCs to be characterized, and discuss the potential roles of these CTC characteristics in the formation of metastatic lesions. Moreover, we discuss the use of circulating tumor DNA and exosomes as markers for early cancer diagnosis and for the monitoring of cancer progression. Next-generation technologies and biomarkers for invasive cancers should allow for the unequivocal detn. of the metastatic potential of CTCs, and for the meaningful anal. of circulating tumor DNA and exosomes.
- 9Alix-Panabieres, C.; Pantel, K. Challenges in circulating tumour cell research. Nat. Rev. Cancer 2014, 14 (9), 623– 31, DOI: 10.1038/nrc3820Google Scholar9https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhsVWktrfM&md5=c75b540d167fe120266521b8cbf9a6c6Challenges in circulating tumour cell researchAlix-Panabieres, Catherine; Pantel, KlausNature Reviews Cancer (2014), 14 (9), 623-631CODEN: NRCAC4; ISSN:1474-175X. (Nature Publishing Group)A review. During the past ten years, circulating tumor cells (CTCs) have received enormous attention as new biomarkers and the subject of basic research. Although CTCs are already used in numerous clin. trials, their clin. utility is still under investigation. Many issues regarding the detection and characterization of CTCs remain unknown. In this Opinion article, we propose a conceptual framework of CTC assays and point out current challenges of CTC research, which might structure this dynamic field of translational cancer research.
- 10Nagrath, S.; Sequist, L. V.; Maheswaran, S.; Bell, D. W.; Irimia, D.; Ulkus, L.; Smith, M. R.; Kwak, E. L.; Digumarthy, S.; Muzikansky, A.; Ryan, P.; Balis, U. J.; Tompkins, R. G.; Haber, D. A.; Toner, M. Isolation of rare circulating tumour cells in cancer patients by microchip technology. Nature 2007, 450 (7173), 1235– 9, DOI: 10.1038/nature06385Google Scholar10https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2sXhsVGjsL%252FK&md5=499fb32c8eb07f12f4d1298f0557ea30Isolation of rare circulating tumour cells in cancer patients by microchip technologyNagrath, Sunitha; Sequist, Lecia V.; Maheswaran, Shyamala; Bell, Daphne W.; Irimia, Daniel; Ulkus, Lindsey; Smith, Matthew R.; Kwak, Eunice L.; Digumarthy, Subba; Muzikansky, Alona; Ryan, Paula; Balis, Ulysses J.; Tompkins, Ronald G.; Haber, Daniel A.; Toner, MehmetNature (London, United Kingdom) (2007), 450 (7173), 1235-1239CODEN: NATUAS; ISSN:0028-0836. (Nature Publishing Group)Viable tumor-derived epithelial cells (circulating tumor cells or CTCs) have been identified in peripheral blood from cancer patients and are probably the origin of intractable metastatic disease. Although extremely rare, CTCs represent a potential alternative to invasive biopsies as a source of tumor tissue for the detection, characterization and monitoring of nonhematol. cancers. The ability to identify, isolate, propagate and molecularly characterize CTC subpopulations could further the discovery of cancer stem cell biomarkers and expand the understanding of the biol. of metastasis. Current strategies for isolating CTCs are limited to complex analytic approaches that generate very low yield and purity. Here the authors describe the development of a unique microfluidic platform (the 'CTC-chip') capable of efficient and selective sepn. of viable CTCs from peripheral whole blood samples, mediated by the interaction of target CTCs with antibody (EpCAM)-coated microposts under precisely controlled laminar flow conditions, and without requisite pre-labeling or processing of samples. The CTC-chip successfully identified CTCs in the peripheral blood of patients with metastatic lung, prostate, pancreatic, breast and colon cancer in 115 of 116 (99%) samples, with a range of 5-1,281 CTCs per mL and approx. 50% purity. In addn., CTCs were isolated in 7/7 patients with early-stage prostate cancer. Given the high sensitivity and specificity of the CTC-chip, the authors tested its potential utility in monitoring response to anti-cancer therapy. In a small cohort of patients with metastatic cancer undergoing systemic treatment, temporal changes in CTC nos. correlated reasonably well with the clin. course of disease as measured by std. radiog. methods. Thus, the CTC-chip provides a new and effective tool for accurate identification and measurement of CTCs in patients with cancer. It has broad implications in advancing both cancer biol. research and clin. cancer management, including the detection, diagnosis and monitoring of cancer.
- 11Stott, S. L.; Hsu, C. H.; Tsukrov, D. I.; Yu, M.; Miyamoto, D. T.; Waltman, B. A.; Rothenberg, S. M.; Shah, A. M.; Smas, M. E.; Korir, G. K.; Floyd, F. P., Jr.; Gilman, A. J.; Lord, J. B.; Winokur, D.; Springer, S.; Irimia, D.; Nagrath, S.; Sequist, L. V.; Lee, R. J.; Isselbacher, K. J.; Maheswaran, S.; Haber, D. A.; Toner, M. Isolation of circulating tumor cells using a microvortex-generating herringbone-chip. Proc. Natl. Acad. Sci. U. S. A. 2010, 107 (43), 18392– 7, DOI: 10.1073/pnas.1012539107Google Scholar11https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXhtl2ktr3N&md5=f0896b84a56d0dcdcdb0207be71650ceIsolation of circulating tumor cells using a microvortex-generating herringbone-chipStott, Shannon L.; Hsua, Chia-Hsien; Tsukrov, Dina I.; Yu, Min; Miyamoto, David T.; Waltman, Belinda A.; Rothenberg, S. Michael; Shah, Ajay M.; Smas, Malgorzata E.; Korir, George K.; Floyd, Frederick P., Jr.; Gilman, Anna J.; Lord, Jenna B.; Winokur, Daniel; Springer, Simeon; Irimia, Daniel; Nagrath, Sunitha; Sequist, Lecia V.; Lee, Richard J.; Isselbacher, Kurt J.; Maheswaran, Shyamala; Haber, Daniel A.; Toner, MehmetProceedings of the National Academy of Sciences of the United States of America (2010), 107 (43), 18392-18397, S18392/1-S18392/9CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)Rare circulating tumor cells (CTCs) present in the bloodstream of patients with cancer provide a potentially accessible source for detection, characterization, and monitoring of nonhematol. cancers. We previously demonstrated the effectiveness of a microfluidic device, the CTC-Chip, in capturing these epithelial cell adhesion mol. (EpCAM)-expressing cells using antibody-coated microposts. Here, we describe a high-throughput microfluidic mixing device, the herringbone-chip, or "HB-Chip," which provides an enhanced platform for CTC isolation. The HB-Chip design applies passive mixing of blood cells through the generation of microvortices to significantly increase the no. of interactions between target CTCs and the antibody-coated chip surface. Efficient cell capture was validated using defined nos. of cancer cells spiked into control blood, and clin. utility was demonstrated in specimens from patients with prostate cancer. CTCs were detected in 14 of 15 (93%) patients with metastatic disease (median = 63 CTCs/mL, mean = 386 ± 238 CTCs/mL), and the tumor-specific TMPRSS2-ERG translocation was readily identified following RNA isolation and RT-PCR anal. The use of transparent materials allowed for imaging of the captured CTCs using std. clin. histopathol. stains, in addn. to immunofluorescence-conjugated antibodies. In a subset of patient samples, the low shear design of the HB-Chip revealed microclusters of CTCs, previously unappreciated tumor cell aggregates that may contribute to the hematogenous dissemination of cancer.
- 12Yoon, H. J.; Kim, T. H.; Zhang, Z.; Azizi, E.; Pham, T. M.; Paoletti, C.; Lin, J.; Ramnath, N.; Wicha, M. S.; Hayes, D. F.; Simeone, D. M.; Nagrath, S. Sensitive capture of circulating tumour cells by functionalized graphene oxide nanosheets. Nat. Nanotechnol. 2013, 8 (10), 735– 41, DOI: 10.1038/nnano.2013.194Google Scholar12https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhsFaksbzL&md5=a6ac26c2fdf8b2e6c93bd63f6634a934Sensitive capture of circulating tumor cells by functionalized graphene oxide nanosheetsYoon, Hyeun Joong; Kim, Tae Hyun; Zhang, Zhuo; Azizi, Ebrahim; Pham, Trinh M.; Paoletti, Costanza; Lin, Jules; Ramnath, Nithya; Wicha, Max S.; Hayes, Daniel F.; Simeone, Diane M.; Nagrath, SunithaNature Nanotechnology (2013), 8 (10), 735-741CODEN: NNAABX; ISSN:1748-3387. (Nature Publishing Group)The spread of cancer throughout the body is driven by circulating tumor cells (CTCs). These cells detach from the primary tumor and move from the bloodstream to a new site of subsequent tumor growth. They also carry information about the primary tumor and have the potential to be valuable biomarkers for disease diagnosis and progression, and for the mol. characterization of certain biol. properties of the tumor. However, the limited sensitivity and specificity of current methods for measuring and studying these cells in patient blood samples prevents the realization of their full clin. potential. The use of microfluidic devices is a promising method for isolating CTCs. However, the devices are reliant on three-dimensional structures, which limits further characterization and expansion of cells on the chip. Here we demonstrate an effective approach to isolating CTCs from blood samples of pancreatic, breast and lung cancer patients, by using functionalized graphene oxide nanosheets on a patterned gold surface. CTCs were captured with high sensitivity at a low concn. of target cells (73 ± 32.4% at 3-5 cells per mL blood).
- 13Zhang, Y.; Wu, M.; Han, X.; Wang, P.; Qin, L. High-Throughput, Label-Free Isolation of Cancer Stem Cells on the Basis of Cell Adhesion Capacity. Angew. Chem., Int. Ed. 2015, 54 (37), 10838– 42, DOI: 10.1002/anie.201505294Google Scholar13https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhtFyqtb%252FK&md5=f328026d6af1e22c6b23800d804e0559High-Throughput, Label-Free Isolation of Cancer Stem Cells on the Basis of Cell Adhesion CapacityZhang, Yuanqing; Wu, Minhao; Han, Xin; Wang, Ping; Qin, LidongAngewandte Chemie, International Edition (2015), 54 (37), 10838-10842CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)Herein the authors report a microfluidics method that enriches cancer stem cells (CSCs) or tumor-initiating cells on the basis of cell adhesion properties. In the authors' on-chip enrichment system, cancer cells were driven by hydrodynamic forces to flow through microchannels coated with basement membrane ext. Highly adhesive cells were captured by the functionalized microchannels, and less adhesive cells were collected from the outlets. Two heterogeneous breast cancer cell lines (SUM-149 and SUM-159) were successfully sepd. into enriched subpopulations according to their adhesive capacity, and the enrichment of the cancer stem cells was confirmed by flow cytometry biomarker anal. and tumor-formation assays. The authors' findings show that the less adhesive phenotype is assocd. with a higher percentage of CSCs, higher cancer-cell motility, and higher resistance to chemotherapeutic drugs.
- 14Witek, M. A.; Aufforth, R. D.; Wang, H.; Kamande, J. W.; Jackson, J. M.; Pullagurla, S. R.; Hupert, M. L.; Usary, J.; Wysham, W. Z.; Hilliard, D.; Montgomery, S.; Bae-Jump, V.; Carey, L. A.; Gehrig, P. A.; Milowsky, M. I.; Perou, C. M.; Soper, J. T.; Whang, Y. E.; Yeh, J. J.; Martin, G.; Soper, S. A. Discrete microfluidics for the isolation of circulating tumor cell subpopulations targeting fibroblast activation protein alpha and epithelial cell adhesion molecule. npj Precis. Oncol. 2017, 1, 24, DOI: 10.1038/s41698-017-0028-8Google ScholarThere is no corresponding record for this reference.
- 15Liu, Z.; Lee, Y.; Jang, J.; Li, Y.; Han, X.; Yokoi, K.; Ferrari, M.; Zhou, L.; Qin, L. Microfluidic cytometric analysis of cancer cell transportability and invasiveness. Sci. Rep. 2015, 5, 14272, DOI: 10.1038/srep14272Google Scholar15https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhsFOktr%252FE&md5=67492ced562b1ce9c51d65ab4f285772Microfluidic cytometric analysis of cancer cell transportability and invasivenessLiu, Zongbin; Lee, Yeonju; Jang, Joon hee; Li, Ying; Han, Xin; Yokoi, Kenji; Ferrari, Mauro; Zhou, Ledu; Qin, LidongScientific Reports (2015), 5 (), 14272CODEN: SRCEC3; ISSN:2045-2322. (Nature Publishing Group)The extensive phenotypic and functional heterogeneity of cancer cells plays an important role in tumor progression and therapeutic resistance. Characterizing this heterogeneity and identifying invasive phenotype may provide possibility to improve chemotherapy treatment. By mimicking cancer cell perfusion through circulatory system in metastasis, we develop a unique microfluidic cytometry (MC) platform to sep. cancer cells at high throughput, and further derive a phys. parameter 'transportability' to characterize the ability to pass through micro-constrictions. The transportability is detd. by cell stiffness and cell-surface frictional property, and can be used to probe tumor heterogeneity, discriminate more invasive phenotypes and correlate with biomarker expressions in breast cancer cells. Decreased cell stiffness and cell-surface frictional force leads to an increase in transportability and may be a feature of invasive cancer cells by promoting cell perfusion through narrow spaces in circulatory system. The MC-Chip provides a promising microfluidic platform for studying cell mechanics and transportability could be used as a novel marker for probing tumor heterogeneity and detg. invasive phenotypes.
- 16Yoneda, K.; Kuwata, T.; Chikaishi, Y.; Mori, M.; Kanayama, M.; Takenaka, M.; Oka, S.; Hirai, A.; Imanishi, N.; Kuroda, K.; Ichiki, Y.; Ohnaga, T.; Tanaka, F. Detection of circulating tumor cells with a novel microfluidic system in malignant pleural mesothelioma. Cancer Sci. 2019, 110 (2), 726– 733, DOI: 10.1111/cas.13895Google Scholar16https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1MXks1WmtQ%253D%253D&md5=21dfae540fa78ad82545b48665c809a0Detection of circulating tumor cells with a novel microfluidic system in malignant pleural mesotheliomaYoneda, Kazue; Kuwata, Taiji; Chikaishi, Yasuhiro; Mori, Masataka; Kanayama, Masatoshi; Takenaka, Masaru; Oka, Soichi; Hirai, Ayako; Imanishi, Naoko; Kuroda, Koji; Ichiki, Yoshinobu; Ohnaga, Takashi; Tanaka, FumihiroCancer Science (2019), 110 (2), 726-733CODEN: CSACCM; ISSN:1349-7006. (Wiley-Blackwell)Detection of rare tumor cells circulating in the blood (CTCs) presents tech. challenges. CellSearch, the only approved system for clin. use, fails to capture epithelial cell adhesion mol.-neg. CTCs such as malignant pleural mesothelioma (MPM). We have developed a novel microfluidic device (CTC-chip) in which any Ab to capture CTCs is conjugated. The CTC-chip was coated with an Ab against podoplanin that is abundantly expressed on MPM. Circulating tumor cell-detection performance was evaluated in exptl. models in which MPM cells were spiked in blood sampled from a healthy volunteer and in clin. samples drawn from MPM patients. The CTC-chip showed superior CTC-detection performance over CellSearch in exptl. models (sensitivity, 63.3%-64.5% vs 0%-1.1%; P < .001) and in clin. samples (CTC-positivity, 68.8% vs 6.3%; P < .001). A receiver operating characteristic (ROC) anal. showed that the CTC test provided a significant diagnostic performance in discrimination of unresectable disease from resectable disease (area under the ROC curve, 0.851; P = .003). The higher CTC count (=2 cells/mL) was significantly assocd. with a poor prognosis (P = .030). The novel CTC-chip enabled sensitive detection of CTCs, which provided significant diagnostic and prognostic information in MPM.
- 17Byun, S.; Son, S.; Amodei, D.; Cermak, N.; Shaw, J.; Kang, J. H.; Hecht, V. C.; Winslow, M. M.; Jacks, T.; Mallick, P.; Manalis, S. R. Characterizing deformability and surface friction of cancer cells. Proc. Natl. Acad. Sci. U. S. A. 2013, 110 (19), 7580– 5, DOI: 10.1073/pnas.1218806110Google Scholar17https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXptFGrtrc%253D&md5=518975ec5375f00df12e70e6567b64f4Characterizing deformability and surface friction of cancer cellsByun, Sangwon; Son, Sungmin; Amodei, Dario; Cermak, Nathan; Shaw, Josephine; Kang, Joon Ho; Hecht, Vivian C.; Winslow, Monte M.; Jacks, Tyler; Mallick, Parag; Manalis, Scott R.Proceedings of the National Academy of Sciences of the United States of America (2013), 110 (19), 7580-7585, S7580/1-S7580/10CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)Metastasis requires the penetration of cancer cells through tight spaces, which is mediated by the phys. properties of the cells as well as their interactions with the confined environment. Various microfluidic approaches have been devised to mimic traversal in vitro by measuring the time required for cells to pass through a constriction. Although a cell's passage time is expected to depend on its deformability, measurements from existing approaches are confounded by a cell's size and its frictional properties with the channel wall. Here, we introduce a device that enables the precise measurement of (i) the size of a single cell, given by its buoyant mass, (ii) the velocity of the cell entering a constricted microchannel (entry velocity), and (iii) the velocity of the cell as it transits through the constriction (transit velocity). Changing the deformability of the cell by perturbing its cytoskeleton primarily alters the entry velocity, whereas changing the surface friction by immobilizing pos. charges on the constriction's walls primarily alters the transit velocity, indicating that these parameters can give insight into the factors affecting the passage of each cell. When accounting for cell buoyant mass, we find that cells possessing higher metastatic potential exhibit faster entry velocities than cells with lower metastatic potential. We addnl. find that some cell types with higher metastatic potential exhibit greater than expected changes in transit velocities, suggesting that not only the increased deformability but reduced friction may be a factor in enabling invasive cancer cells to efficiently squeeze through tight spaces.
- 18Shaw Bagnall, J.; Byun, S.; Begum, S.; Miyamoto, D. T.; Hecht, V. C.; Maheswaran, S.; Stott, S. L.; Toner, M.; Hynes, R. O.; Manalis, S. R. Deformability of Tumor Cells versus Blood Cells. Sci. Rep. 2015, 5, 18542, DOI: 10.1038/srep18542Google Scholar18https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXitVKrsLbF&md5=36365ef18dad7735c0b6177cc56d5a8fDeformability of Tumor Cells versus Blood CellsShaw Bagnall, Josephine; Byun, Sangwon; Begum, Shahinoor; Miyamoto, David T.; Hecht, Vivian C.; Maheswaran, Shyamala; Stott, Shannon L.; Toner, Mehmet; Hynes, Richard O.; Manalis, Scott R.Scientific Reports (2015), 5 (), 18542CODEN: SRCEC3; ISSN:2045-2322. (Nature Publishing Group)The potential for circulating tumor cells (CTCs) to elucidate the process of cancer metastasis and inform clin. decision-making has made their isolation of great importance. However, CTCs are rare in the blood, and universal properties with which to identify them remain elusive. As technol. advancements have made single-cell deformability measurements increasingly routine, the assessment of phys. distinctions between tumor cells and blood cells may provide insight into the feasibility of deformability-based methods for identifying CTCs in patient blood. To this end, we present an initial study assessing deformability differences between tumor cells and blood cells, indicated by the length of time required for them to pass through a microfluidic constriction. Here, we demonstrate that deformability changes in tumor cells that have undergone phenotypic shifts are small compared to differences between tumor cell lines and blood cells. Addnl., in a syngeneic mouse tumor model, cells that are able to exit a tumor and enter circulation are not required to be more deformable than the cells that were first injected into the mouse. However, a limited study of metastatic prostate cancer patients provides evidence that some CTCs may be more mech. similar to blood cells than to typical tumor cell lines.
- 19Sarioglu, A. F.; Aceto, N.; Kojic, N.; Donaldson, M. C.; Zeinali, M.; Hamza, B.; Engstrom, A.; Zhu, H.; Sundaresan, T. K.; Miyamoto, D. T.; Luo, X.; Bardia, A.; Wittner, B. S.; Ramaswamy, S.; Shioda, T.; Ting, D. T.; Stott, S. L.; Kapur, R.; Maheswaran, S.; Haber, D. A.; Toner, M. A microfluidic device for label-free, physical capture of circulating tumor cell clusters. Nat. Methods 2015, 12 (7), 685– 91, DOI: 10.1038/nmeth.3404Google Scholar19https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhtFWqtbbE&md5=6ce523882376f88875f506e33cb04b22A microfluidic device for label-free, physical capture of circulating tumor cell clustersSarioglu, A. Fatih; Aceto, Nicola; Kojic, Nikola; Donaldson, Maria C.; Zeinali, Mahnaz; Hamza, Bashar; Engstrom, Amanda; Zhu, Huili; Sundaresan, Tilak K.; Miyamoto, David T.; Luo, Xi; Bardia, Aditya; Wittner, Ben S.; Ramaswamy, Sridhar; Shioda, Toshi; Ting, David T.; Stott, Shannon L.; Kapur, Ravi; Maheswaran, Shyamala; Haber, Daniel A.; Toner, MehmetNature Methods (2015), 12 (7), 685-691CODEN: NMAEA3; ISSN:1548-7091. (Nature Publishing Group)Cancer cells metastasize through the bloodstream either as single migratory circulating tumor cells (CTCs) or as multicellular groupings (CTC clusters). Existing technologies for CTC enrichment are designed to isolate single CTCs, and although CTC clusters are detectable in some cases, their true prevalence and significance remain to be detd. Here we developed a microchip technol. (the Cluster-Chip) to capture CTC clusters independently of tumor-specific markers from unprocessed blood. CTC clusters are isolated through specialized bifurcating traps under low-shear stress conditions that preserve their integrity, and even two-cell clusters are captured efficiently. Using the Cluster-Chip, we identified CTC clusters in 30-40% of patients with metastatic breast or prostate cancer or with melanoma. RNA sequencing of CTC clusters confirmed their tumor origin and identified tissue-derived macrophages within the clusters. Efficient capture of CTC clusters will enable the detailed characterization of their biol. properties and role in metastasis.
- 20Poudineh, M.; Aldridge, P. M.; Ahmed, S.; Green, B. J.; Kermanshah, L.; Nguyen, V.; Tu, C.; Mohamadi, R. M.; Nam, R. K.; Hansen, A.; Sridhar, S. S.; Finelli, A.; Fleshner, N. E.; Joshua, A. M.; Sargent, E. H.; Kelley, S. O. Tracking the dynamics of circulating tumour cell phenotypes using nanoparticle-mediated magnetic ranking. Nat. Nanotechnol. 2017, 12 (3), 274– 281, DOI: 10.1038/nnano.2016.239Google Scholar20https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhvFahtLbL&md5=484c655c7f6d0f4ef882e913d52ca5d4Tracking the dynamics of circulating tumour cell phenotypes using nanoparticle-mediated magnetic rankingPoudineh, Mahla; Aldridge, Peter M.; Ahmed, Sharif; Green, Brenda J.; Kermanshah, Leyla; Nguyen, Vivian; Tu, Carmen; Mohamadi, Reza M.; Nam, Robert K.; Hansen, Aaron; Sridhar, Srikala S.; Finelli, Antonio; Fleshner, Neil E.; Joshua, Anthony M.; Sargent, Edward H.; Kelley, Shana O.Nature Nanotechnology (2017), 12 (3), 274-281CODEN: NNAABX; ISSN:1748-3387. (Nature Publishing Group)Profiling the heterogeneous phenotypes of rare circulating tumor cells (CTCs) in whole blood is crit. to unraveling the complex and dynamic properties of these potential clin. markers. This task is challenging because these cells are present at ppb levels among normal blood cells. Here the authors report a new nanoparticle-enabled method for CTC characterization, called magnetic ranking cytometry, which profiles CTCs on the basis of their surface expression phenotype. The authors achieve this using a microfluidic chip that successfully processes whole blood samples. The approach classifies CTCs with single-cell resoln. in accordance with their expression of phenotypic surface markers, which is read out using magnetic nanoparticles. The authors deploy this new technique to reveal the dynamic phenotypes of CTCs in unprocessed blood from mice as a function of tumor growth and aggressiveness. The authors also test magnetic ranking cytometry using blood samples collected from cancer patients.
- 21Labib, M.; Mohamadi, R. M.; Poudineh, M.; Ahmed, S. U.; Ivanov, I.; Huang, C. L.; Moosavi, M.; Sargent, E. H.; Kelley, S. O. Single-cell mRNA cytometry via sequence-specific nanoparticle clustering and trapping. Nat. Chem. 2018, 10 (5), 489– 495, DOI: 10.1038/s41557-018-0025-8Google Scholar21https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXos1Smtr8%253D&md5=dd6ba074a864f04d2ef145e4f8ef1131Single-cell mRNA cytometry via sequence-specific nanoparticle clustering and trappingLabib, Mahmoud; Mohamadi, Reza M.; Poudineh, Mahla; Ahmed, Sharif U.; Ivanov, Ivaylo; Huang, Ching-Lung; Moosavi, Maral; Sargent, Edward H.; Kelley, Shana O.Nature Chemistry (2018), 10 (5), 489-495CODEN: NCAHBB; ISSN:1755-4330. (Nature Research)Cell-to-cell variation in gene expression creates a need for techniques that can characterize expression at the level of individual cells. This is particularly true for rare circulating tumor cells, in which subtyping and drug resistance are of intense interest. Here the authors describe a method for cell anal.-single-cell mRNA cytometry-that enables the isolation of rare cells from whole blood as a function of target mRNA sequences. This approach uses two classes of magnetic particles that are labeled to selectively hybridize with different regions of the target mRNA. Hybridization leads to the formation of large magnetic clusters that remain localized within the cells of interest, thereby enabling the cells to be magnetically sepd. Targeting specific intracellular mRNAs enables circulating tumor cells to be distinguished from normal hematopoietic cells. No polymerase chain reaction amplification is required to det. RNA expression levels and genotype at the single-cell level, and minimal cell manipulation is required. To demonstrate this approach the authors use single-cell mRNA cytometry to detect clin. important sequences in prostate cancer specimens.
- 22Pantel, K.; Brakenhoff, R. H.; Brandt, B. Detection, clinical relevance and specific biological properties of disseminating tumour cells. Nat. Rev. Cancer 2008, 8 (5), 329– 40, DOI: 10.1038/nrc2375Google Scholar22https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1cXltVGlurs%253D&md5=a8b4dea66d28b6705e67933ed7e65a82Detection, clinical relevance and specific biological properties of disseminating tumour cellsPantel, Klaus; Brakenhoff, Ruud H.; Brandt, BurkhardNature Reviews Cancer (2008), 8 (5), 329-340CODEN: NRCAC4; ISSN:1474-175X. (Nature Publishing Group)A review. Most cancer deaths are caused by hematogenous metastatic spread and subsequent growth of tumor cells at distant organs. Disseminating tumor cells present in the peripheral blood and bone marrow can now be detected and characterized at the single-cell level. These cells are highly relevant to the study of the biol. of early metastatic spread and provide a diagnostic source in patients with overt metastases. Here we review the evidence that disseminating tumor cells have a variety of uses for understanding tumor biol. and improving cancer treatment.
- 23Ligthart, S. T.; Coumans, F. A. W.; Attard, G.; Mulick Cassidy, A.; de Bono, J. S.; Terstappen, L. W. M. M. Unbiased and automated identification of a circulating tumour cell definition that associates with overall survival. PLoS One 2011, 6 (11), e27419, DOI: 10.1371/journal.pone.0027419Google Scholar23https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXhsFemtrzJ&md5=cc3f4373514ac79444d0179d5343dcd9Unbiased and automated identification of a circulating tumour cell definition that associates with overall survivalLigthart, Sjoerd T.; Coumans, Frank A. W.; Attard, Gerhardt; Cassidy, Amy Mulick; de Bono, Johann S.; Terstappen, Leon W. M. M.PLoS One (2011), 6 (11), e27419CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Circulating tumor cells (CTC) in patients with metastatic carcinomas are assocd. with poor survival and can be used to guide therapy. Classification of CTC however remains subjective, as they are morphol. heterogeneous. We acquired digital images, using the CellSearch system, from blood of 185 castration resistant prostate cancer (CRPC) patients and 68 healthy subjects to define CTC by computer algorithms. Patient survival data was used as the training parameter for the computer to define CTC. The computer-generated CTC definition was validated on a sep. CRPC dataset comprising 100 patients. The optimal definition of the computer defined CTC (aCTC) was stricter as compared to the manual CellSearch CTC (mCTC) definition and as a consequence aCTC were less frequent. The computer-generated CTC definition resulted in hazard ratios (HRs) of 2.8 for baseline and 3.9 for follow-up samples, which is comparable to the mCTC definition (baseline HR 2.9, follow-up HR 4.5). Validation resulted in HRs at baseline/follow-up of 3.9/5.4 for computer and 4.8/5.8 for manual definitions. In conclusion, we have defined and validated CTC by clin. outcome using a perfectly reproducing automated algorithm.
- 24Watson, P. A.; Arora, V. K.; Sawyers, C. L. Emerging mechanisms of resistance to androgen receptor inhibitors in prostate cancer. Nat. Rev. Cancer 2015, 15 (12), 701– 11, DOI: 10.1038/nrc4016Google Scholar24https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhvVCqtLnN&md5=98aaa704d576f2954cc1d83e474c0863Emerging mechanisms of resistance to androgen receptor inhibitors in prostate cancerWatson, Philip A.; Arora, Vivek K.; Sawyers, Charles L.Nature Reviews Cancer (2015), 15 (12), 701-711CODEN: NRCAC4; ISSN:1474-175X. (Nature Publishing Group)A review. During the past 10 years, preclin. studies implicating sustained androgen receptor (AR) signaling as the primary driver of castration-resistant prostate cancer (CRPC) have led to the development of novel agents targeting the AR pathway that are now in widespread clin. use. These drugs prolong the survival of patients with late-stage prostate cancer but are not curative. In this Review, we highlight emerging mechanisms of acquired resistance to these contemporary therapies, which fall into the three broad categories of restored AR signaling, AR bypass signaling and complete AR independence. This diverse range of resistance mechanisms presents new challenges for long-term disease control, which may be addressable through early use of combination therapies guided by recent insights from genomic landscape studies of CRPC.
- 25Lowes, L. E.; Goodale, D.; Xia, Y.; Postenka, C.; Piaseczny, M. M.; Paczkowski, F.; Allan, A. L. Epithelial-to-mesenchymal transition leads to disease-stage differences in circulating tumor cell detection and metastasis in pre-clinical models of prostate cancer. Oncotarget 2016, 7 (46), 76125– 76139, DOI: 10.18632/oncotarget.12682Google Scholar25https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC2srksFWlsg%253D%253D&md5=f6037b90eac2d99d28bbc5ea4ce47493Epithelial-to-mesenchymal transition leads to disease-stage differences in circulating tumor cell detection and metastasis in pre-clinical models of prostate cancerLowes Lori E; Piaseczny Matthew M; Allan Alison L; Goodale David; Xia Ying; Postenka Carl; Paczkowski Freeman; Allan Alison L; Allan Alison L; Allan Alison LOncotarget (2016), 7 (46), 76125-76139 ISSN:.Metastasis is the cause of most prostate cancer (PCa) deaths and has been associated with circulating tumor cells (CTCs). The presence of ≥5 CTCs/7.5mL of blood is a poor prognosis indicator in metastatic PCa when assessed by the CellSearch® system, the "gold standard" clinical platform. However, ~35% of metastatic PCa patients assessed by CellSearch® have undetectable CTCs. We hypothesize that this is due to epithelial-to-mesenchymal transition (EMT) and subsequent loss of necessary CTC detection markers, with important implications for PCa metastasis. Two pre-clinical assays were developed to assess human CTCs in xenograft models; one comparable to CellSearch® (EpCAM-based) and one detecting CTCs semi-independent of EMT status via combined staining with EpCAM/HLA (human leukocyte antigen). In vivo differences in CTC generation, kinetics, metastasis and EMT status were determined using 4 PCa models with progressive epithelial (LNCaP, LNCaP-C42B) to mesenchymal (PC-3, PC-3M) phenotypes. Assay validation demonstrated that the CellSearch®-based assay failed to detect a significant number (~40-50%) of mesenchymal CTCs. In vivo, PCa with an increasingly mesenchymal phenotype shed greater numbers of CTCs more quickly and with greater metastatic capacity than PCa with an epithelial phenotype. Notably, the CellSearch®-based assay captured the majority of CTCs shed during early-stage disease in vivo, and only after establishment of metastases were a significant number of undetectable CTCs present. This study provides important insight into the influence of EMT on CTC generation and subsequent metastasis, and highlights that novel technologies aimed at capturing mesenchymal CTCs may only be useful in the setting of advanced metastatic disease.
- 26Antonarakis, E. S.; Lu, C.; Luber, B.; Wang, H.; Chen, Y.; Zhu, Y.; Silberstein, J. L.; Taylor, M. N.; Maughan, B. L.; Denmeade, S. R.; Pienta, K. J.; Paller, C. J.; Carducci, M. A.; Eisenberger, M. A.; Luo, J. Clinical Significance of Androgen Receptor Splice Variant-7 mRNA Detection in Circulating Tumor Cells of Men With Metastatic Castration-Resistant Prostate Cancer Treated With First- and Second-Line Abiraterone and Enzalutamide. J. Clin. Oncol. 2017, 35 (19), 2149– 2156, DOI: 10.1200/JCO.2016.70.1961Google Scholar26https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXhvFKnsL%252FK&md5=471447974648fa52958ea71a7959637eClinical Significance of androgen receptor splice variant-7 mRNA detection in circulating tumor cells of men with metastatic castration-resistant prostate cancer treated With first- and second-line abiraterone and enzalutamideAntonarakis, Emmanuel S.; Lu, Changxue; Luber, Brandon; Wang, Hao; Chen, Yan; Zhu, Yezi; Silberstein, John L.; Taylor, Maritza N.; Maughan, Benjamin L.; Denmeade, Samuel R.; Pienta, Kenneth J.; Paller, Channing J.; Carducci, Michael A.; Eisenberger, Mario A.; Luo, JunJournal of Clinical Oncology (2017), 35 (19), 2149-2157CODEN: JCONDN; ISSN:1527-7755. (American Society of Clinical Oncology)Purpose We reported previously that the detection of androgen receptor splice variant-7 (AR-V7) mRNA in circulating tumor cells (CTCs) correlated with poor outcomes from the use of abiraterone and enzalutamide in patients with castration-resistant prostate cancer (CRPC). Here, we expanded our cohort size to better characterize the prognostic significance of AR-V7 in this setting. Methods We prospectively enrolled 202 patients with CRPC starting abiraterone or enzalutamide and investigated the prognostic value of CTC detection (+ v -) and AR-V7 detection (+ v -) using a CTCbased AR-V7 mRNA assay. We examd. $ 50% prostate-specific antigen (PSA) responses, PSA progression-free survival, clin. and radiol. progression-free survival, and overall survival. We constructed multivariable models adjusting for PSA, Gleason sum, no. of prior hormone therapies, prior abiraterone or enzalutamide use, prior taxane use, presence of visceral metastases, and Eastern Cooperative Oncol. Group score. We also sep. examd. the first-line and second-line novel hormonal therapy (NHT) settings. Results Median follow-up times were 15.0, 21.7, and 14.6months for CTC-, CTC+/AR-V7- and CTC+/AR-V7+ patients, resp. CTC+/AR-V7+ patients were more likely to have Gleason scores ≥8 (P =.05), metastatic disease at diagnosis (P =.01), higher PSA (P <.01), prior abiraterone or enzalutamide use (P =.03), prior taxane use (P =.02), and Eastern Cooperative Oncol. Group ≥1 (P =.01). Outcomes for the overall cohort (and sep. for the first-line and second-line NHT cohorts) were best for CTC- patients, intermediate for CTC+/AR-V7- patients, andworse for CTC+/AR-V7+ patients. These correlations remained significant in multivariable models. Conclusion This expanded anal. further characterizes the importance of CTC-based AR-V7 mRNA detection in predicting outcomes in patients with CRPC receiving first- and second-line NHT and, to the best of our knowledge, is the first to suggest that this assay be interpreted using three sep. prognostic categories: CTC-, CTC+/AR-V7-, and CTC+/AR-V7+.
- 27Satelli, A.; Batth, I.; Brownlee, Z.; Mitra, A.; Zhou, S.; Noh, H.; Rojas, C. R.; Li, H.; Meng, Q. H.; Li, S. EMT circulating tumor cells detected by cell-surface vimentin are associated with prostate cancer progression. Oncotarget 2017, 8 (30), 49329– 49337, DOI: 10.18632/oncotarget.17632Google Scholar27https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1crovFOktQ%253D%253D&md5=c167a5a77af07996b0f8b7f76ce658ecEMT circulating tumor cells detected by cell-surface vimentin are associated with prostate cancer progressionSatelli Arun; Batth Izhar; Brownlee Zachary; Mitra Abhishek; Noh Hyangsoon; Rojas Christina R; Li Heming; Li Shulin; Zhou Shouhao; Meng Qing HOncotarget (2017), 8 (30), 49329-49337 ISSN:.Recent advances in the field of circulating tumor cells (CTC) have shown promise in this liquid biopsy-based prognosis of patient outcome. However, not all of the circulating cells are tumor cells, as evidenced by a lack of tumor-specific markers. The current FDA standard for capturing CTCs (CellSearch) relies on an epithelial marker and cells captured via CellSearch cannot be considered to have undergone EMT. Therefore, it is difficult to ascertain the presence and relevance of any mesenchymal or EMT-like CTCs. To address this gap in technology, we recently discovered the utility of cell-surface vimentin (CSV) as a marker for detecting mesenchymal CTCs from sarcoma, breast, and colon cancer. Here we studied peripheral blood samples of 48 prostate cancer (PCA) patients including hormone sensitive and castration resistant sub-groups. Blood samples were analyzed for three different properties including our own CSV-based CTC enumeration (using 84-1 mAb against CSV), CellSearch-based epithelial CTC counts, and serum prostate-specific antigen (PSA) quantification. Our data demonstrated that in comparison with CellSearch, the CSV-based method had greater sensitivity and specificity. Further, we observed significantly greater numbers of CTCs in castration resistant patients as measured by our CSV method but not CellSearch. Our data suggests CSV-guided CTC enumeration may hold prognostic value and should be further validated as a possible measurement of PCA progression towards the deadly, androgen-independent form.
- 28Armstrong, A. J.; Marengo, M. S.; Oltean, S.; Kemeny, G.; Bitting, R. L.; Turnbull, J. D.; Herold, C. I.; Marcom, P. K.; George, D. J.; Garcia-Blanco, M. A. Circulating tumor cells from patients with advanced prostate and breast cancer display both epithelial and mesenchymal markers. Mol. Cancer Res. 2011, 9 (8), 997– 1007, DOI: 10.1158/1541-7786.MCR-10-0490Google Scholar28https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXhtVajtbvL&md5=78bab32e33ba992ed65beb9acbfb0acdCirculating Tumor Cells from Patients with Advanced Prostate and Breast Cancer Display Both Epithelial and Mesenchymal MarkersArmstrong, Andrew J.; Marengo, Matthew S.; Oltean, Sebastian; Kemeny, Gabor; Bitting, Rhonda L.; Turnbull, James D.; Herold, Christina I.; Marcom, Paul K.; George, Daniel J.; Garcia-Blanco, Mariano A.Molecular Cancer Research (2011), 9 (8), 997-1007CODEN: MCROC5; ISSN:1541-7786. (American Association for Cancer Research)During cancer progression, malignant cells undergo epithelial-mesenchymal transitions (EMT) and mesenchymal-epithelial transitions (MET) as part of a broad invasion and metastasis program. We previously obsd. MET events among lung metastases in a preclin. model of prostate adenocarcinoma that suggested a relationship between epithelial plasticity and metastatic spread. We thus sought to translate these findings into clin. evidence by examg. the existence of EMT in circulating tumor cells (CTC) from patients with progressive metastatic solid tumors, with a focus on men with castration-resistant prostate cancer (CRPC) and women with metastatic breast cancer. We showed that the majority (>80%) of these CTCs in patients with metastatic CRPC coexpress epithelial proteins such as epithelial cell adhesion mol. (EpCAM), cytokeratins (CK), and E-cadherin, with mesenchymal proteins including vimentin, N-cadherin and O-cadherin, and the stem cell marker CD133. Equally, we found that more than 75% of CTCs from women with metastatic breast cancer coexpress CK, vimentin, and N-cadherin. The existence and high frequency of these CTCs coexpressing epithelial, mesenchymal, and stem cell markers in patients with progressive metastases has important implications for the application and interpretation of approved methods to detect CTCs.
- 29Chen, C. L.; Mahalingam, D.; Osmulski, P.; Jadhav, R. R.; Wang, C. M.; Leach, R. J.; Chang, T. C.; Weitman, S. D.; Kumar, A. P.; Sun, L.; Gaczynska, M. E.; Thompson, I. M.; Huang, T. H. Single-cell analysis of circulating tumor cells identifies cumulative expression patterns of EMT-related genes in metastatic prostate cancer. Prostate 2013, 73 (8), 813– 26, DOI: 10.1002/pros.22625Google Scholar29https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXmslWnu74%253D&md5=13a5248089ac2b8796a6893728b27512Single-cell analysis of circulating tumor cells identifies cumulative expression patterns of EMT-related genes in metastatic prostate cancerChen, Chun-Liang; Mahalingam, Devalingam; Osmulski, Pawel; Jadhav, Rohit R.; Wang, Chiou-Miin; Leach, Robin J.; Chang, Tien-Cheng; Weitman, Steven D.; Kumar, Addanki Pratap; Sun, Lu Zhe; Gaczynska, Maria E.; Thompson, Ian M.; Huang, Tim Hui-MingProstate (Hoboken, NJ, United States) (2013), 73 (8), 813-826CODEN: PRSTDS; ISSN:0270-4137. (Wiley-Blackwell)BACKGROUND : Prostate tumors shed circulating tumor cells (CTCs) into the blood stream. Increased evidence shows that CTCs are often present in metastatic prostate cancer and can be alternative sources for disease profiling and prognostication. Here we postulate that CTCs expressing genes related to epithelial-mesenchymal transition (EMT) are strong predictors of metastatic prostate cancer. METHODS : A microfiltration system was used to trap CTCs from peripheral blood based on size selection of large epithelial-like cells without CD45 leukocyte marker. These cells individually retrieved with a micromanipulator device were assessed for cell membrane phys. properties using at. force microscopy. Addnl., 38 CTCs from eight prostate cancer patients were used to det. expression profiles of 84 EMT-related and ref. genes using a microfluidics-based PCR system. RESULTS : Increased cell elasticity and membrane smoothness were found in CTCs compared to noncancerous cells, highlighting their potential invasiveness and mobility in the peripheral circulation. Despite heterogeneous expression patterns of individual CTCs, genes that promote mesenchymal transitioning into a more malignant state, including IGF1, IGF2, EGFR, FOXP3, and TGFB3, were commonly obsd. in these cells. An addnl. subset of EMT-related genes (e.g., PTPRN2, ALDH1, ESR2, and WNT5A) were expressed in CTCs of castration-resistant cancer, but less frequently in castration-sensitive cancer. CONCLUSIONS : The study suggests that an incremental expression of EMT-related genes in CTCs is assocd. with metastatic castration-resistant cancer. Although CTCs represent a group of highly heterogeneous cells, their unique EMT-related gene signatures provide a new opportunity for personalized treatments with targeted inhibitors in advanced prostate cancer patients. Prostate 73: 813-826, 2013. © 2012 Wiley Periodicals, Inc.
- 30Mitra, A.; Mishra, L.; Li, S. EMT, CTCs and CSCs in tumor relapse and drug-resistance. Oncotarget 2015, 6 (13), 10697– 10711, DOI: 10.18632/oncotarget.4037Google Scholar30https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC2MfltVyktw%253D%253D&md5=db1d641e9116b4527b6fb733c2a40a81EMT, CTCs and CSCs in tumor relapse and drug-resistanceMitra Abhisek; Li Shulin; Mishra LopaOncotarget (2015), 6 (13), 10697-711 ISSN:.Tumor relapse and metastasis are the primary causes of poor survival rates in patients with advanced cancer despite successful resection or chemotherapeutic treatment. A primary cause of relapse and metastasis is the persistence of cancer stem cells (CSCs), which are highly resistant to chemotherapy. Although highly efficacious drugs suppressing several subpopulations of CSCs in various tissue-specific cancers are available, recurrence is still common in patients. To find more suitable therapy for relapse, the mechanisms underlying metastasis and drug-resistance associated with relapse-initiating CSCs need to be identified. Recent studies in circulating tumor cells (CTCs) of some cancer patients manifest phenotypes of both CSCs and epithelial-mesenchymal transition (EMT). These patients are unresponsive to standard chemotherapies and have low progression free survival, suggesting that EMT-positive CTCs are related to co-occur with or transform into relapse-initiating CSCs. Furthermore, EMT programming in cancer cells enables in the remodeling of extracellular matrix to break the dormancy of relapse-initiating CSCs. In this review, we extensively discuss the association of the EMT program with CTCs and CSCs to characterize a subpopulation of patients prone to relapses. Identifying the mechanisms by which EMT-transformed CTCs and CSCs initiate relapse could facilitate the development of new or enhanced personalized therapeutic regimens.
- 31Chang, L.; Graham, P. H.; Hao, J.; Ni, J.; Bucci, J.; Cozzi, P. J.; Kearsley, J. H.; Li, Y. Acquisition of epithelial-mesenchymal transition and cancer stem cell phenotypes is associated with activation of the PI3K/Akt/mTOR pathway in prostate cancer radioresistance. Cell Death Dis. 2013, 4, e875, DOI: 10.1038/cddis.2013.407Google Scholar31https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhs1Olt7jK&md5=0f7e376b9cf9e3e5cd96544c405371e0Acquisition of epithelial-mesenchymal transition and cancer stem cell phenotypes is associated with activation of the PI3K/Akt/mTOR pathway in prostate cancer radioresistanceChang, L.; Graham, P. H.; Hao, J.; Ni, J.; Bucci, J.; Cozzi, P. J.; Kearsley, J. H.; Li, Y.Cell Death & Disease (2013), 4 (Oct.), e875CODEN: CDDEA4; ISSN:2041-4889. (Nature Publishing Group)Radioresistance is a major challenge in prostate cancer (CaP) radiotherapy (RT). In this study, we investigated the role and assocn. of epithelial-mesenchymal transition (EMT), cancer stem cells (CSCs) and the PI3K/Akt/mTOR signaling pathway in CaP radioresistance. We developed three novel CaP radioresistant (RR) cell lines (PC-3RR, DU145RR and LNCaPRR) by radiation treatment and confirmed their radioresistance using a clonogenic survival assay. Compared with untreated CaP-control cells, the CaP-RR cells had increased colony formation, invasion ability and spheroid formation capability (P<0.05). In addn., enhanced EMT/CSC phenotypes and activation of the checkpoint proteins (Chk1 and Chk2) and the PI3K/Akt/mTOR signaling pathway proteins were also found in CaP-RR cells using immunofluorescence, western blotting and quant. real-time PCR (qRT-PCR). Furthermore, combination of a dual PI3K/mTOR inhibitor (BEZ235) with RT effectively increased radiosensitivity and induced more apoptosis in CaP-RR cells, concomitantly correlated with the reduced expression of EMT/CSC markers and the PI3K/Akt/mTOR signaling pathway proteins compared with RT alone. Our findings indicate that CaP radioresistance is assocd. with EMT and enhanced CSC phenotypes via activation of the PI3K/Akt/mTOR signaling pathway, and that the combination of BEZ235 with RT is a promising modality to overcome radioresistance in the treatment of CaP. This combination approach warrants future in vivo animal study and clin. trials.
- 32Scher, H. I.; Morris, M. J.; Stadler, W. M.; Higano, C.; Basch, E.; Fizazi, K.; Antonarakis, E. S.; Beer, T. M.; Carducci, M. A.; Chi, K. N.; Corn, P. G.; de Bono, J. S.; Dreicer, R.; George, D. J.; Heath, E. I.; Hussain, M.; Kelly, W. K.; Liu, G.; Logothetis, C.; Nanus, D.; Stein, M. N.; Rathkopf, D. E.; Slovin, S. F.; Ryan, C. J.; Sartor, O.; Small, E. J.; Smith, M. R.; Sternberg, C. N.; Taplin, M. E.; Wilding, G.; Nelson, P. S.; Schwartz, L. H.; Halabi, S.; Kantoff, P. W.; Armstrong, A. J. Trial Design and Objectives for Castration-Resistant Prostate Cancer: Updated Recommendations From the Prostate Cancer Clinical Trials Working Group 3. J. Clin. Oncol. 2016, 34 (12), 1402– 18, DOI: 10.1200/JCO.2015.64.2702Google Scholar32https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC28jjt1KgsA%253D%253D&md5=ca0a8f257cabae86b5434b97079b2552Trial Design and Objectives for Castration-Resistant Prostate Cancer: Updated Recommendations From the Prostate Cancer Clinical Trials Working Group 3Scher Howard I; Morris Michael J; Stadler Walter M; Higano Celestia; Basch Ethan; Fizazi Karim; Antonarakis Emmanuel S; Beer Tomasz M; Carducci Michael A; Chi Kim N; Corn Paul G; de Bono Johann S; Dreicer Robert; George Daniel J; Heath Elisabeth I; Hussain Maha; Kelly Wm Kevin; Liu Glenn; Logothetis Christopher; Nanus David; Stein Mark N; Rathkopf Dana E; Slovin Susan F; Ryan Charles J; Sartor Oliver; Small Eric J; Smith Matthew Raymond; Sternberg Cora N; Taplin Mary-Ellen; Wilding George; Nelson Peter S; Schwartz Lawrence H; Halabi Susan; Kantoff Philip W; Armstrong Andrew JJournal of clinical oncology : official journal of the American Society of Clinical Oncology (2016), 34 (12), 1402-18 ISSN:.PURPOSE: Evolving treatments, disease phenotypes, and biology, together with a changing drug development environment, have created the need to revise castration-resistant prostate cancer (CRPC) clinical trial recommendations to succeed those from prior Prostate Cancer Clinical Trials Working Groups. METHODS: An international expert committee of prostate cancer clinical investigators (the Prostate Cancer Clinical Trials Working Group 3 [PCWG3]) was reconvened and expanded and met in 2012-2015 to formulate updated criteria on the basis of emerging trial data and validation studies of the Prostate Cancer Clinical Trials Working Group 2 recommendations. RESULTS: PCWG3 recommends that baseline patient assessment include tumor histology, detailed records of prior systemic treatments and responses, and a detailed reporting of disease subtypes based on an anatomic pattern of metastatic spread. New recommendations for trial outcome measures include the time to event end point of symptomatic skeletal events, as well as time to first metastasis and time to progression for trials in the nonmetastatic CRPC state. PCWG3 introduces the concept of no longer clinically benefiting to underscore the distinction between first evidence of progression and the clinical need to terminate or change treatment, and the importance of documenting progression in existing lesions as distinct from the development of new lesions. Serial biologic profiling using tumor samples from biopsies, blood-based diagnostics, and/or imaging is also recommended to gain insight into mechanisms of resistance and to identify predictive biomarkers of sensitivity for use in prospective trials. CONCLUSION: PCWG3 moves drug development closer to unmet needs in clinical practice by focusing on disease manifestations most likely to affect prognosis adversely for therapeutics tested in both nonmetastatic and metastatic CRPC populations. Consultation with regulatory authorities is recommended if a trial is intended to seek support for drug approval.
- 33Tucci, M.; Scagliotti, G. V.; Vignani, F. Metastatic castration-resistant prostate cancer: time for innovation. Future Oncol. 2015, 11 (1), 91– 106, DOI: 10.2217/fon.14.145Google Scholar33https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXkvFGhsA%253D%253D&md5=f891f272b109eb0d6a7523d8376ce2b7Metastatic castration-resistant prostate cancer: time for innovationTucci, Marcello; Scagliotti, Giorgio Vittorio; Vignani, FrancescaFuture Oncology (2015), 11 (1), 91-106CODEN: FOUNBN; ISSN:1479-6694. (Future Medicine Ltd.)A review. Androgen deprivation is the mainstay of advanced prostate cancer treatment. Despite initial responses, almost all patients progress to castration-resistant prostate cancer (CRPC). The understanding of the biol. of CRPC and the evidence that CRPC still remains driven by androgen receptor signaling led to the discovery of new therapeutic targets. In the last few years, large Phase III trials showed improvements in survival and outcomes and led to the approval of a CYP17 inhibitor (abiraterone), an androgen receptor antagonist (enzalutamide), the taxane cabazitaxel, an α-emitter (radium-223), the bone resorption-targeting drug denosumab and an immunotherapy (sipuleucel-T). This article describes the mol. mechanisms underlying castration resistance, discusses recent and ongoing trials and offers some insights into identifying the best sequence of new drugs.
- 34Liu, W.; Yin, B.; Wang, X.; Yu, P.; Duan, X.; Liu, C.; Wang, B.; Tao, Z. Circulating tumor cells in prostate cancer: Precision diagnosis and therapy. Oncol. Lett. 2017, 14 (2), 1223– 1232, DOI: 10.3892/ol.2017.6332Google Scholar34https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1cfls12qtg%253D%253D&md5=535eace292cffa2d2738134434a51cf2Circulating tumor cells in prostate cancer: Precision diagnosis and therapyLiu Weiwei; Yin Binbin; Wang Xuchu; Yu Pan; Duan Xiuzhi; Liu Chunhua; Wang Ben; Tao ZhihuaOncology letters (2017), 14 (2), 1223-1232 ISSN:1792-1074.The primary cause of tumor-associated mortality in prostate cancer (PCa) remains distant metastasis. The dissemination of tumor cells from the primary tumor to distant sites through the bloodstream cannot be detected early by standard imaging methods. Circulating tumor cells (CTCs) represent an effective prognostic and predictive biomarker, which are able to monitor efficacy of adjuvant therapies, detect early development of metastases, and finally, assess therapeutic responses of advanced disease earlier than traditional diagnostic methods. In addition, since repeated tissue biopsies are invasive, costly and not always feasible, the assessment of tumor characteristics on CTCs, by a peripheral blood sample as a liquid biopsy, represents an attractive opportunity. The implementation of molecular and genomic characterization of CTCs may contribute to improve the treatment selection and thus, to move toward more precise diagnosis and therapy in PCa. The present study summarizes the current advances in CTC enrichment and detection strategies and reviews how CTCs may contribute to significant insights in the metastatic process, as well as how they may be utilized in clinical application in PCa. Although it is proposed that CTCs may offer insights into the prognosis and management of PCa, there are a number of challenges in the study of circulating tumor cells, and their clinical utility remains under investigation.
- 35De Laere, B.; Oeyen, S.; Van Oyen, P.; Ghysel, C.; Ampe, J.; Ost, P.; Demey, W.; Hoekx, L.; Schrijvers, D.; Brouwers, B.; Lybaert, W.; Everaert, E.; Van Kerckhove, P.; De Maeseneer, D.; Strijbos, M.; Bols, A.; Fransis, K.; Beije, N.; de Kruijff, I.; van Dam, V.; Brouwer, A.; van Dam, P. J.; Van den Eynden, G.; Rutten, A.; Sleijfer, S.; Vandebroek, J.; Van Laere, S.; Dirix, L. Circulating tumor cells and survival in abiraterone- and enzalutamide-treated patients with castration-resistant prostate cancer. Prostate 2018, 78 (6), 435– 445, DOI: 10.1002/pros.23488Google Scholar35https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXks1ylsb0%253D&md5=bb35fa656326b4fec6aaabc65bc3f222Circulating tumor cells and survival in abiraterone- and enzalutamide-treated patients with castration-resistant prostate cancerDe Laere, Bram; Oeyen, Steffi; Van Oyen, Peter; Ghysel, Christophe; Ampe, Jozef; Ost, Piet; Demey, Wim; Hoekx, Lucien; Schrijvers, Dirk; Brouwers, Barbara; Lybaert, Willem; Everaert, Els; Van Kerckhove, Piet; De Maeseneer, Daan; Strijbos, Michiel; Bols, Alain; Fransis, Karen; Beije, Nick; de Kruijff, Inge; van Dam, Valerie; Brouwer, Anja; van Dam, Pieter-Jan; Van den Eynden, Gert; Rutten, Annemie; Sleijfer, Stefan; Vandebroek, Jean; Van Laere, Steven; Dirix, LucProstate (Hoboken, NJ, United States) (2018), 78 (6), 435-445CODEN: PRSTDS; ISSN:0270-4137. (Wiley-Blackwell)Background : The outcome to treatment administered to patients with metastatic castration-resistant prostate cancer (mCRPC) greatly differs between individuals, underlining the need for biomarkers guiding treatment decision making. Objective : To investigate the prognostic value of circulating tumor cell (CTC) enumeration and dynamics, in the context of second-line endocrine therapies (ie, abiraterone acetate or enzalutamide), irresp. of prior systemic therapies. Design, Settings, and Participants : In a prospective, multicentre study blood samples for CTC enumeration were collected from patients with mCRPC at baseline (n = 174). In patients who responded for minimally 10-12 wk a follow-up sample was collected. Outcome Measurements and Statistical Anal. : For baseline anal., patients were stratified in <5 or ≥5 CTCs/7.5 mL, whereas for the anal. of CTC dynamics at 10-12 wk, in patients with stable, increasing or decreasing CTC counts. Progression-free survival (PFS), overall survival (OS), and PSA changes at 10-12 wk were compared between groups. Results : Patients demonstrating increasing CTCs on therapy had a shorter median PFS (4.03 vs 12.98 vs 13.67 mo, HR 3.6, 95%CI 1.9-6.8; P < 0.0001) and OS (11.2 mo vs not reached, HR 9.5, 95%CI 3.7-24; P < 0.0001), compared to patients with decreasing or stable CTCs. Multivariable Cox regression showed that prior chemotherapy (HR 4.1, 95%CI 1.9-8.9; P = 0.0003), a high baseline CTC count (HR 1.5, 95%CI 1.2-1.9; P = 0.002) and increasing CTCs at follow-up (HR 3.3, 95%CI 1.4-7.6; P = 0.005) were independent predictors of worse PFS. Previous chemotherapy (HR 7, 95%CI 1.9-25; P = 0.003), high baseline CTC counts (HR 2.2, 95%CI 1.4-3.7; P = 0.002) and increasing CTCs during therapy (HR 4.6, 95%CI 1.4-15; P = 0.01) were independently assocd. with shorter OS. ≥30% and ≥50% PSA responses less frequently occurred in patients with CTC inclines at 10-12 wk on therapy (χ2 test: P < 0.01). Conclusions : CTC dynamics during therapy are assocd. with PSA response and provide independent clin. prognostication over PSA declines. Hence the study demonstrates the pharmacodynamic properties of CTCs.
- 36Scher, H. I.; Fizazi, K.; Saad, F.; Taplin, M. E.; Sternberg, C. N.; Miller, K.; de Wit, R.; Mulders, P.; Chi, K. N.; Shore, N. D.; Armstrong, A. J.; Flaig, T. W.; Flechon, A.; Mainwaring, P.; Fleming, M.; Hainsworth, J. D.; Hirmand, M.; Selby, B.; Seely, L.; de Bono, J. S. Increased survival with enzalutamide in prostate cancer after chemotherapy. N. Engl. J. Med. 2012, 367 (13), 1187– 97, DOI: 10.1056/NEJMoa1207506Google Scholar36https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhsVKhsLvK&md5=02fcfdf5b35766d91a2ca023558acd74Increased survival with enzalutamide in prostate cancer after chemotherapyScher, Howard I.; Fizazi, Karim; Saad, Fred; Taplin, Mary-Ellen; Sternberg, Cora N.; Miller, Kurt; de Wit, Ronald; Mulders, Peter; Chi, Kim N.; Shore, Neal D.; Armstrong, Andrew J.; Flaig, Thomas W.; Flechon, Aude; Mainwaring, Paul; Fleming, Mark; Hainsworth, John D.; Hirmand, Mohammad; Selby, Bryan; Seely, Lynn; de Bono, Johann S.New England Journal of Medicine (2012), 367 (13), 1187-1197CODEN: NEJMAG; ISSN:0028-4793. (Massachusetts Medical Society)Background: Enzalutamide (formerly called MDV3100) targets multiple steps in the androgen-receptor-signaling pathway, the major driver of prostate-cancer growth. We aimed to evaluate whether enzalutamide prolongs survival in men with castration-resistant prostate cancer after chemotherapy. Methods: In our phase 3, double-blind, placebo-controlled trial, we stratified 1199 men with castration-resistant prostate cancer after chemotherapy according to the Eastern Cooperative Oncol. Group performance-status score and pain intensity. We randomly assigned them, in a 2:1 ratio, to receive oral enzalutamide at a dose of 160 mg per day (800 patients) or placebo (399 patients). The primary end point was overall survival. Results: The study was stopped after a planned interim anal. at the time of 520 deaths. The median overall survival was 18.4 mo (95% confidence interval [CI], 17.3 to not yet reached) in the enzalutamide group vs. 13.6 mo (95% CI, 11.3 to 15.8) in the placebo group (hazard ratio for death in the enzalutamide group, 0.63; 95% CI, 0.53 to 0.75; P<0.001). The superiority of enzalutamide over placebo was shown with respect to all secondary end points: the proportion of patients with a redn. in the prostate-specific antigen (PSA) level by 50% or more (54% vs. 2%, P<0.001), the soft-tissue response rate (29% vs. 4%, P<0.001), the quality-of-life response rate (43% vs. 18%, P<0.001), the time to PSA progression (8.3 vs. 3.0 mo; hazard ratio, 0.25; P<0.001), radiog. progression-free survival (8.3 vs. 2.9 mo; hazard ratio, 0.40; P<0.001), and the time to the first skeletal-related event (16.7 vs. 13.3 mo; hazard ratio, 0.69; P<0.001). Rates of fatigue, diarrhea, and hot flashes were higher in the enzalutamide group. Seizures were reported in five patients (0.6%) receiving enzalutamide. Conclusions: Enzalutamide significantly prolonged the survival of men with metastatic castration-resistant prostate cancer after chemotherapy.
- 37Loriot, Y.; Bianchini, D.; Ileana, E.; Sandhu, S.; Patrikidou, A.; Pezaro, C.; Albiges, L.; Attard, G.; Fizazi, K.; De Bono, J. S.; Massard, C. Antitumour activity of abiraterone acetate against metastatic castration-resistant prostate cancer progressing after docetaxel and enzalutamide (MDV3100). Ann. Oncol. 2013, 24 (7), 1807– 12, DOI: 10.1093/annonc/mdt136Google Scholar37https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC3srktF2muw%253D%253D&md5=c9a3e24cb46d2f5d99e1b52b3eed39a7Antitumour activity of abiraterone acetate against metastatic castration-resistant prostate cancer progressing after docetaxel and enzalutamide (MDV3100)Loriot Y; Bianchini D; Ileana E; Sandhu S; Patrikidou A; Pezaro C; Albiges L; Attard G; Fizazi K; De Bono J S; Massard CAnnals of oncology : official journal of the European Society for Medical Oncology (2013), 24 (7), 1807-12 ISSN:.BACKGROUND: Androgen receptor (AR) signalling remains critically important in metastatic castration-resistant prostate cancer (mCRPC) as confirmed by recent phase III trials, showing a survival advantage for abiraterone acetate and enzalutamide (MDV3100). The antitumour activity of abiraterone and prednisolone in patients pre-treated with enzalutamide is as yet unknown. PATIENTS AND METHODS: We investigated the antitumour activity of abiraterone and prednisolone in patients with mCRPC who had progressed following treatment with docetaxel (Taxotere) and enzalutamide. Clinical data were retrospectively analysed for prostate-specific antigen (PSA) and RECIST responses, clinical benefit and survival. RESULTS: Thirty-eight patients were included in the analysis. The median age was 71 years (range 52-84); metastatic sites included bone disease in 37 patients (97%), lymph nodes in 15 patients (39%) and visceral disease in 10 patients (26%). Abiraterone was well tolerated. Three patients (8%) attained a PSA response, defined as ≥50% decline in PSA confirmed after ≥4 weeks, while seven patients (18%) had a ≥30% PSA decline. The median progression-free survival (PFS) was 2.7 months (95% CI 2.3-4.1). Of the 12 patients assessable radiologically, only 1 (8%) attained a confirmed partial response. CONCLUSION: Abiraterone and prednisolone have modest antitumour activities in patients with mCRPC pretreated with docetaxel and enzalutamide.
- 38Brand, L. J.; Dehm, S. M. Androgen receptor gene rearrangements: new perspectives on prostate cancer progression. Curr. Drug Targets 2013, 14 (4), 441– 449, DOI: 10.2174/1389450111314040005Google Scholar38https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXotlyrsLs%253D&md5=e5d8a285f32674aaf05282a17f799ec7Androgen receptor gene rearrangements: new perspectives on prostate cancer progressionBrand, Lucas J.; Dehm, Scott M.Current Drug Targets (2013), 14 (4), 441-449CODEN: CDTUAU; ISSN:1389-4501. (Bentham Science Publishers Ltd.)A review. The androgen receptor (AR) is a master regulator transcription factor in normal and cancerous prostate cells. Canonical AR activation requires binding of androgen ligand to the AR ligand binding domain, translocation to the nucleus, and transcriptional activation of AR target genes. This regulatory axis is targeted for systemic therapy of advanced prostate cancer. However, a new paradigm for AR activation in castration-resistant prostate cancer (CRPC) has emerged wherein alternative splicing of AR mRNA promotes synthesis of constitutively active AR variants that lack the AR ligand binding domain (LBD). Recent work has indicated that structural alteration of the AR gene locus represents a key mechanism by which alterations in AR mRNA splicing arise. In this review, we examine the role of truncated AR variants (ARVs) and their corresponding genomic origins in models of prostate cancer progression, as well as the challenges they pose to the current std. of prostate cancer therapies targeting the AR ligand binding domain. Since ARVs lack the COOH-terminal LBD, the genesis of these AR gene rearrangements and their resulting ARVs provides strong rationale for the pursuit of new avenues of therapeutic intervention targeted at the AR NH2-terminal domain. We further suggest that genomic events leading to ARV expression could act as novel biomarkers of disease progression that may guide the optimal use of current and next-generation AR-targeted therapy.
- 39van der Steen, T.; Tindall, D. J.; Huang, H. Posttranslational modification of the androgen receptor in prostate cancer. Int. J. Mol. Sci. 2013, 14 (7), 14833– 59, DOI: 10.3390/ijms140714833Google Scholar39https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXht1OntLzK&md5=f72323de1ac843834793fbf4d22c4a87Posttranslational modification of the androgen receptor in prostate cancervan der Steen, Travis; Tindall, Donald J.; Huang, HaojieInternational Journal of Molecular Sciences (2013), 14 (7), 14833-14859, 27 pp.CODEN: IJMCFK; ISSN:1422-0067. (MDPI AG)A review. The androgen receptor (AR) is important in the development of the prostate by regulating transcription, cellular proliferation, and apoptosis. AR undergoes posttranslational modifications that alter its transcription activity, translocation to the nucleus and stability. The posttranslational modifications that regulate these events are of utmost importance to understand the functional role of AR and its activity. The majority of these modifications occur in the activation function-1 (AF1) region of the AR, which contains the transcriptional activation unit 1 (TAU1) and 5 (TAU5). Identification of the modifications that occur to these regions may increase our understanding of AR activation in prostate cancer and the role of AR in the progression from androgen-dependent to castration-resistant prostate cancer (CRPC). Most of the posttranslational modifications identified to date have been detd. using the full-length AR in androgen dependent cells. Further investigations into the role of posttranslational modifications in androgen-independent activation of full-length AR and constitutively active splicing variants are warranted, findings from which may provide new therapeutic options for CRPC.
- 40Mohamadi, R. M.; Besant, J. D.; Mepham, A.; Green, B.; Mahmoudian, L.; Gibbs, T.; Ivanov, I.; Malvea, A.; Stojcic, J.; Allan, A. L.; Lowes, L. E.; Sargent, E. H.; Nam, R. K.; Kelley, S. O. Nanoparticle-mediated binning and profiling of heterogeneous circulating tumor cell subpopulations. Angew. Chem., Int. Ed. 2015, 54 (1), 139– 43, DOI: 10.1002/anie.201409376Google Scholar40https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhvVKlsLjI&md5=668e003fea2719f7bfbc02172f1a7150Nanoparticle-Mediated Binning and Profiling of Heterogeneous Circulating Tumor Cell SubpopulationsMohamadi, Reza M.; Besant, Justin D.; Mepham, Adam; Green, Brenda; Mahmoudian, Laili; Gibbs, Thaddeus; Ivanov, Ivaylo; Malvea, Anahita; Stojcic, Jessica; Allan, Alison L.; Lowes, Lori E.; Sargent, Edward H.; Nam, Robert K.; Kelley, Shana O.Angewandte Chemie, International Edition (2015), 54 (1), 139-143CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)The anal. of circulating tumor cells (CTCs) is an important capability that may lead to new approaches for cancer management. CTC capture devices developed to date isolate a bulk population of CTCs and do not differentiate subpopulations that may have varying phenotypes with different levels of clin. relevance. Here, the authors present a new device for CTC spatial sorting and profiling that sequesters blood-borne tumor cells with different phenotypes into discrete spatial bins. Validation data are presented showing that cancer cell lines with varying surface expression generate different binning profiles within the device. Working with patient blood samples, the authors obtain profiles that elucidate the heterogeneity of CTC populations present in cancer patients and also report on the status of CTCs within the epithelial-to-mesenchymal transition (EMT).
- 41Green, B. J.; Kermanshah, L.; Labib, M.; Ahmed, S. U.; Silva, P. N.; Mahmoudian, L.; Chang, I. H.; Mohamadi, R. M.; Rocheleau, J. V.; Kelley, S. O. Isolation of Phenotypically Distinct Cancer Cells Using Nanoparticle-Mediated Sorting. ACS Appl. Mater. Interfaces 2017, 9 (24), 20435– 20443, DOI: 10.1021/acsami.7b05253Google Scholar41https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXosVOktbw%253D&md5=869f093a722ed0245c2518483762ca0cIsolation of Phenotypically Distinct Cancer Cells Using Nanoparticle-Mediated SortingGreen, Brenda J.; Kermanshah, Leyla; Labib, Mahmoud; Ahmed, Sharif U.; Silva, Pamuditha N.; Mahmoudian, Laili; Chang, I-Hsin; Mohamadi, Reza M.; Rocheleau, Jonathan V.; Kelley, Shana O.ACS Applied Materials & Interfaces (2017), 9 (24), 20435-20443CODEN: AAMICK; ISSN:1944-8244. (American Chemical Society)Isolating subpopulations of heterogeneous cancer cells is an important capability for the meaningful characterization of circulating tumor cells at different stages of tumor progression and during the epithelial-to-mesenchymal transition. Here, the authors present a microfluidic device that can sep. phenotypically distinct subpopulations of cancer cells. Magnetic nanoparticles coated with antibodies against the epithelial cell adhesion mol. (EpCAM) were used to sep. breast cancer cells in the microfluidic platform. Cells are sorted into different zones on the basis of the levels of EpCAM expression, which enables the detection of cells that are losing epithelial character and becoming more mesenchymal. The phenotypic properties of the isolated cells with low and high EpCAM are then assessed using matrix-coated surfaces for collagen uptake anal. and an NAD(P)H assay that assesses metabolic activity. Low-EpCAM expressing cells have higher collagen uptake and higher folate-induced NAD(P)H responses compared to those of high-EpCAM expressing cells. In addn., the authors tested SKBR3 cancer cells undergoing chem. induced hypoxia. The induced cells have reduced expression of EpCAM, and these cells have higher collagen uptake and NAD(P)H metab. relative to noninduced cells. Nanoparticle-mediated binning facilitates the isolation of functionally distinct cell subpopulations and allows surface marker expression to be assocd. with invasiveness, including collagen uptake and metabolic activity.
- 42Poudineh, M.; Labib, M.; Ahmed, S.; Nguyen, L. N.; Kermanshah, L.; Mohamadi, R. M.; Sargent, E. H.; Kelley, S. O. Profiling Functional and Biochemical Phenotypes of Circulating Tumor Cells Using a Two-Dimensional Sorting Device. Angew. Chem., Int. Ed. 2017, 56 (1), 163– 168, DOI: 10.1002/anie.201608983Google Scholar42https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhvFGqurbJ&md5=6b03fdf7980b9b6a31e6bd1334a4f1d5Profiling Functional and Biochemical Phenotypes of Circulating Tumor Cells Using a Two-Dimensional Sorting DevicePoudineh, Mahla; Labib, Mahmoud; Ahmed, Sharif; Nguyen, L. N. Matthew; Kermanshah, Leyla; Mohamadi, Reza M.; Sargent, Edward H.; Kelley, Shana O.Angewandte Chemie, International Edition (2017), 56 (1), 163-168CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)During cancer progression, tumors shed circulating tumor cells (CTCs) into the bloodstream. CTCs that originate from the same primary tumor can have heterogeneous phenotypes and, while some CTCs possess benign properties, others have high metastatic potential. Deconstructing the heterogeneity of CTCs is challenging and new methods are needed that can sort small nos. of cancer cells according to their phenotypic properties. Here the authors describe a new microfluidic approach that profiles, along two independent phenotypic axes, the behavior of heterogeneous cell subpopulations. Cancer cells are first profiled according to expression of a surface marker using a nanoparticle-enabled approach. Along the second dimension, these subsets are further sepd. into subpopulations corresponding to migration profiles generated in response to a chemotactic agent. The authors deploy this new technique and find a strong correlation between the surface expression and migration potential of CTCs present in blood from mice with xenografted tumors. This system provides an important new means to characterize functional diversity in circulating tumor cells.
- 43Xu, Y.; Zhao, H.; Hou, J. Correlation between overexpression of EpCAM in prostate tissues and genesis of androgen-dependent prostate cancer. Tumor Biol. 2014, 35 (7), 6695– 700, DOI: 10.1007/s13277-014-1892-2Google Scholar43https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXht1egu77F&md5=9f51be7711935b9164ac60da6b4d4980Correlation between overexpression of EpCAM in prostate tissues and genesis of androgen-dependent prostate cancerXu, Yuan; Zhao, Hu; Hou, JianquanTumor Biology (2014), 35 (7), 6695-6700CODEN: TUMBEA; ISSN:1010-4283. (Springer)The objective of this study was to investigate the role of epithelial cell adhesion mol. (EpCAM) in the genesis and the progress of prostate cancer, esp. of castration-resistant prostate cancer. Protein expression of EpCAM in ten pairs of prostate cancer tissues and normal adjacent tissues, plus three cell lines, was examd. Short hairpin RNA (shRNA) interference technique was employed to silence the expression of EpCAM in prostate cancer cell LNCaP and construct a stable transfected cell line. In vitro assay was conducted to analyze the effect of EpCAM expression on the expressions of Androgen receptor (AR), Prostate specific antigen (PSA), and cellular proliferation and invasion. EpCAM was found significantly expressed higher in prostate cancer tissues than in normal adjacent tissues. In three cell lines (DU-145, PC-3, and LNCaP), the expression of EpCAM in LNCaP, androgen-dependent prostate cancer cells, was significantly higher than that in the other two. As EpCAM was silenced in LNCaP, the expression levels of AR and PSA obviously descended, and cellular abilities of proliferation and invasion were obviously inhibited. The overexpression of EpCAM has correlation with the genesis of prostate cancer, esp. androgen-dependent prostate cancer. As the expression of AR is facilitated, prostate cancer cells' abilities to proliferate and invade are consequently enhanced.
- 44Wilbaux, M.; Tod, M.; De Bono, J.; Lorente, D.; Mateo, J.; Freyer, G.; You, B.; Henin, E. A Joint Model for the Kinetics of CTC Count and PSA Concentration During Treatment in Metastatic Castration-Resistant Prostate Cancer. CPT: Pharmacometrics Syst. Pharmacol. 2015, 4 (5), 277– 85, DOI: 10.1002/psp4.34Google Scholar44https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXnsVWrtrw%253D&md5=1094f9d797a067a8607f301896c25f90A Joint Model for the Kinetics of CTC Count and PSA Concentration During Treatment in Metastatic Castration-Resistant Prostate CancerWilbaux, M.; Tod, M.; De Bono, J.; Lorente, D.; Mateo, J.; Freyer, G.; You, B.; Henin, E.CPT: Pharmacometrics & Systems Pharmacology (2015), 4 (5), 277-285CODEN: CPSPBR; ISSN:2163-8306. (Wiley-Blackwell)Assessment of treatment efficacy in metastatic castration-resistant prostate cancer (mCRPC) is limited by frequent nonmeasurable bone metastases. The count of circulating tumor cells (CTCs) is a promising surrogate marker that may replace the widely used prostate-specific antigen (PSA). The purpose of this study was to quantify the dynamic relationships between the longitudinal kinetics of these markers during treatment in patients with mCRPC. Data from 223 patients with mCRPC treated by chemotherapy and/or hormonotherapy were analyzed for up to 6 mo of treatment. A semimechanistic model was built, combining the following several pharmacometric advanced features: (1) Kinetic-Pharmacodynamic (K-PD) compartments for treatments (chemotherapy and hormonotherapy); (2) a latent variable linking both marker kinetics; (3) modeling of CTC kinetics with a cell lifespan model; and (4) a neg. binomial distribution for the CTC random sampling. Linked with survival, this model would potentially be useful for predicting treatment efficacy during drug development or for therapeutic adjustment in treated patients.
- 45Kermanshah, L.; Poudineh, M.; Ahmed, S.; Nguyen, L. N. M.; Srikant, S.; Makonnen, R.; Pena Cantu, F.; Corrigan, M.; Kelley, S. O. Dynamic CTC phenotypes in metastatic prostate cancer models visualized using magnetic ranking cytometry. Lab Chip 2018, 18 (14), 2055– 2064, DOI: 10.1039/C8LC00310FGoogle Scholar45https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXhtFeiurvN&md5=452584c4e071201252af5367831a3fcbDynamic CTC phenotypes in metastatic prostate cancer models visualized using magnetic ranking cytometryKermanshah, Leyla; Poudineh, Mahla; Ahmed, Sharif; Nguyen, L. N. Matthew; Srikant, Sanjana; Makonnen, Rhema; Pena Cantu, Fernando; Corrigan, Michael; Kelley, Shana O.Lab on a Chip (2018), 18 (14), 2055-2064CODEN: LCAHAM; ISSN:1473-0189. (Royal Society of Chemistry)Tumors can shed thousands of cells into the circulation daily. These circulating tumor cells (CTCs) are heterogeneous, and their phenotypes change dynamically. Real-time monitoring of CTC phenotypes is crucial to elucidate the role of CTCs in the metastatic cascade. Here, we monitor phenotypic changes in CTCs in mice xenografted with tumors with varying aggressiveness during cancer progression and a course of chemotherapy to study the metastatic potential of CTCs and changes in the properties of these cells in response to treatment. A new device that enables magnetic ranking cytometry (MagRC) is employed to profile the phenotypic properties of CTCs. Overall, CTCs from metastatic xenografts in mice display dynamic and heterogeneous profiles while non-metastatic models had static profiles. Decreased heterogeneity followed by a redn. in metastasis incidence was obsd. after a course of chemotherapy administered to highly metastatic xenografts. Phenotypic profiling of CTCs could be employed to monitor disease progression and predict therapeutic responses.
- 46Muhanna, N.; Mepham, A.; Mohamadi, R. M.; Chan, H.; Khan, T.; Akens, M.; Besant, J. D.; Irish, J.; Kelley, S. O. Nanoparticle-based sorting of circulating tumor cells by epithelial antigen expression during disease progression in an animal model. Nanomedicine 2015, 11 (7), 1613– 20, DOI: 10.1016/j.nano.2015.04.017Google Scholar46https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXoslCmt7w%253D&md5=613449490a485edcfcbf3df0db75e004Nanoparticle-based sorting of circulating tumor cells by epithelial antigen expression during disease progression in an animal modelMuhanna, Nidal; Mepham, Adam; Mohamadi, Reza M.; Chan, Harley; Khan, Tahsin; Akens, Margarete; Besant, Justin D.; Irish, Jonathan; Kelley, Shana O.Nanomedicine (New York, NY, United States) (2015), 11 (7), 1613-1620CODEN: NANOBF; ISSN:1549-9634. (Elsevier)Circulating tumor cells (CTCs) can be used as markers for the detection, characterization, and targeted therapeutic management of cancer. We recently developed a nanoparticle-mediated approach for capture and sorting of CTCs based on their specific epithelial phenotype. In the current study, we investigate the phenotypic transition of tumor cells in an animal model and show the correlation of this transition with tumor progression. VX2 tumor cells were injected into rabbits, and CTCs were evaluated during tumor progression and correlated with computerized tomog. (CT) measurements of tumor vol. The results showed a dramatic increase of CTCs during the four weeks of tumor growth. Following resection, CTC levels dropped but then rebounded, likely due to lymph node metastases. Addnl., CTCs showed a marked loss of the epithelial cell adhesion mol. (EpCAM) relative to precursor cells. In conclusion, the device accurately traces disease progression and CTC phenotypic shift in an animal model.
- 47Scher, H. I.; Lu, D.; Schreiber, N. A.; Louw, J.; Graf, R. P.; Vargas, H. A.; Johnson, A.; Jendrisak, A.; Bambury, R.; Danila, D.; McLaughlin, B.; Wahl, J.; Greene, S. B.; Heller, G.; Marrinucci, D.; Fleisher, M.; 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 Oncol 2016, 2 (11), 1441– 1449, DOI: 10.1001/jamaoncol.2016.1828Google Scholar47https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC2s%252Fms1Kitw%253D%253D&md5=92be71b45282c41a743d5afc03813167Association of AR-V7 on Circulating Tumor Cells as a Treatment-Specific Biomarker With Outcomes and Survival in Castration-Resistant Prostate CancerScher Howard I; Danila Daniel; Lu David; Louw Jessica; Graf Ryon P; Johnson Ann; Jendrisak Adam; Wahl Justin; Greene Stephanie B; Marrinucci Dena; Dittamore Ryan; Schreiber Nicole A; McLaughlin Brigit; Vargas Hebert A; Bambury Richard; Heller Glenn; Fleisher MartinJAMA oncology (2016), 2 (11), 1441-1449 ISSN:.IMPORTANCE: A critical decision in the management of metastatic castration-resistant prostate cancer (mCRPC) is when to administer an androgen receptor signaling (ARS) inhibitor or a taxane. OBJECTIVE: To determine if pretherapy nuclear androgen-receptor splice variant 7 (AR-V7) protein expression and localization on circulating tumor cells (CTCs) is a treatment-specific marker for response and outcomes between ARS inhibitors and taxanes. DESIGN, SETTING, AND PARTICIPANTS: For this cross-sectional cohort study at Memorial Sloan Kettering Cancer Center, 265 men with progressive mCRPC undergoing a change in treatment were considered; 86 were excluded because they were not initiating ARS or taxane therapy; and 18 were excluded for processing time constraints, leaving 161 patients for analysis. Between December 2012 and March 2015, blood was collected and processed from patients with progressive mCRPC immediately prior to new line of systemic therapy. Patients were followed up to 3 years. MAIN OUTCOMES AND MEASURES: Prostate-specific antigen (PSA) response, time receiving therapy, radiographic progression-free survival (rPFS), and overall survival (OS). RESULTS: Overall, of 193 prospectively collected blood samples from 161 men with mCRPC, 191 were evaluable (128 pre-ARS inhibitor and 63 pretaxane). AR-V7-positive CTCs were found in 34 samples (18%), including 3% of first-line, 18% of second-line, and 31% of third- or greater line samples. Patients whose samples had AR-V7-positive CTCs before ARS inhibition had resistant posttherapy PSA changes (PTPC), shorter rPFS, shorter time on therapy, and shorter OS than those without AR-V7-positive CTCs. Overall, resistant PTPC were seen in 65 of 112 samples (58%) without detectable AR-V7-positive CTCs prior to ARS inhibition. There were statistically significant differences in OS but not in PTPC, time on therapy, or rPFS for patients with or without pretherapy AR-V7-positive CTCs treated with a taxane. A multivariable model adjusting for baseline factors associated with survival showed superior OS with taxanes relative to ARS inhibitors when AR-V7-positive CTCs were detected pretherapy (hazard ratio, 0.24; 95% CI, 0.10-0.57; P = .035). CONCLUSIONS AND RELEVANCE: The results validate CTC nuclear expression of AR-V7 protein in men with mCRPC as a treatment-specific biomarker that is associated with superior survival on taxane therapy over ARS-directed therapy in a clinical practice setting. Continued examination of this biomarker in prospective studies will further aid clinical utility.
- 48Antonarakis, E. S.; Lu, C.; Wang, H.; Luber, B.; Nakazawa, M.; Roeser, J. C.; Chen, Y.; Mohammad, T. A.; Chen, Y.; Fedor, H. L.; Lotan, T. L.; Zheng, Q.; De Marzo, A. M.; Isaacs, J. T.; Isaacs, W. B.; Nadal, R.; Paller, C. J.; Denmeade, S. R.; Carducci, M. A.; Eisenberger, M. A.; Luo, J. AR-V7 and resistance to enzalutamide and abiraterone in prostate cancer. N. Engl. J. Med. 2014, 371 (11), 1028– 38, DOI: 10.1056/NEJMoa1315815Google Scholar48https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhs1Olt73L&md5=c9e06bd306abae0616d963ed82c443eeAR-V7 and resistance to enzalutamide and abiraterone in prostate cancerAntonarakis, Emmanuel S.; Lu, Changxue; Wang, Hao; Luber, Brandon; Nakazawa, Mary; Roeser, Jeffrey C.; Chen, Yan; Mohammad, Tabrez A.; Chen, Yidong; Fedor, Helen L.; Lotan, Tamara L.; Zheng, Qizhi; De Marzo, Angelo M.; Isaacs, John T.; Isaacs, William B.; Nadal, Rosa; Paller, Channing J.; Denmeade, Samuel R.; Carducci, Michael A.; Eisenberger, Mario A.; Luo, JunNew England Journal of Medicine (2014), 371 (11), 1028-1038, 11 pp.CODEN: NEJMAG; ISSN:1533-4406. (Massachusetts Medical Society)Background The androgen-receptor isoform encoded by splice variant 7 lacks the ligand-binding domain, which is the target of enzalutamide and abiraterone, but remains constitutively active as a transcription factor. We hypothesized that detection of androgen-receptor splice variant 7 mRNA (AR-V7) in circulating tumor cells from men with advanced prostate cancer would be assocd. with resistance to enzalutamide and abiraterone. Methods We used a quant. reverse-transcriptase-polymerase-chain-reaction assay to evaluate AR-V7 in circulating tumor cells from prospectively enrolled patients with metastatic castration-resistant prostate cancer who were initiating treatment with either enzalutamide or abiraterone. We examd. assocns. between AR-V7 status (pos. vs. neg.) and prostate-specific antigen (PSA) response rates (the primary end point), freedom from PSA progression (PSA progression-free survival), clin. or radiog. progression-free survival, and overall survival. Results A total of 31 enzalutamide-treated patients and 31 abiraterone-treated patients were enrolled, of whom 39% and 19%, resp., had detectable AR-V7 in circulating tumor cells. Among men receiving enzalutamide, AR-V7-pos. patients had lower PSA response rates than AR-V7-neg. patients (0% vs. 53%, P = 0.004) and shorter PSA progression-free survival (median, 1.4 mo vs. 6.0 mo; P<0.001), clin. or radiog. progression-free survival (median, 2.1 mo vs. 6.1 mo; P<0.001), and overall survival (median, 5.5 mo vs. not reached; P = 0.002). Similarly, among men receiving abiraterone, AR-V7-pos. patients had lower PSA response rates than AR-V7-neg. patients (0% vs. 68%, P = 0.004) and shorter PSA progression-free survival (median, 1.3 mo vs. not reached; P<0.001), clin. or radiog. progression-free survival (median, 2.3 mo vs. not reached; P<0.001), and overall survival (median, 10.6 mo vs. not reached, P = 0.006). The assocn. between AR-V7 detection and therapeutic resistance was maintained after adjustment for expression of full-length androgen receptor mRNA. Conclusions Detection of AR-V7 in circulating tumor cells from patients with castration-resistant prostate cancer may be assocd. with resistance to enzalutamide and abiraterone. These findings require large-scale prospective validation.
- 49Antonarakis, E. S.; Lu, C.; Luber, B.; Wang, H.; Chen, Y.; Nakazawa, M.; Nadal, R.; Paller, C. J.; Denmeade, S. R.; Carducci, M. A.; Eisenberger, M. A.; Luo, J. Androgen Receptor Splice Variant 7 and Efficacy of Taxane Chemotherapy in Patients With Metastatic Castration-Resistant Prostate Cancer. JAMA Oncol 2015, 1 (5), 582– 91, DOI: 10.1001/jamaoncol.2015.1341Google Scholar49https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC28%252FjsVGquw%253D%253D&md5=445419defe501788b64fea6426b58231Androgen Receptor Splice Variant 7 and Efficacy of Taxane Chemotherapy in Patients With Metastatic Castration-Resistant Prostate CancerAntonarakis Emmanuel S; Luber Brandon; Wang Hao; Nadal Rosa; Paller Channing J; Denmeade Samuel R; Carducci Michael A; Eisenberger Mario A; Lu Changxue; Chen Yan; Nakazawa Mary; Luo JunJAMA oncology (2015), 1 (5), 582-91 ISSN:.IMPORTANCE: We previously showed that detection of androgen receptor splice variant 7 (AR-V7) in circulating tumor cells (CTCs) from men with castration-resistant prostate cancer (CRPC) was associated with primary resistance to enzalutamide and abiraterone therapy, but the relevance of AR-V7 status in the context of chemotherapy is unknown. OBJECTIVE: To investigate whether AR-V7-positive patients would retain sensitivity to taxane chemotherapy and whether AR-V7 status would have a differential impact on taxane-treated men compared with enzalutamide- or abiraterone-treated men. DESIGN, SETTING, AND PARTICIPANTS: We examined CTCs for AR-V7 mRNA using a reverse-transcription polymerase chain reaction assay. From January 2013 to July 2014, we prospectively enrolled patients with metastatic CRPC initiating taxane chemotherapy (docetaxel or cabazitaxel) at a single academic institution (Johns Hopkins). Our prespecified statistical plan required a sample size of 36 taxane-treated men. MAIN OUTCOMES AND MEASURES: We evaluated associations between AR-V7 status and prostate-specific antigen (PSA) response rates, PSA progression-free survival (PSA PFS), and clinical and/or radiographic progression-free survival (PFS). After incorporating updated data from our prior study of 62 patients treated with enzalutamide or abiraterone, we also investigated the interaction between AR-V7 status (positive or negative) and treatment type (taxane vs enzalutamide or abiraterone). RESULTS: Of 37 taxane-treated patients enrolled, 17 (46%) had detectable AR-V7 in CTCs. Prostate-specific antigen responses were achieved in both AR-V7-positive and AR-V7-negative men (41% vs 65%; P = .19). Similarly, PSA PFS (hazard ratio [HR], 1.7, 95% CI, 0.6-5.0; P = .32) and PFS (HR, 2.7, 95% CI, 0.8-8.8; P = .11) were comparable in AR-V7-positive and AR-V7-negative patients. A significant interaction was observed between AR-V7 status and treatment type (P < .001). Clinical outcomes were superior with taxanes compared with enzalutamide or abiraterone therapy in AR-V7-positive men, whereas outcomes did not differ by treatment type in AR-V7-negative men. In AR-V7-positive patients, PSA responses were higher in taxane-treated vs enzalutamide- or abiraterone-treated men (41% vs 0%; P < .001), and PSA PFS and PFS were significantly longer in taxane-treated men (HR, 0.19 [95% CI, 0.07-0.52] for PSA PFS, P = .001; HR, 0.21 [95% CI, 0.07-0.59] for PFS, P = .003). CONCLUSIONS AND RELEVANCE: Detection of AR-V7 in CTCs from men with metastatic CRPC is not associated with primary resistance to taxane chemotherapy. In AR-V7-positive men, taxanes appear to be more efficacious than enzalutamide or abiraterone therapy, whereas in AR-V7-negative men, taxanes and enzalutamide or abiraterone may have comparable efficacy. Circulating tumor cell-based AR-V7 detection may serve as a treatment selection biomarker in CRPC.
- 50Crespo, M.; van Dalum, G.; Ferraldeschi, R.; Zafeiriou, Z.; Sideris, S.; Lorente, D.; Bianchini, D.; Rodrigues, D. N.; Riisnaes, R.; Miranda, S.; Figueiredo, I.; Flohr, P.; Nowakowska, K.; de Bono, J. S.; Terstappen, L. W.; Attard, G. Androgen receptor expression in circulating tumour cells from castration-resistant prostate cancer patients treated with novel endocrine agents. Br. J. Cancer 2015, 112 (7), 1166– 74, DOI: 10.1038/bjc.2015.63Google Scholar50https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXls1artrk%253D&md5=95c7a159574eb10d17eec49acbd8c7d8Androgen receptor expression in circulating tumour cells from castration-resistant prostate cancer patients treated with novel endocrine agentsCrespo, M.; van Dalum, G.; Ferraldeschi, R.; Zafeiriou, Z.; Sideris, S.; Lorente, D.; Bianchini, D.; Rodrigues, D. N.; Riisnaes, R.; Miranda, S.; Figueiredo, I.; Flohr, P.; Nowakowska, K.; de Bono, J. S.; Terstappen, L. W. M. M.; Attard, G.British Journal of Cancer (2015), 112 (7), 1166-1174CODEN: BJCAAI; ISSN:0007-0920. (Nature Publishing Group)Background: Abiraterone and enzalutamide are novel endocrine treatments that abrogate androgen receptor (AR) signalling in castration-resistant prostate cancer (CRPC). Here, we developed a circulating tumor cells (CTCs)-based assay to evaluate AR expression in real-time in CRPC and investigated nuclear AR expression in CTCs in patients treated with enzalutamide and abiraterone. Methods: CTCs were captured and characterized using the CellSearch system. An automated algorithm to identify CTCs and quantify AR expression was employed. The primary aim was to evaluate the assocn. between CTC AR expression and prior treatment with abiraterone or enzalutamide. Results: AR expression in CTCs was evaluated in 94 samples from 48 metastatic CRPC patients. We obsd. large intra-patient heterogeneity of AR expression in CTCs. Prior exposure to abiraterone or enzalutamide was not assocd. with a change in CTCs AR expression (median intensity and distribution of AR-pos. classes). In support of this, we also confirmed maintained nuclear AR expression in tissue samples collected after progression on abiraterone. AR staining also identified addnl. AR-pos. CD45-neg. circulating cells that were CK-neg./weak and therefore missed using std. protocols. The no. of these events correlated with traditional CTCs and was assocd. with worse outcome on univariate anal. Conclusions: We developed a non-invasive method to monitor AR nuclear expression in CTCs. Our studies confirm nuclear AR expression in CRPC patients progressing on novel endocrine treatments. Owing to the significant heterogeneity of AR expression in CTCs, studies in larger cohorts of patients are required to identify assocns. with outcome.
- 51Zeinali, M.; Murlidhar, V.; Fouladdel, S.; Shao, S.; Zhao, L.; Cameron, H.; Bankhead, A., III; Shi, J.; Cuneo, K. C.; Sahai, V.; Azizi, E.; Wicha, M. S.; Hafner, M.; Simeone, D. M.; Nagrath, S. Profiling heterogeneous circulating tumor cells (CTC) populations in pancreatic cancer using a serial microfluidic CTC carpet chip. Adv. Biosystems 2018, 2 (12), 1800228, DOI: 10.1002/adbi.201800228Google Scholar51https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXisFSnu7bE&md5=56c8b468d9e306f32d1d240b5d9cc7b1Profiling Heterogeneous Circulating Tumor Cells (CTC) Populations in Pancreatic Cancer Using a Serial Microfluidic CTC Carpet ChipZeinali, Mina; Murlidhar, Vasudha; Fouladdel, Shamileh; Shao, Shimeng; Zhao, Lili; Cameron, Heather; Bankhead, Armand, III; Shi, Jiaqi; Cuneo, Kyle C.; Sahai, Vaibhav; Azizi, Ebrahim; Wicha, Max S.; Hafner, Mathias; Simeone, Diane M.; Nagrath, SunithaAdvanced Biosystems (2018), 2 (12), 1800228/1-1800228/13CODEN: ABDIHL; ISSN:2366-7478. (Wiley-VCH Verlag GmbH & Co. KGaA)Although isolation of circulating tumor cells (CTCs) from pancreatic adenocarcinoma patients is feasible, investigating their clin. utility has proven less successful than other cancers due to the limitations of epithelial cellular adhesion mol. (EpCAM)-only based CTC assays. An integrated technol.- and biol.-based approach using a microfluidic "Carpet Chip" is presented to study the biol. relevance of heterogeneous CTC populations. Both epithelial CTCs (EpCs) and epithelial-to-mesenchymal transition (EMT)-like CTCs (EMTCs) are isolated simultaneously from the whole blood of pancreatic cancer (PaCa) patients (n = 35) by sep. targeting two surface markers: EpCAM and CD133. Recovery of cancer cell lines spiked into whole blood is ≥97% with >76% purity. Thirty-four patients had ≥5 EpCs mL-1 and 35 patients had ≥15 EMTCs mL-1. Overall, significantly higher nos. of EMTCs than EpCs are recovered, reflecting the aggressive nature of PaCa. Furthermore, higher nos. of EMTCs are obsd. in patients with lymph node involvement compared to patients without. Gene expression profiling of CTCs from 17 patients reveals that CXCR1 is significantly upregulated in EpCs, while known stem cell markers POU5F1/Oct-4 and MYC are upregulated in EMTCs. In conclusion, successful isolation and genomic profiling of heterogeneous CTC populations are demonstrated, revealing genetic signatures relevant to patient outcomes.
- 52Kang, Y.-T.; Kim, Y. J.; Bu, J.; Chen, S.; Cho, Y.-H.; Lee, H. M.; Ryu, C. J.; Lim, Y.; Han, S.-W. Epithelial and mesenchymal circulating tumor cell isolation and discrimination using dual-immunopatterned device with newly-developed anti-63B6 and anti-EpCAM. Sens. Actuators, B 2018, 260, 320– 330, DOI: 10.1016/j.snb.2017.12.157Google Scholar52https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXmt1Oisg%253D%253D&md5=cd74b3e899b6fc273801b147fa8899b8Epithelial and mesenchymal circulating tumor cell isolation and discrimination using dual-immunopatterned device with newly-developed anti-63B6 and anti-EpCAMKang, Yoon-Tae; Kim, Young Jun; Bu, Jiyoon; Chen, Shilin; Cho, Young-Ho; Lee, Hyun Min; Ryu, Chun Jeih; Lim, Yoojoo; Han, Sae-WonSensors and Actuators, B: Chemical (2018), 260 (), 320-330CODEN: SABCEB; ISSN:0925-4005. (Elsevier B.V.)We report a dual-immunopatterned microfluidic device for capturing both epithelial and mesenchymal circulating tumor cells (CTCs) simultaneously in a single device using the two antibodies of anti-epithelial cell adhesion mol. (EpCAM) antibody and newly developed anti-63B6 antibody. In addn. to the conventional epithelial antibody of anti-EpCAM, our inhouse produced anti-63B6 antibody targets mesenchymal stem cell-like cancer cells and intermediate cancer cells, thus overcoming limitation of the conventional single anti-EpCAM based CTC isolations. These two antibodies are immobilized to each top and bottom layer of the present device and the combination of two layers forms a circular chamber with the center inlet. The mutual complementary cell isolation through competing reaction in different velocity zones between mesenchymal and epithelial antibodies enables the precise screening and profiling of CTCs depending on their positivity and degree of epithelial/mesenchymal surface antigen expression, which may differ in compliance with disease status. From the expts. using epithelial and mesenchymal-like cancer cells, the present device captures 94.47% of cancer cells, which is substantially higher than the EpCAM-only-based method in terms of heterogeneous cancer cell isolation including mesenchymal-like cancer cells. Patient blood samples were employed to assess the clin. application of the present device for examg. the patient status based on their CTC heterogeneity. The CTCs captured by both antibodies exhibit considerably varied expression profiles of epithelial protein including cytokeratin and EpCAM. The present device facilitates the isolation of heterogeneous subtypes of CTCs. These undiscovered heterogeneities would be helpful for precise anal. of CTCs to find their underlying meaning in cancer progression.
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Abstract
Figure 1
Figure 1. High-efficiency capture of low-EpCAM CTCs from prostate cancer patients. (A,B) Schematic of patient sample collection and processing through the microfluidic device. CTCs are captured from metastatic castrate resistant prostate cancer (mCRPC) patients (n = 36) receiving enzalutamide or abiraterone. Briefly, whole blood is incubated with magnetic nanoparticles (MNPs) conjugated to EpCAM and introduced into the device at a flow rate of 600 μL/h in the presence of an external magnetic field. CTCs are trapped in different zones based on their expression level of EpCAM. (C) LnCaP and PC3 cells were profiled with the microfluidic device. (n = 292 LnCaP cells and n = 343 PC3 cells). High-EpCAM LnCaP cells are trapped primarily in zone 1, whereas low-EpCAM expressing PC3 cells are trapped in zones 3 and 4. (D) Capture efficiency of LnCaP and PC3 cells in the microfluidic device compared to CellSearch (n = 138 LnCaP cells and n = 96 PC3 cells). Cells were spiked in 1 mL of whole blood. Statistics are performed with two-tailed t test, *p < 0.05.
Figure 2
Figure 2. CTC profiles from prostate cancer patients. CTC counts were profiled with the microfluidic device (CTCs/mL) and compared with CellSearch (CTCs/mL) over (A) 0 weeks and (B) 9–22 weeks on-treatment. CTCs were identified as DAPI+/CK+/CD45–. Microfluidic device CTC counts were divided into CTCs captured in low-EpCAM zones 1 and 2 (light gray) and high-EpCAM zones 3 and 4 (black). CTC profiles were obtained from 36 mCRPC patients receiving enzalutamide or abiraterone. The horizontal dashed line indicates data is not available for a particular patient. No bar indicates 0 CTC count. The dotted line separates PSA responsive vs progressive patients, as defined according to PCWG3 criteria. (32) PSA response is defined as a >50% decline from baseline measured twice 3 to 4 weeks apart. PSA progression is defined as the first PSA rise that is ≥25% and ≥2 ng/mL above the nadir, confirmed 3 or more weeks later. If there is no initial decline from baseline, PSA progression is defined as ≥25% and ≥2 ng/mL after 12 weeks.
Figure 3
Figure 3. mCRPC CTCs exhibit a low-EpCAM phenotype. CTCs were profiled with the microfluidic device (CTCs/mL) and separated into (A) high-EpCAM (zone 1 and zone 2) and (B) low-EpCAM (zone 3 and zone 4) counts. (C) CellSearch CTC counts (CTCs/mL). Patients were profiled at 0 weeks and 9–22 weeks on-treatment. The dotted line separates PSA responsive vs progressive patients (refer to definition in the Materials and Methods). CTC counts were obtained from 36 mCRPC patients receiving abiraterone or enzalutamide. CTCs were identified as DAPI+/CK+/CD45–. Each dot represents a patient CTC count. Box plots represent standard error of the mean. The central square represents the mean, and the line represents the median.
Figure 4
Figure 4. Low-EpCAM CTCs exhibit increased ARV7 levels and reduced epithelial properties. (A) Representative immunofluorescent images obtained from patient CTCs isolated from the four zones of the microfluidic device. CTCs are identified as DAPI+/CK+/ARV7+/CD45– and obtained with a 50× objective (CD45 image is not shown in the panel). (B,C) Relative fluorescent intensity of androgen receptor variant 7 (ARV7) and cytokeratin CTCs captured in the four zones of the microfluidic device (n = 76 CTCs, n = 104 CTCs, respectively). Fluorescent intensities are obtained using immunofluorescence image analysis. (D) Percentage of androgen receptor variant 7 (ARV7) CTCs: cytokeratin positive CTCs for responsive versus progressive patients. (E) Percentage of N-cadherin positive CTCs: cytokeratin positive CTCs for responsive versus progressive patients. Zone 3 and zone 4 counts were considered for CTC ratio analysis. CTCs are captured with EpCAM-MNPs in separate microfluidic devices: in the first device, CTCs are identified as DAPI+/CK+/CD45–; in the second device, CTCs are identified as DAPI+/CK+/ARV7+/CD45–; and in the third device, CTCs are identified as DAPI+/NCad+/CD45–. CTC counts are obtained at 0- and 9–22-weeks on-treatment. Statistics were performed with the Mann–Whitney test. *p < 0.05 is significant.
References
ARTICLE SECTIONSThis article references 52 other publications.
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- 6Green, B. J.; Panagiotakopoulou, M.; Pramotton, F. M.; Stefopoulos, G.; Kelley, S. O.; Poulikakos, D.; Ferrari, A. Pore shape defines paths of metastatic cell migration. Nano Lett. 2018, 18, 2140, DOI: 10.1021/acs.nanolett.8b00431Google Scholar6https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXjtlOnur8%253D&md5=c2bc204cfa33e9f955ff9ddf7ac9f537Pore shape defines paths of metastatic cell migrationGreen, Brenda J.; Panagiotakopoulou, Magdalini; Pramotton, Francesca Michela; Stefopoulos, Georgios; Kelley, Shana O.; Poulikakos, Dimos; Ferrari, AldoNano Letters (2018), 18 (3), 2140-2147CODEN: NALEFD; ISSN:1530-6984. (American Chemical Society)Invasion of dense tissues by cancer cells involves the interplay between the penetration resistance offered by interstitial pores and the deformability of cells. Metastatic cancer cells find optimal paths of minimal resistance through an adaptive path-finding process, which leads to successful dissemination. The phys. limit of nuclear deformation is related to the minimal cross section of pores that can be successfully penetrated. However, this single biophys. parameter does not fully describe the architectural complexity of tissues featuring pores of variable area and shape. Here, employing laser nanolithog., we fabricate pore microenvironment models with well-controlled pore shapes, through which human breast cells (MCF10A) and their metastatic offspring (MCF10CA1a.cl1) could pervade. In these exptl. settings, we demonstrate that the actual pore shape, and not only the cross section, is a major and independent determinant of cancer penetration efficiency. In complex architectures contg. pores demanding large deformations from invading cells, tall and narrow rectangular openings facilitate cancer migration. In addn., we highlight the characteristic traits of the explorative behavior enabling metastatic cells to identify and select such pore shapes in a complex multishape pore environment, pinpointing paths of least resistance to invasion.
- 7Green, B. J.; Saberi Safaei, T.; Mepham, A.; Labib, M.; Mohamadi, R. M.; Kelley, S. O. Beyond the Capture of Circulating Tumor Cells: Next-Generation Devices and Materials. Angew. Chem., Int. Ed. 2016, 55 (4), 1252– 65, DOI: 10.1002/anie.201505100Google Scholar7https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhvFKiurvM&md5=44ad3d40a3090e30b791c01cec97beefBeyond the Capture of Circulating Tumor Cells: Next-Generation Devices and MaterialsGreen, Brenda J.; Saberi Safaei, Tina; Mepham, Adam; Labib, Mahmoud; Mohamadi, Reza M.; Kelley, Shana O.Angewandte Chemie, International Edition (2016), 55 (4), 1252-1265CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)A review. Over the last decade, significant progress has been made towards the development of approaches that enable the capture of rare circulating tumor cells (CTCs) from the blood of cancer patients, a crit. capability for noninvasive tumor profiling. These advances have leveraged new insights in materials chem. and microfluidics and allowed the capture and enumeration of CTCs with unprecedented sensitivity. However, it has become increasingly clear that simply capturing and counting tumor cells launched into the bloodstream may not provide the information needed to advance our understanding of the biol. of these rare cells, or to allow us to better exploit them in medicine. A variety of advances have now emerged demonstrating that more information can be extd. from CTCs with next-generation devices and materials featuring tailored phys. and chem. properties. In this Minireview, the last ten years of work in this area will be discussed, with an emphasis on the groundbreaking work of the last five years, during which the focus has moved beyond the simple capture of CTCs and gravitated towards approaches that enable in-depth anal.
- 8Poudineh, M.; Sargent, E. H.; Pantel, K.; Kelley, S. O. Profiling circulating tumour cells and other biomarkers of invasive cancers. Nat. Biomed. Eng. 2018, 2 (2), 72– 84, DOI: 10.1038/s41551-018-0190-5Google Scholar8https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1MXhtFegu7fN&md5=2dff83b64b0f2b1feaeeed716a21eb27Profiling circulating tumour cells and other biomarkers of invasive cancersPoudineh, Mahla; Sargent, Edward H.; Pantel, Klaus; Kelley, Shana O.Nature Biomedical Engineering (2018), 2 (2), 72-84CODEN: NBEAB3; ISSN:2157-846X. (Nature Research)A review. During cancer progression, many tumors shed circulating tumor cells (CTCs) and other biomarkers into the bloodstream. The anal. of CTCs offers the prospect of collecting a liq. biopsy from a patients blood to predict and monitor therapeutic responses and tumor recurrence. In this Review, we discuss progress towards the isolation and recovery of bulk CTCs from whole blood samples for the identification of cells with high metastatic potential. We provide an overview of the techniques that initially pointed to the clin. significance of CTCs and describe the key requirements for clin. applications of CTC anal. We also summarize recent advances that permit the functional and biochem. phenotypes of CTCs to be characterized, and discuss the potential roles of these CTC characteristics in the formation of metastatic lesions. Moreover, we discuss the use of circulating tumor DNA and exosomes as markers for early cancer diagnosis and for the monitoring of cancer progression. Next-generation technologies and biomarkers for invasive cancers should allow for the unequivocal detn. of the metastatic potential of CTCs, and for the meaningful anal. of circulating tumor DNA and exosomes.
- 9Alix-Panabieres, C.; Pantel, K. Challenges in circulating tumour cell research. Nat. Rev. Cancer 2014, 14 (9), 623– 31, DOI: 10.1038/nrc3820Google Scholar9https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhsVWktrfM&md5=c75b540d167fe120266521b8cbf9a6c6Challenges in circulating tumour cell researchAlix-Panabieres, Catherine; Pantel, KlausNature Reviews Cancer (2014), 14 (9), 623-631CODEN: NRCAC4; ISSN:1474-175X. (Nature Publishing Group)A review. During the past ten years, circulating tumor cells (CTCs) have received enormous attention as new biomarkers and the subject of basic research. Although CTCs are already used in numerous clin. trials, their clin. utility is still under investigation. Many issues regarding the detection and characterization of CTCs remain unknown. In this Opinion article, we propose a conceptual framework of CTC assays and point out current challenges of CTC research, which might structure this dynamic field of translational cancer research.
- 10Nagrath, S.; Sequist, L. V.; Maheswaran, S.; Bell, D. W.; Irimia, D.; Ulkus, L.; Smith, M. R.; Kwak, E. L.; Digumarthy, S.; Muzikansky, A.; Ryan, P.; Balis, U. J.; Tompkins, R. G.; Haber, D. A.; Toner, M. Isolation of rare circulating tumour cells in cancer patients by microchip technology. Nature 2007, 450 (7173), 1235– 9, DOI: 10.1038/nature06385Google Scholar10https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD2sXhsVGjsL%252FK&md5=499fb32c8eb07f12f4d1298f0557ea30Isolation of rare circulating tumour cells in cancer patients by microchip technologyNagrath, Sunitha; Sequist, Lecia V.; Maheswaran, Shyamala; Bell, Daphne W.; Irimia, Daniel; Ulkus, Lindsey; Smith, Matthew R.; Kwak, Eunice L.; Digumarthy, Subba; Muzikansky, Alona; Ryan, Paula; Balis, Ulysses J.; Tompkins, Ronald G.; Haber, Daniel A.; Toner, MehmetNature (London, United Kingdom) (2007), 450 (7173), 1235-1239CODEN: NATUAS; ISSN:0028-0836. (Nature Publishing Group)Viable tumor-derived epithelial cells (circulating tumor cells or CTCs) have been identified in peripheral blood from cancer patients and are probably the origin of intractable metastatic disease. Although extremely rare, CTCs represent a potential alternative to invasive biopsies as a source of tumor tissue for the detection, characterization and monitoring of nonhematol. cancers. The ability to identify, isolate, propagate and molecularly characterize CTC subpopulations could further the discovery of cancer stem cell biomarkers and expand the understanding of the biol. of metastasis. Current strategies for isolating CTCs are limited to complex analytic approaches that generate very low yield and purity. Here the authors describe the development of a unique microfluidic platform (the 'CTC-chip') capable of efficient and selective sepn. of viable CTCs from peripheral whole blood samples, mediated by the interaction of target CTCs with antibody (EpCAM)-coated microposts under precisely controlled laminar flow conditions, and without requisite pre-labeling or processing of samples. The CTC-chip successfully identified CTCs in the peripheral blood of patients with metastatic lung, prostate, pancreatic, breast and colon cancer in 115 of 116 (99%) samples, with a range of 5-1,281 CTCs per mL and approx. 50% purity. In addn., CTCs were isolated in 7/7 patients with early-stage prostate cancer. Given the high sensitivity and specificity of the CTC-chip, the authors tested its potential utility in monitoring response to anti-cancer therapy. In a small cohort of patients with metastatic cancer undergoing systemic treatment, temporal changes in CTC nos. correlated reasonably well with the clin. course of disease as measured by std. radiog. methods. Thus, the CTC-chip provides a new and effective tool for accurate identification and measurement of CTCs in patients with cancer. It has broad implications in advancing both cancer biol. research and clin. cancer management, including the detection, diagnosis and monitoring of cancer.
- 11Stott, S. L.; Hsu, C. H.; Tsukrov, D. I.; Yu, M.; Miyamoto, D. T.; Waltman, B. A.; Rothenberg, S. M.; Shah, A. M.; Smas, M. E.; Korir, G. K.; Floyd, F. P., Jr.; Gilman, A. J.; Lord, J. B.; Winokur, D.; Springer, S.; Irimia, D.; Nagrath, S.; Sequist, L. V.; Lee, R. J.; Isselbacher, K. J.; Maheswaran, S.; Haber, D. A.; Toner, M. Isolation of circulating tumor cells using a microvortex-generating herringbone-chip. Proc. Natl. Acad. Sci. U. S. A. 2010, 107 (43), 18392– 7, DOI: 10.1073/pnas.1012539107Google Scholar11https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3cXhtl2ktr3N&md5=f0896b84a56d0dcdcdb0207be71650ceIsolation of circulating tumor cells using a microvortex-generating herringbone-chipStott, Shannon L.; Hsua, Chia-Hsien; Tsukrov, Dina I.; Yu, Min; Miyamoto, David T.; Waltman, Belinda A.; Rothenberg, S. Michael; Shah, Ajay M.; Smas, Malgorzata E.; Korir, George K.; Floyd, Frederick P., Jr.; Gilman, Anna J.; Lord, Jenna B.; Winokur, Daniel; Springer, Simeon; Irimia, Daniel; Nagrath, Sunitha; Sequist, Lecia V.; Lee, Richard J.; Isselbacher, Kurt J.; Maheswaran, Shyamala; Haber, Daniel A.; Toner, MehmetProceedings of the National Academy of Sciences of the United States of America (2010), 107 (43), 18392-18397, S18392/1-S18392/9CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)Rare circulating tumor cells (CTCs) present in the bloodstream of patients with cancer provide a potentially accessible source for detection, characterization, and monitoring of nonhematol. cancers. We previously demonstrated the effectiveness of a microfluidic device, the CTC-Chip, in capturing these epithelial cell adhesion mol. (EpCAM)-expressing cells using antibody-coated microposts. Here, we describe a high-throughput microfluidic mixing device, the herringbone-chip, or "HB-Chip," which provides an enhanced platform for CTC isolation. The HB-Chip design applies passive mixing of blood cells through the generation of microvortices to significantly increase the no. of interactions between target CTCs and the antibody-coated chip surface. Efficient cell capture was validated using defined nos. of cancer cells spiked into control blood, and clin. utility was demonstrated in specimens from patients with prostate cancer. CTCs were detected in 14 of 15 (93%) patients with metastatic disease (median = 63 CTCs/mL, mean = 386 ± 238 CTCs/mL), and the tumor-specific TMPRSS2-ERG translocation was readily identified following RNA isolation and RT-PCR anal. The use of transparent materials allowed for imaging of the captured CTCs using std. clin. histopathol. stains, in addn. to immunofluorescence-conjugated antibodies. In a subset of patient samples, the low shear design of the HB-Chip revealed microclusters of CTCs, previously unappreciated tumor cell aggregates that may contribute to the hematogenous dissemination of cancer.
- 12Yoon, H. J.; Kim, T. H.; Zhang, Z.; Azizi, E.; Pham, T. M.; Paoletti, C.; Lin, J.; Ramnath, N.; Wicha, M. S.; Hayes, D. F.; Simeone, D. M.; Nagrath, S. Sensitive capture of circulating tumour cells by functionalized graphene oxide nanosheets. Nat. Nanotechnol. 2013, 8 (10), 735– 41, DOI: 10.1038/nnano.2013.194Google Scholar12https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhsFaksbzL&md5=a6ac26c2fdf8b2e6c93bd63f6634a934Sensitive capture of circulating tumor cells by functionalized graphene oxide nanosheetsYoon, Hyeun Joong; Kim, Tae Hyun; Zhang, Zhuo; Azizi, Ebrahim; Pham, Trinh M.; Paoletti, Costanza; Lin, Jules; Ramnath, Nithya; Wicha, Max S.; Hayes, Daniel F.; Simeone, Diane M.; Nagrath, SunithaNature Nanotechnology (2013), 8 (10), 735-741CODEN: NNAABX; ISSN:1748-3387. (Nature Publishing Group)The spread of cancer throughout the body is driven by circulating tumor cells (CTCs). These cells detach from the primary tumor and move from the bloodstream to a new site of subsequent tumor growth. They also carry information about the primary tumor and have the potential to be valuable biomarkers for disease diagnosis and progression, and for the mol. characterization of certain biol. properties of the tumor. However, the limited sensitivity and specificity of current methods for measuring and studying these cells in patient blood samples prevents the realization of their full clin. potential. The use of microfluidic devices is a promising method for isolating CTCs. However, the devices are reliant on three-dimensional structures, which limits further characterization and expansion of cells on the chip. Here we demonstrate an effective approach to isolating CTCs from blood samples of pancreatic, breast and lung cancer patients, by using functionalized graphene oxide nanosheets on a patterned gold surface. CTCs were captured with high sensitivity at a low concn. of target cells (73 ± 32.4% at 3-5 cells per mL blood).
- 13Zhang, Y.; Wu, M.; Han, X.; Wang, P.; Qin, L. High-Throughput, Label-Free Isolation of Cancer Stem Cells on the Basis of Cell Adhesion Capacity. Angew. Chem., Int. Ed. 2015, 54 (37), 10838– 42, DOI: 10.1002/anie.201505294Google Scholar13https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhtFyqtb%252FK&md5=f328026d6af1e22c6b23800d804e0559High-Throughput, Label-Free Isolation of Cancer Stem Cells on the Basis of Cell Adhesion CapacityZhang, Yuanqing; Wu, Minhao; Han, Xin; Wang, Ping; Qin, LidongAngewandte Chemie, International Edition (2015), 54 (37), 10838-10842CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)Herein the authors report a microfluidics method that enriches cancer stem cells (CSCs) or tumor-initiating cells on the basis of cell adhesion properties. In the authors' on-chip enrichment system, cancer cells were driven by hydrodynamic forces to flow through microchannels coated with basement membrane ext. Highly adhesive cells were captured by the functionalized microchannels, and less adhesive cells were collected from the outlets. Two heterogeneous breast cancer cell lines (SUM-149 and SUM-159) were successfully sepd. into enriched subpopulations according to their adhesive capacity, and the enrichment of the cancer stem cells was confirmed by flow cytometry biomarker anal. and tumor-formation assays. The authors' findings show that the less adhesive phenotype is assocd. with a higher percentage of CSCs, higher cancer-cell motility, and higher resistance to chemotherapeutic drugs.
- 14Witek, M. A.; Aufforth, R. D.; Wang, H.; Kamande, J. W.; Jackson, J. M.; Pullagurla, S. R.; Hupert, M. L.; Usary, J.; Wysham, W. Z.; Hilliard, D.; Montgomery, S.; Bae-Jump, V.; Carey, L. A.; Gehrig, P. A.; Milowsky, M. I.; Perou, C. M.; Soper, J. T.; Whang, Y. E.; Yeh, J. J.; Martin, G.; Soper, S. A. Discrete microfluidics for the isolation of circulating tumor cell subpopulations targeting fibroblast activation protein alpha and epithelial cell adhesion molecule. npj Precis. Oncol. 2017, 1, 24, DOI: 10.1038/s41698-017-0028-8Google ScholarThere is no corresponding record for this reference.
- 15Liu, Z.; Lee, Y.; Jang, J.; Li, Y.; Han, X.; Yokoi, K.; Ferrari, M.; Zhou, L.; Qin, L. Microfluidic cytometric analysis of cancer cell transportability and invasiveness. Sci. Rep. 2015, 5, 14272, DOI: 10.1038/srep14272Google Scholar15https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhsFOktr%252FE&md5=67492ced562b1ce9c51d65ab4f285772Microfluidic cytometric analysis of cancer cell transportability and invasivenessLiu, Zongbin; Lee, Yeonju; Jang, Joon hee; Li, Ying; Han, Xin; Yokoi, Kenji; Ferrari, Mauro; Zhou, Ledu; Qin, LidongScientific Reports (2015), 5 (), 14272CODEN: SRCEC3; ISSN:2045-2322. (Nature Publishing Group)The extensive phenotypic and functional heterogeneity of cancer cells plays an important role in tumor progression and therapeutic resistance. Characterizing this heterogeneity and identifying invasive phenotype may provide possibility to improve chemotherapy treatment. By mimicking cancer cell perfusion through circulatory system in metastasis, we develop a unique microfluidic cytometry (MC) platform to sep. cancer cells at high throughput, and further derive a phys. parameter 'transportability' to characterize the ability to pass through micro-constrictions. The transportability is detd. by cell stiffness and cell-surface frictional property, and can be used to probe tumor heterogeneity, discriminate more invasive phenotypes and correlate with biomarker expressions in breast cancer cells. Decreased cell stiffness and cell-surface frictional force leads to an increase in transportability and may be a feature of invasive cancer cells by promoting cell perfusion through narrow spaces in circulatory system. The MC-Chip provides a promising microfluidic platform for studying cell mechanics and transportability could be used as a novel marker for probing tumor heterogeneity and detg. invasive phenotypes.
- 16Yoneda, K.; Kuwata, T.; Chikaishi, Y.; Mori, M.; Kanayama, M.; Takenaka, M.; Oka, S.; Hirai, A.; Imanishi, N.; Kuroda, K.; Ichiki, Y.; Ohnaga, T.; Tanaka, F. Detection of circulating tumor cells with a novel microfluidic system in malignant pleural mesothelioma. Cancer Sci. 2019, 110 (2), 726– 733, DOI: 10.1111/cas.13895Google Scholar16https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1MXks1WmtQ%253D%253D&md5=21dfae540fa78ad82545b48665c809a0Detection of circulating tumor cells with a novel microfluidic system in malignant pleural mesotheliomaYoneda, Kazue; Kuwata, Taiji; Chikaishi, Yasuhiro; Mori, Masataka; Kanayama, Masatoshi; Takenaka, Masaru; Oka, Soichi; Hirai, Ayako; Imanishi, Naoko; Kuroda, Koji; Ichiki, Yoshinobu; Ohnaga, Takashi; Tanaka, FumihiroCancer Science (2019), 110 (2), 726-733CODEN: CSACCM; ISSN:1349-7006. (Wiley-Blackwell)Detection of rare tumor cells circulating in the blood (CTCs) presents tech. challenges. CellSearch, the only approved system for clin. use, fails to capture epithelial cell adhesion mol.-neg. CTCs such as malignant pleural mesothelioma (MPM). We have developed a novel microfluidic device (CTC-chip) in which any Ab to capture CTCs is conjugated. The CTC-chip was coated with an Ab against podoplanin that is abundantly expressed on MPM. Circulating tumor cell-detection performance was evaluated in exptl. models in which MPM cells were spiked in blood sampled from a healthy volunteer and in clin. samples drawn from MPM patients. The CTC-chip showed superior CTC-detection performance over CellSearch in exptl. models (sensitivity, 63.3%-64.5% vs 0%-1.1%; P < .001) and in clin. samples (CTC-positivity, 68.8% vs 6.3%; P < .001). A receiver operating characteristic (ROC) anal. showed that the CTC test provided a significant diagnostic performance in discrimination of unresectable disease from resectable disease (area under the ROC curve, 0.851; P = .003). The higher CTC count (=2 cells/mL) was significantly assocd. with a poor prognosis (P = .030). The novel CTC-chip enabled sensitive detection of CTCs, which provided significant diagnostic and prognostic information in MPM.
- 17Byun, S.; Son, S.; Amodei, D.; Cermak, N.; Shaw, J.; Kang, J. H.; Hecht, V. C.; Winslow, M. M.; Jacks, T.; Mallick, P.; Manalis, S. R. Characterizing deformability and surface friction of cancer cells. Proc. Natl. Acad. Sci. U. S. A. 2013, 110 (19), 7580– 5, DOI: 10.1073/pnas.1218806110Google Scholar17https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXptFGrtrc%253D&md5=518975ec5375f00df12e70e6567b64f4Characterizing deformability and surface friction of cancer cellsByun, Sangwon; Son, Sungmin; Amodei, Dario; Cermak, Nathan; Shaw, Josephine; Kang, Joon Ho; Hecht, Vivian C.; Winslow, Monte M.; Jacks, Tyler; Mallick, Parag; Manalis, Scott R.Proceedings of the National Academy of Sciences of the United States of America (2013), 110 (19), 7580-7585, S7580/1-S7580/10CODEN: PNASA6; ISSN:0027-8424. (National Academy of Sciences)Metastasis requires the penetration of cancer cells through tight spaces, which is mediated by the phys. properties of the cells as well as their interactions with the confined environment. Various microfluidic approaches have been devised to mimic traversal in vitro by measuring the time required for cells to pass through a constriction. Although a cell's passage time is expected to depend on its deformability, measurements from existing approaches are confounded by a cell's size and its frictional properties with the channel wall. Here, we introduce a device that enables the precise measurement of (i) the size of a single cell, given by its buoyant mass, (ii) the velocity of the cell entering a constricted microchannel (entry velocity), and (iii) the velocity of the cell as it transits through the constriction (transit velocity). Changing the deformability of the cell by perturbing its cytoskeleton primarily alters the entry velocity, whereas changing the surface friction by immobilizing pos. charges on the constriction's walls primarily alters the transit velocity, indicating that these parameters can give insight into the factors affecting the passage of each cell. When accounting for cell buoyant mass, we find that cells possessing higher metastatic potential exhibit faster entry velocities than cells with lower metastatic potential. We addnl. find that some cell types with higher metastatic potential exhibit greater than expected changes in transit velocities, suggesting that not only the increased deformability but reduced friction may be a factor in enabling invasive cancer cells to efficiently squeeze through tight spaces.
- 18Shaw Bagnall, J.; Byun, S.; Begum, S.; Miyamoto, D. T.; Hecht, V. C.; Maheswaran, S.; Stott, S. L.; Toner, M.; Hynes, R. O.; Manalis, S. R. Deformability of Tumor Cells versus Blood Cells. Sci. Rep. 2015, 5, 18542, DOI: 10.1038/srep18542Google Scholar18https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXitVKrsLbF&md5=36365ef18dad7735c0b6177cc56d5a8fDeformability of Tumor Cells versus Blood CellsShaw Bagnall, Josephine; Byun, Sangwon; Begum, Shahinoor; Miyamoto, David T.; Hecht, Vivian C.; Maheswaran, Shyamala; Stott, Shannon L.; Toner, Mehmet; Hynes, Richard O.; Manalis, Scott R.Scientific Reports (2015), 5 (), 18542CODEN: SRCEC3; ISSN:2045-2322. (Nature Publishing Group)The potential for circulating tumor cells (CTCs) to elucidate the process of cancer metastasis and inform clin. decision-making has made their isolation of great importance. However, CTCs are rare in the blood, and universal properties with which to identify them remain elusive. As technol. advancements have made single-cell deformability measurements increasingly routine, the assessment of phys. distinctions between tumor cells and blood cells may provide insight into the feasibility of deformability-based methods for identifying CTCs in patient blood. To this end, we present an initial study assessing deformability differences between tumor cells and blood cells, indicated by the length of time required for them to pass through a microfluidic constriction. Here, we demonstrate that deformability changes in tumor cells that have undergone phenotypic shifts are small compared to differences between tumor cell lines and blood cells. Addnl., in a syngeneic mouse tumor model, cells that are able to exit a tumor and enter circulation are not required to be more deformable than the cells that were first injected into the mouse. However, a limited study of metastatic prostate cancer patients provides evidence that some CTCs may be more mech. similar to blood cells than to typical tumor cell lines.
- 19Sarioglu, A. F.; Aceto, N.; Kojic, N.; Donaldson, M. C.; Zeinali, M.; Hamza, B.; Engstrom, A.; Zhu, H.; Sundaresan, T. K.; Miyamoto, D. T.; Luo, X.; Bardia, A.; Wittner, B. S.; Ramaswamy, S.; Shioda, T.; Ting, D. T.; Stott, S. L.; Kapur, R.; Maheswaran, S.; Haber, D. A.; Toner, M. A microfluidic device for label-free, physical capture of circulating tumor cell clusters. Nat. Methods 2015, 12 (7), 685– 91, DOI: 10.1038/nmeth.3404Google Scholar19https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhtFWqtbbE&md5=6ce523882376f88875f506e33cb04b22A microfluidic device for label-free, physical capture of circulating tumor cell clustersSarioglu, A. Fatih; Aceto, Nicola; Kojic, Nikola; Donaldson, Maria C.; Zeinali, Mahnaz; Hamza, Bashar; Engstrom, Amanda; Zhu, Huili; Sundaresan, Tilak K.; Miyamoto, David T.; Luo, Xi; Bardia, Aditya; Wittner, Ben S.; Ramaswamy, Sridhar; Shioda, Toshi; Ting, David T.; Stott, Shannon L.; Kapur, Ravi; Maheswaran, Shyamala; Haber, Daniel A.; Toner, MehmetNature Methods (2015), 12 (7), 685-691CODEN: NMAEA3; ISSN:1548-7091. (Nature Publishing Group)Cancer cells metastasize through the bloodstream either as single migratory circulating tumor cells (CTCs) or as multicellular groupings (CTC clusters). Existing technologies for CTC enrichment are designed to isolate single CTCs, and although CTC clusters are detectable in some cases, their true prevalence and significance remain to be detd. Here we developed a microchip technol. (the Cluster-Chip) to capture CTC clusters independently of tumor-specific markers from unprocessed blood. CTC clusters are isolated through specialized bifurcating traps under low-shear stress conditions that preserve their integrity, and even two-cell clusters are captured efficiently. Using the Cluster-Chip, we identified CTC clusters in 30-40% of patients with metastatic breast or prostate cancer or with melanoma. RNA sequencing of CTC clusters confirmed their tumor origin and identified tissue-derived macrophages within the clusters. Efficient capture of CTC clusters will enable the detailed characterization of their biol. properties and role in metastasis.
- 20Poudineh, M.; Aldridge, P. M.; Ahmed, S.; Green, B. J.; Kermanshah, L.; Nguyen, V.; Tu, C.; Mohamadi, R. M.; Nam, R. K.; Hansen, A.; Sridhar, S. S.; Finelli, A.; Fleshner, N. E.; Joshua, A. M.; Sargent, E. H.; Kelley, S. O. Tracking the dynamics of circulating tumour cell phenotypes using nanoparticle-mediated magnetic ranking. Nat. Nanotechnol. 2017, 12 (3), 274– 281, DOI: 10.1038/nnano.2016.239Google Scholar20https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhvFahtLbL&md5=484c655c7f6d0f4ef882e913d52ca5d4Tracking the dynamics of circulating tumour cell phenotypes using nanoparticle-mediated magnetic rankingPoudineh, Mahla; Aldridge, Peter M.; Ahmed, Sharif; Green, Brenda J.; Kermanshah, Leyla; Nguyen, Vivian; Tu, Carmen; Mohamadi, Reza M.; Nam, Robert K.; Hansen, Aaron; Sridhar, Srikala S.; Finelli, Antonio; Fleshner, Neil E.; Joshua, Anthony M.; Sargent, Edward H.; Kelley, Shana O.Nature Nanotechnology (2017), 12 (3), 274-281CODEN: NNAABX; ISSN:1748-3387. (Nature Publishing Group)Profiling the heterogeneous phenotypes of rare circulating tumor cells (CTCs) in whole blood is crit. to unraveling the complex and dynamic properties of these potential clin. markers. This task is challenging because these cells are present at ppb levels among normal blood cells. Here the authors report a new nanoparticle-enabled method for CTC characterization, called magnetic ranking cytometry, which profiles CTCs on the basis of their surface expression phenotype. The authors achieve this using a microfluidic chip that successfully processes whole blood samples. The approach classifies CTCs with single-cell resoln. in accordance with their expression of phenotypic surface markers, which is read out using magnetic nanoparticles. The authors deploy this new technique to reveal the dynamic phenotypes of CTCs in unprocessed blood from mice as a function of tumor growth and aggressiveness. The authors also test magnetic ranking cytometry using blood samples collected from cancer patients.
- 21Labib, M.; Mohamadi, R. M.; Poudineh, M.; Ahmed, S. U.; Ivanov, I.; Huang, C. L.; Moosavi, M.; Sargent, E. H.; Kelley, S. O. Single-cell mRNA cytometry via sequence-specific nanoparticle clustering and trapping. Nat. Chem. 2018, 10 (5), 489– 495, DOI: 10.1038/s41557-018-0025-8Google Scholar21https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXos1Smtr8%253D&md5=dd6ba074a864f04d2ef145e4f8ef1131Single-cell mRNA cytometry via sequence-specific nanoparticle clustering and trappingLabib, Mahmoud; Mohamadi, Reza M.; Poudineh, Mahla; Ahmed, Sharif U.; Ivanov, Ivaylo; Huang, Ching-Lung; Moosavi, Maral; Sargent, Edward H.; Kelley, Shana O.Nature Chemistry (2018), 10 (5), 489-495CODEN: NCAHBB; ISSN:1755-4330. (Nature Research)Cell-to-cell variation in gene expression creates a need for techniques that can characterize expression at the level of individual cells. This is particularly true for rare circulating tumor cells, in which subtyping and drug resistance are of intense interest. Here the authors describe a method for cell anal.-single-cell mRNA cytometry-that enables the isolation of rare cells from whole blood as a function of target mRNA sequences. This approach uses two classes of magnetic particles that are labeled to selectively hybridize with different regions of the target mRNA. Hybridization leads to the formation of large magnetic clusters that remain localized within the cells of interest, thereby enabling the cells to be magnetically sepd. Targeting specific intracellular mRNAs enables circulating tumor cells to be distinguished from normal hematopoietic cells. No polymerase chain reaction amplification is required to det. RNA expression levels and genotype at the single-cell level, and minimal cell manipulation is required. To demonstrate this approach the authors use single-cell mRNA cytometry to detect clin. important sequences in prostate cancer specimens.
- 22Pantel, K.; Brakenhoff, R. H.; Brandt, B. Detection, clinical relevance and specific biological properties of disseminating tumour cells. Nat. Rev. Cancer 2008, 8 (5), 329– 40, DOI: 10.1038/nrc2375Google Scholar22https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BD1cXltVGlurs%253D&md5=a8b4dea66d28b6705e67933ed7e65a82Detection, clinical relevance and specific biological properties of disseminating tumour cellsPantel, Klaus; Brakenhoff, Ruud H.; Brandt, BurkhardNature Reviews Cancer (2008), 8 (5), 329-340CODEN: NRCAC4; ISSN:1474-175X. (Nature Publishing Group)A review. Most cancer deaths are caused by hematogenous metastatic spread and subsequent growth of tumor cells at distant organs. Disseminating tumor cells present in the peripheral blood and bone marrow can now be detected and characterized at the single-cell level. These cells are highly relevant to the study of the biol. of early metastatic spread and provide a diagnostic source in patients with overt metastases. Here we review the evidence that disseminating tumor cells have a variety of uses for understanding tumor biol. and improving cancer treatment.
- 23Ligthart, S. T.; Coumans, F. A. W.; Attard, G.; Mulick Cassidy, A.; de Bono, J. S.; Terstappen, L. W. M. M. Unbiased and automated identification of a circulating tumour cell definition that associates with overall survival. PLoS One 2011, 6 (11), e27419, DOI: 10.1371/journal.pone.0027419Google Scholar23https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXhsFemtrzJ&md5=cc3f4373514ac79444d0179d5343dcd9Unbiased and automated identification of a circulating tumour cell definition that associates with overall survivalLigthart, Sjoerd T.; Coumans, Frank A. W.; Attard, Gerhardt; Cassidy, Amy Mulick; de Bono, Johann S.; Terstappen, Leon W. M. M.PLoS One (2011), 6 (11), e27419CODEN: POLNCL; ISSN:1932-6203. (Public Library of Science)Circulating tumor cells (CTC) in patients with metastatic carcinomas are assocd. with poor survival and can be used to guide therapy. Classification of CTC however remains subjective, as they are morphol. heterogeneous. We acquired digital images, using the CellSearch system, from blood of 185 castration resistant prostate cancer (CRPC) patients and 68 healthy subjects to define CTC by computer algorithms. Patient survival data was used as the training parameter for the computer to define CTC. The computer-generated CTC definition was validated on a sep. CRPC dataset comprising 100 patients. The optimal definition of the computer defined CTC (aCTC) was stricter as compared to the manual CellSearch CTC (mCTC) definition and as a consequence aCTC were less frequent. The computer-generated CTC definition resulted in hazard ratios (HRs) of 2.8 for baseline and 3.9 for follow-up samples, which is comparable to the mCTC definition (baseline HR 2.9, follow-up HR 4.5). Validation resulted in HRs at baseline/follow-up of 3.9/5.4 for computer and 4.8/5.8 for manual definitions. In conclusion, we have defined and validated CTC by clin. outcome using a perfectly reproducing automated algorithm.
- 24Watson, P. A.; Arora, V. K.; Sawyers, C. L. Emerging mechanisms of resistance to androgen receptor inhibitors in prostate cancer. Nat. Rev. Cancer 2015, 15 (12), 701– 11, DOI: 10.1038/nrc4016Google Scholar24https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXhvVCqtLnN&md5=98aaa704d576f2954cc1d83e474c0863Emerging mechanisms of resistance to androgen receptor inhibitors in prostate cancerWatson, Philip A.; Arora, Vivek K.; Sawyers, Charles L.Nature Reviews Cancer (2015), 15 (12), 701-711CODEN: NRCAC4; ISSN:1474-175X. (Nature Publishing Group)A review. During the past 10 years, preclin. studies implicating sustained androgen receptor (AR) signaling as the primary driver of castration-resistant prostate cancer (CRPC) have led to the development of novel agents targeting the AR pathway that are now in widespread clin. use. These drugs prolong the survival of patients with late-stage prostate cancer but are not curative. In this Review, we highlight emerging mechanisms of acquired resistance to these contemporary therapies, which fall into the three broad categories of restored AR signaling, AR bypass signaling and complete AR independence. This diverse range of resistance mechanisms presents new challenges for long-term disease control, which may be addressable through early use of combination therapies guided by recent insights from genomic landscape studies of CRPC.
- 25Lowes, L. E.; Goodale, D.; Xia, Y.; Postenka, C.; Piaseczny, M. M.; Paczkowski, F.; Allan, A. L. Epithelial-to-mesenchymal transition leads to disease-stage differences in circulating tumor cell detection and metastasis in pre-clinical models of prostate cancer. Oncotarget 2016, 7 (46), 76125– 76139, DOI: 10.18632/oncotarget.12682Google Scholar25https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC2srksFWlsg%253D%253D&md5=f6037b90eac2d99d28bbc5ea4ce47493Epithelial-to-mesenchymal transition leads to disease-stage differences in circulating tumor cell detection and metastasis in pre-clinical models of prostate cancerLowes Lori E; Piaseczny Matthew M; Allan Alison L; Goodale David; Xia Ying; Postenka Carl; Paczkowski Freeman; Allan Alison L; Allan Alison L; Allan Alison LOncotarget (2016), 7 (46), 76125-76139 ISSN:.Metastasis is the cause of most prostate cancer (PCa) deaths and has been associated with circulating tumor cells (CTCs). The presence of ≥5 CTCs/7.5mL of blood is a poor prognosis indicator in metastatic PCa when assessed by the CellSearch® system, the "gold standard" clinical platform. However, ~35% of metastatic PCa patients assessed by CellSearch® have undetectable CTCs. We hypothesize that this is due to epithelial-to-mesenchymal transition (EMT) and subsequent loss of necessary CTC detection markers, with important implications for PCa metastasis. Two pre-clinical assays were developed to assess human CTCs in xenograft models; one comparable to CellSearch® (EpCAM-based) and one detecting CTCs semi-independent of EMT status via combined staining with EpCAM/HLA (human leukocyte antigen). In vivo differences in CTC generation, kinetics, metastasis and EMT status were determined using 4 PCa models with progressive epithelial (LNCaP, LNCaP-C42B) to mesenchymal (PC-3, PC-3M) phenotypes. Assay validation demonstrated that the CellSearch®-based assay failed to detect a significant number (~40-50%) of mesenchymal CTCs. In vivo, PCa with an increasingly mesenchymal phenotype shed greater numbers of CTCs more quickly and with greater metastatic capacity than PCa with an epithelial phenotype. Notably, the CellSearch®-based assay captured the majority of CTCs shed during early-stage disease in vivo, and only after establishment of metastases were a significant number of undetectable CTCs present. This study provides important insight into the influence of EMT on CTC generation and subsequent metastasis, and highlights that novel technologies aimed at capturing mesenchymal CTCs may only be useful in the setting of advanced metastatic disease.
- 26Antonarakis, E. S.; Lu, C.; Luber, B.; Wang, H.; Chen, Y.; Zhu, Y.; Silberstein, J. L.; Taylor, M. N.; Maughan, B. L.; Denmeade, S. R.; Pienta, K. J.; Paller, C. J.; Carducci, M. A.; Eisenberger, M. A.; Luo, J. Clinical Significance of Androgen Receptor Splice Variant-7 mRNA Detection in Circulating Tumor Cells of Men With Metastatic Castration-Resistant Prostate Cancer Treated With First- and Second-Line Abiraterone and Enzalutamide. J. Clin. Oncol. 2017, 35 (19), 2149– 2156, DOI: 10.1200/JCO.2016.70.1961Google Scholar26https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXhvFKnsL%252FK&md5=471447974648fa52958ea71a7959637eClinical Significance of androgen receptor splice variant-7 mRNA detection in circulating tumor cells of men with metastatic castration-resistant prostate cancer treated With first- and second-line abiraterone and enzalutamideAntonarakis, Emmanuel S.; Lu, Changxue; Luber, Brandon; Wang, Hao; Chen, Yan; Zhu, Yezi; Silberstein, John L.; Taylor, Maritza N.; Maughan, Benjamin L.; Denmeade, Samuel R.; Pienta, Kenneth J.; Paller, Channing J.; Carducci, Michael A.; Eisenberger, Mario A.; Luo, JunJournal of Clinical Oncology (2017), 35 (19), 2149-2157CODEN: JCONDN; ISSN:1527-7755. (American Society of Clinical Oncology)Purpose We reported previously that the detection of androgen receptor splice variant-7 (AR-V7) mRNA in circulating tumor cells (CTCs) correlated with poor outcomes from the use of abiraterone and enzalutamide in patients with castration-resistant prostate cancer (CRPC). Here, we expanded our cohort size to better characterize the prognostic significance of AR-V7 in this setting. Methods We prospectively enrolled 202 patients with CRPC starting abiraterone or enzalutamide and investigated the prognostic value of CTC detection (+ v -) and AR-V7 detection (+ v -) using a CTCbased AR-V7 mRNA assay. We examd. $ 50% prostate-specific antigen (PSA) responses, PSA progression-free survival, clin. and radiol. progression-free survival, and overall survival. We constructed multivariable models adjusting for PSA, Gleason sum, no. of prior hormone therapies, prior abiraterone or enzalutamide use, prior taxane use, presence of visceral metastases, and Eastern Cooperative Oncol. Group score. We also sep. examd. the first-line and second-line novel hormonal therapy (NHT) settings. Results Median follow-up times were 15.0, 21.7, and 14.6months for CTC-, CTC+/AR-V7- and CTC+/AR-V7+ patients, resp. CTC+/AR-V7+ patients were more likely to have Gleason scores ≥8 (P =.05), metastatic disease at diagnosis (P =.01), higher PSA (P <.01), prior abiraterone or enzalutamide use (P =.03), prior taxane use (P =.02), and Eastern Cooperative Oncol. Group ≥1 (P =.01). Outcomes for the overall cohort (and sep. for the first-line and second-line NHT cohorts) were best for CTC- patients, intermediate for CTC+/AR-V7- patients, andworse for CTC+/AR-V7+ patients. These correlations remained significant in multivariable models. Conclusion This expanded anal. further characterizes the importance of CTC-based AR-V7 mRNA detection in predicting outcomes in patients with CRPC receiving first- and second-line NHT and, to the best of our knowledge, is the first to suggest that this assay be interpreted using three sep. prognostic categories: CTC-, CTC+/AR-V7-, and CTC+/AR-V7+.
- 27Satelli, A.; Batth, I.; Brownlee, Z.; Mitra, A.; Zhou, S.; Noh, H.; Rojas, C. R.; Li, H.; Meng, Q. H.; Li, S. EMT circulating tumor cells detected by cell-surface vimentin are associated with prostate cancer progression. Oncotarget 2017, 8 (30), 49329– 49337, DOI: 10.18632/oncotarget.17632Google Scholar27https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1crovFOktQ%253D%253D&md5=c167a5a77af07996b0f8b7f76ce658ecEMT circulating tumor cells detected by cell-surface vimentin are associated with prostate cancer progressionSatelli Arun; Batth Izhar; Brownlee Zachary; Mitra Abhishek; Noh Hyangsoon; Rojas Christina R; Li Heming; Li Shulin; Zhou Shouhao; Meng Qing HOncotarget (2017), 8 (30), 49329-49337 ISSN:.Recent advances in the field of circulating tumor cells (CTC) have shown promise in this liquid biopsy-based prognosis of patient outcome. However, not all of the circulating cells are tumor cells, as evidenced by a lack of tumor-specific markers. The current FDA standard for capturing CTCs (CellSearch) relies on an epithelial marker and cells captured via CellSearch cannot be considered to have undergone EMT. Therefore, it is difficult to ascertain the presence and relevance of any mesenchymal or EMT-like CTCs. To address this gap in technology, we recently discovered the utility of cell-surface vimentin (CSV) as a marker for detecting mesenchymal CTCs from sarcoma, breast, and colon cancer. Here we studied peripheral blood samples of 48 prostate cancer (PCA) patients including hormone sensitive and castration resistant sub-groups. Blood samples were analyzed for three different properties including our own CSV-based CTC enumeration (using 84-1 mAb against CSV), CellSearch-based epithelial CTC counts, and serum prostate-specific antigen (PSA) quantification. Our data demonstrated that in comparison with CellSearch, the CSV-based method had greater sensitivity and specificity. Further, we observed significantly greater numbers of CTCs in castration resistant patients as measured by our CSV method but not CellSearch. Our data suggests CSV-guided CTC enumeration may hold prognostic value and should be further validated as a possible measurement of PCA progression towards the deadly, androgen-independent form.
- 28Armstrong, A. J.; Marengo, M. S.; Oltean, S.; Kemeny, G.; Bitting, R. L.; Turnbull, J. D.; Herold, C. I.; Marcom, P. K.; George, D. J.; Garcia-Blanco, M. A. Circulating tumor cells from patients with advanced prostate and breast cancer display both epithelial and mesenchymal markers. Mol. Cancer Res. 2011, 9 (8), 997– 1007, DOI: 10.1158/1541-7786.MCR-10-0490Google Scholar28https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3MXhtVajtbvL&md5=78bab32e33ba992ed65beb9acbfb0acdCirculating Tumor Cells from Patients with Advanced Prostate and Breast Cancer Display Both Epithelial and Mesenchymal MarkersArmstrong, Andrew J.; Marengo, Matthew S.; Oltean, Sebastian; Kemeny, Gabor; Bitting, Rhonda L.; Turnbull, James D.; Herold, Christina I.; Marcom, Paul K.; George, Daniel J.; Garcia-Blanco, Mariano A.Molecular Cancer Research (2011), 9 (8), 997-1007CODEN: MCROC5; ISSN:1541-7786. (American Association for Cancer Research)During cancer progression, malignant cells undergo epithelial-mesenchymal transitions (EMT) and mesenchymal-epithelial transitions (MET) as part of a broad invasion and metastasis program. We previously obsd. MET events among lung metastases in a preclin. model of prostate adenocarcinoma that suggested a relationship between epithelial plasticity and metastatic spread. We thus sought to translate these findings into clin. evidence by examg. the existence of EMT in circulating tumor cells (CTC) from patients with progressive metastatic solid tumors, with a focus on men with castration-resistant prostate cancer (CRPC) and women with metastatic breast cancer. We showed that the majority (>80%) of these CTCs in patients with metastatic CRPC coexpress epithelial proteins such as epithelial cell adhesion mol. (EpCAM), cytokeratins (CK), and E-cadherin, with mesenchymal proteins including vimentin, N-cadherin and O-cadherin, and the stem cell marker CD133. Equally, we found that more than 75% of CTCs from women with metastatic breast cancer coexpress CK, vimentin, and N-cadherin. The existence and high frequency of these CTCs coexpressing epithelial, mesenchymal, and stem cell markers in patients with progressive metastases has important implications for the application and interpretation of approved methods to detect CTCs.
- 29Chen, C. L.; Mahalingam, D.; Osmulski, P.; Jadhav, R. R.; Wang, C. M.; Leach, R. J.; Chang, T. C.; Weitman, S. D.; Kumar, A. P.; Sun, L.; Gaczynska, M. E.; Thompson, I. M.; Huang, T. H. Single-cell analysis of circulating tumor cells identifies cumulative expression patterns of EMT-related genes in metastatic prostate cancer. Prostate 2013, 73 (8), 813– 26, DOI: 10.1002/pros.22625Google Scholar29https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXmslWnu74%253D&md5=13a5248089ac2b8796a6893728b27512Single-cell analysis of circulating tumor cells identifies cumulative expression patterns of EMT-related genes in metastatic prostate cancerChen, Chun-Liang; Mahalingam, Devalingam; Osmulski, Pawel; Jadhav, Rohit R.; Wang, Chiou-Miin; Leach, Robin J.; Chang, Tien-Cheng; Weitman, Steven D.; Kumar, Addanki Pratap; Sun, Lu Zhe; Gaczynska, Maria E.; Thompson, Ian M.; Huang, Tim Hui-MingProstate (Hoboken, NJ, United States) (2013), 73 (8), 813-826CODEN: PRSTDS; ISSN:0270-4137. (Wiley-Blackwell)BACKGROUND : Prostate tumors shed circulating tumor cells (CTCs) into the blood stream. Increased evidence shows that CTCs are often present in metastatic prostate cancer and can be alternative sources for disease profiling and prognostication. Here we postulate that CTCs expressing genes related to epithelial-mesenchymal transition (EMT) are strong predictors of metastatic prostate cancer. METHODS : A microfiltration system was used to trap CTCs from peripheral blood based on size selection of large epithelial-like cells without CD45 leukocyte marker. These cells individually retrieved with a micromanipulator device were assessed for cell membrane phys. properties using at. force microscopy. Addnl., 38 CTCs from eight prostate cancer patients were used to det. expression profiles of 84 EMT-related and ref. genes using a microfluidics-based PCR system. RESULTS : Increased cell elasticity and membrane smoothness were found in CTCs compared to noncancerous cells, highlighting their potential invasiveness and mobility in the peripheral circulation. Despite heterogeneous expression patterns of individual CTCs, genes that promote mesenchymal transitioning into a more malignant state, including IGF1, IGF2, EGFR, FOXP3, and TGFB3, were commonly obsd. in these cells. An addnl. subset of EMT-related genes (e.g., PTPRN2, ALDH1, ESR2, and WNT5A) were expressed in CTCs of castration-resistant cancer, but less frequently in castration-sensitive cancer. CONCLUSIONS : The study suggests that an incremental expression of EMT-related genes in CTCs is assocd. with metastatic castration-resistant cancer. Although CTCs represent a group of highly heterogeneous cells, their unique EMT-related gene signatures provide a new opportunity for personalized treatments with targeted inhibitors in advanced prostate cancer patients. Prostate 73: 813-826, 2013. © 2012 Wiley Periodicals, Inc.
- 30Mitra, A.; Mishra, L.; Li, S. EMT, CTCs and CSCs in tumor relapse and drug-resistance. Oncotarget 2015, 6 (13), 10697– 10711, DOI: 10.18632/oncotarget.4037Google Scholar30https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC2MfltVyktw%253D%253D&md5=db1d641e9116b4527b6fb733c2a40a81EMT, CTCs and CSCs in tumor relapse and drug-resistanceMitra Abhisek; Li Shulin; Mishra LopaOncotarget (2015), 6 (13), 10697-711 ISSN:.Tumor relapse and metastasis are the primary causes of poor survival rates in patients with advanced cancer despite successful resection or chemotherapeutic treatment. A primary cause of relapse and metastasis is the persistence of cancer stem cells (CSCs), which are highly resistant to chemotherapy. Although highly efficacious drugs suppressing several subpopulations of CSCs in various tissue-specific cancers are available, recurrence is still common in patients. To find more suitable therapy for relapse, the mechanisms underlying metastasis and drug-resistance associated with relapse-initiating CSCs need to be identified. Recent studies in circulating tumor cells (CTCs) of some cancer patients manifest phenotypes of both CSCs and epithelial-mesenchymal transition (EMT). These patients are unresponsive to standard chemotherapies and have low progression free survival, suggesting that EMT-positive CTCs are related to co-occur with or transform into relapse-initiating CSCs. Furthermore, EMT programming in cancer cells enables in the remodeling of extracellular matrix to break the dormancy of relapse-initiating CSCs. In this review, we extensively discuss the association of the EMT program with CTCs and CSCs to characterize a subpopulation of patients prone to relapses. Identifying the mechanisms by which EMT-transformed CTCs and CSCs initiate relapse could facilitate the development of new or enhanced personalized therapeutic regimens.
- 31Chang, L.; Graham, P. H.; Hao, J.; Ni, J.; Bucci, J.; Cozzi, P. J.; Kearsley, J. H.; Li, Y. Acquisition of epithelial-mesenchymal transition and cancer stem cell phenotypes is associated with activation of the PI3K/Akt/mTOR pathway in prostate cancer radioresistance. Cell Death Dis. 2013, 4, e875, DOI: 10.1038/cddis.2013.407Google Scholar31https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXhs1Olt7jK&md5=0f7e376b9cf9e3e5cd96544c405371e0Acquisition of epithelial-mesenchymal transition and cancer stem cell phenotypes is associated with activation of the PI3K/Akt/mTOR pathway in prostate cancer radioresistanceChang, L.; Graham, P. H.; Hao, J.; Ni, J.; Bucci, J.; Cozzi, P. J.; Kearsley, J. H.; Li, Y.Cell Death & Disease (2013), 4 (Oct.), e875CODEN: CDDEA4; ISSN:2041-4889. (Nature Publishing Group)Radioresistance is a major challenge in prostate cancer (CaP) radiotherapy (RT). In this study, we investigated the role and assocn. of epithelial-mesenchymal transition (EMT), cancer stem cells (CSCs) and the PI3K/Akt/mTOR signaling pathway in CaP radioresistance. We developed three novel CaP radioresistant (RR) cell lines (PC-3RR, DU145RR and LNCaPRR) by radiation treatment and confirmed their radioresistance using a clonogenic survival assay. Compared with untreated CaP-control cells, the CaP-RR cells had increased colony formation, invasion ability and spheroid formation capability (P<0.05). In addn., enhanced EMT/CSC phenotypes and activation of the checkpoint proteins (Chk1 and Chk2) and the PI3K/Akt/mTOR signaling pathway proteins were also found in CaP-RR cells using immunofluorescence, western blotting and quant. real-time PCR (qRT-PCR). Furthermore, combination of a dual PI3K/mTOR inhibitor (BEZ235) with RT effectively increased radiosensitivity and induced more apoptosis in CaP-RR cells, concomitantly correlated with the reduced expression of EMT/CSC markers and the PI3K/Akt/mTOR signaling pathway proteins compared with RT alone. Our findings indicate that CaP radioresistance is assocd. with EMT and enhanced CSC phenotypes via activation of the PI3K/Akt/mTOR signaling pathway, and that the combination of BEZ235 with RT is a promising modality to overcome radioresistance in the treatment of CaP. This combination approach warrants future in vivo animal study and clin. trials.
- 32Scher, H. I.; Morris, M. J.; Stadler, W. M.; Higano, C.; Basch, E.; Fizazi, K.; Antonarakis, E. S.; Beer, T. M.; Carducci, M. A.; Chi, K. N.; Corn, P. G.; de Bono, J. S.; Dreicer, R.; George, D. J.; Heath, E. I.; Hussain, M.; Kelly, W. K.; Liu, G.; Logothetis, C.; Nanus, D.; Stein, M. N.; Rathkopf, D. E.; Slovin, S. F.; Ryan, C. J.; Sartor, O.; Small, E. J.; Smith, M. R.; Sternberg, C. N.; Taplin, M. E.; Wilding, G.; Nelson, P. S.; Schwartz, L. H.; Halabi, S.; Kantoff, P. W.; Armstrong, A. J. Trial Design and Objectives for Castration-Resistant Prostate Cancer: Updated Recommendations From the Prostate Cancer Clinical Trials Working Group 3. J. Clin. Oncol. 2016, 34 (12), 1402– 18, DOI: 10.1200/JCO.2015.64.2702Google Scholar32https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC28jjt1KgsA%253D%253D&md5=ca0a8f257cabae86b5434b97079b2552Trial Design and Objectives for Castration-Resistant Prostate Cancer: Updated Recommendations From the Prostate Cancer Clinical Trials Working Group 3Scher Howard I; Morris Michael J; Stadler Walter M; Higano Celestia; Basch Ethan; Fizazi Karim; Antonarakis Emmanuel S; Beer Tomasz M; Carducci Michael A; Chi Kim N; Corn Paul G; de Bono Johann S; Dreicer Robert; George Daniel J; Heath Elisabeth I; Hussain Maha; Kelly Wm Kevin; Liu Glenn; Logothetis Christopher; Nanus David; Stein Mark N; Rathkopf Dana E; Slovin Susan F; Ryan Charles J; Sartor Oliver; Small Eric J; Smith Matthew Raymond; Sternberg Cora N; Taplin Mary-Ellen; Wilding George; Nelson Peter S; Schwartz Lawrence H; Halabi Susan; Kantoff Philip W; Armstrong Andrew JJournal of clinical oncology : official journal of the American Society of Clinical Oncology (2016), 34 (12), 1402-18 ISSN:.PURPOSE: Evolving treatments, disease phenotypes, and biology, together with a changing drug development environment, have created the need to revise castration-resistant prostate cancer (CRPC) clinical trial recommendations to succeed those from prior Prostate Cancer Clinical Trials Working Groups. METHODS: An international expert committee of prostate cancer clinical investigators (the Prostate Cancer Clinical Trials Working Group 3 [PCWG3]) was reconvened and expanded and met in 2012-2015 to formulate updated criteria on the basis of emerging trial data and validation studies of the Prostate Cancer Clinical Trials Working Group 2 recommendations. RESULTS: PCWG3 recommends that baseline patient assessment include tumor histology, detailed records of prior systemic treatments and responses, and a detailed reporting of disease subtypes based on an anatomic pattern of metastatic spread. New recommendations for trial outcome measures include the time to event end point of symptomatic skeletal events, as well as time to first metastasis and time to progression for trials in the nonmetastatic CRPC state. PCWG3 introduces the concept of no longer clinically benefiting to underscore the distinction between first evidence of progression and the clinical need to terminate or change treatment, and the importance of documenting progression in existing lesions as distinct from the development of new lesions. Serial biologic profiling using tumor samples from biopsies, blood-based diagnostics, and/or imaging is also recommended to gain insight into mechanisms of resistance and to identify predictive biomarkers of sensitivity for use in prospective trials. CONCLUSION: PCWG3 moves drug development closer to unmet needs in clinical practice by focusing on disease manifestations most likely to affect prognosis adversely for therapeutics tested in both nonmetastatic and metastatic CRPC populations. Consultation with regulatory authorities is recommended if a trial is intended to seek support for drug approval.
- 33Tucci, M.; Scagliotti, G. V.; Vignani, F. Metastatic castration-resistant prostate cancer: time for innovation. Future Oncol. 2015, 11 (1), 91– 106, DOI: 10.2217/fon.14.145Google Scholar33https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXkvFGhsA%253D%253D&md5=f891f272b109eb0d6a7523d8376ce2b7Metastatic castration-resistant prostate cancer: time for innovationTucci, Marcello; Scagliotti, Giorgio Vittorio; Vignani, FrancescaFuture Oncology (2015), 11 (1), 91-106CODEN: FOUNBN; ISSN:1479-6694. (Future Medicine Ltd.)A review. Androgen deprivation is the mainstay of advanced prostate cancer treatment. Despite initial responses, almost all patients progress to castration-resistant prostate cancer (CRPC). The understanding of the biol. of CRPC and the evidence that CRPC still remains driven by androgen receptor signaling led to the discovery of new therapeutic targets. In the last few years, large Phase III trials showed improvements in survival and outcomes and led to the approval of a CYP17 inhibitor (abiraterone), an androgen receptor antagonist (enzalutamide), the taxane cabazitaxel, an α-emitter (radium-223), the bone resorption-targeting drug denosumab and an immunotherapy (sipuleucel-T). This article describes the mol. mechanisms underlying castration resistance, discusses recent and ongoing trials and offers some insights into identifying the best sequence of new drugs.
- 34Liu, W.; Yin, B.; Wang, X.; Yu, P.; Duan, X.; Liu, C.; Wang, B.; Tao, Z. Circulating tumor cells in prostate cancer: Precision diagnosis and therapy. Oncol. Lett. 2017, 14 (2), 1223– 1232, DOI: 10.3892/ol.2017.6332Google Scholar34https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC1cfls12qtg%253D%253D&md5=535eace292cffa2d2738134434a51cf2Circulating tumor cells in prostate cancer: Precision diagnosis and therapyLiu Weiwei; Yin Binbin; Wang Xuchu; Yu Pan; Duan Xiuzhi; Liu Chunhua; Wang Ben; Tao ZhihuaOncology letters (2017), 14 (2), 1223-1232 ISSN:1792-1074.The primary cause of tumor-associated mortality in prostate cancer (PCa) remains distant metastasis. The dissemination of tumor cells from the primary tumor to distant sites through the bloodstream cannot be detected early by standard imaging methods. Circulating tumor cells (CTCs) represent an effective prognostic and predictive biomarker, which are able to monitor efficacy of adjuvant therapies, detect early development of metastases, and finally, assess therapeutic responses of advanced disease earlier than traditional diagnostic methods. In addition, since repeated tissue biopsies are invasive, costly and not always feasible, the assessment of tumor characteristics on CTCs, by a peripheral blood sample as a liquid biopsy, represents an attractive opportunity. The implementation of molecular and genomic characterization of CTCs may contribute to improve the treatment selection and thus, to move toward more precise diagnosis and therapy in PCa. The present study summarizes the current advances in CTC enrichment and detection strategies and reviews how CTCs may contribute to significant insights in the metastatic process, as well as how they may be utilized in clinical application in PCa. Although it is proposed that CTCs may offer insights into the prognosis and management of PCa, there are a number of challenges in the study of circulating tumor cells, and their clinical utility remains under investigation.
- 35De Laere, B.; Oeyen, S.; Van Oyen, P.; Ghysel, C.; Ampe, J.; Ost, P.; Demey, W.; Hoekx, L.; Schrijvers, D.; Brouwers, B.; Lybaert, W.; Everaert, E.; Van Kerckhove, P.; De Maeseneer, D.; Strijbos, M.; Bols, A.; Fransis, K.; Beije, N.; de Kruijff, I.; van Dam, V.; Brouwer, A.; van Dam, P. J.; Van den Eynden, G.; Rutten, A.; Sleijfer, S.; Vandebroek, J.; Van Laere, S.; Dirix, L. Circulating tumor cells and survival in abiraterone- and enzalutamide-treated patients with castration-resistant prostate cancer. Prostate 2018, 78 (6), 435– 445, DOI: 10.1002/pros.23488Google Scholar35https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXks1ylsb0%253D&md5=bb35fa656326b4fec6aaabc65bc3f222Circulating tumor cells and survival in abiraterone- and enzalutamide-treated patients with castration-resistant prostate cancerDe Laere, Bram; Oeyen, Steffi; Van Oyen, Peter; Ghysel, Christophe; Ampe, Jozef; Ost, Piet; Demey, Wim; Hoekx, Lucien; Schrijvers, Dirk; Brouwers, Barbara; Lybaert, Willem; Everaert, Els; Van Kerckhove, Piet; De Maeseneer, Daan; Strijbos, Michiel; Bols, Alain; Fransis, Karen; Beije, Nick; de Kruijff, Inge; van Dam, Valerie; Brouwer, Anja; van Dam, Pieter-Jan; Van den Eynden, Gert; Rutten, Annemie; Sleijfer, Stefan; Vandebroek, Jean; Van Laere, Steven; Dirix, LucProstate (Hoboken, NJ, United States) (2018), 78 (6), 435-445CODEN: PRSTDS; ISSN:0270-4137. (Wiley-Blackwell)Background : The outcome to treatment administered to patients with metastatic castration-resistant prostate cancer (mCRPC) greatly differs between individuals, underlining the need for biomarkers guiding treatment decision making. Objective : To investigate the prognostic value of circulating tumor cell (CTC) enumeration and dynamics, in the context of second-line endocrine therapies (ie, abiraterone acetate or enzalutamide), irresp. of prior systemic therapies. Design, Settings, and Participants : In a prospective, multicentre study blood samples for CTC enumeration were collected from patients with mCRPC at baseline (n = 174). In patients who responded for minimally 10-12 wk a follow-up sample was collected. Outcome Measurements and Statistical Anal. : For baseline anal., patients were stratified in <5 or ≥5 CTCs/7.5 mL, whereas for the anal. of CTC dynamics at 10-12 wk, in patients with stable, increasing or decreasing CTC counts. Progression-free survival (PFS), overall survival (OS), and PSA changes at 10-12 wk were compared between groups. Results : Patients demonstrating increasing CTCs on therapy had a shorter median PFS (4.03 vs 12.98 vs 13.67 mo, HR 3.6, 95%CI 1.9-6.8; P < 0.0001) and OS (11.2 mo vs not reached, HR 9.5, 95%CI 3.7-24; P < 0.0001), compared to patients with decreasing or stable CTCs. Multivariable Cox regression showed that prior chemotherapy (HR 4.1, 95%CI 1.9-8.9; P = 0.0003), a high baseline CTC count (HR 1.5, 95%CI 1.2-1.9; P = 0.002) and increasing CTCs at follow-up (HR 3.3, 95%CI 1.4-7.6; P = 0.005) were independent predictors of worse PFS. Previous chemotherapy (HR 7, 95%CI 1.9-25; P = 0.003), high baseline CTC counts (HR 2.2, 95%CI 1.4-3.7; P = 0.002) and increasing CTCs during therapy (HR 4.6, 95%CI 1.4-15; P = 0.01) were independently assocd. with shorter OS. ≥30% and ≥50% PSA responses less frequently occurred in patients with CTC inclines at 10-12 wk on therapy (χ2 test: P < 0.01). Conclusions : CTC dynamics during therapy are assocd. with PSA response and provide independent clin. prognostication over PSA declines. Hence the study demonstrates the pharmacodynamic properties of CTCs.
- 36Scher, H. I.; Fizazi, K.; Saad, F.; Taplin, M. E.; Sternberg, C. N.; Miller, K.; de Wit, R.; Mulders, P.; Chi, K. N.; Shore, N. D.; Armstrong, A. J.; Flaig, T. W.; Flechon, A.; Mainwaring, P.; Fleming, M.; Hainsworth, J. D.; Hirmand, M.; Selby, B.; Seely, L.; de Bono, J. S. Increased survival with enzalutamide in prostate cancer after chemotherapy. N. Engl. J. Med. 2012, 367 (13), 1187– 97, DOI: 10.1056/NEJMoa1207506Google Scholar36https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC38XhsVKhsLvK&md5=02fcfdf5b35766d91a2ca023558acd74Increased survival with enzalutamide in prostate cancer after chemotherapyScher, Howard I.; Fizazi, Karim; Saad, Fred; Taplin, Mary-Ellen; Sternberg, Cora N.; Miller, Kurt; de Wit, Ronald; Mulders, Peter; Chi, Kim N.; Shore, Neal D.; Armstrong, Andrew J.; Flaig, Thomas W.; Flechon, Aude; Mainwaring, Paul; Fleming, Mark; Hainsworth, John D.; Hirmand, Mohammad; Selby, Bryan; Seely, Lynn; de Bono, Johann S.New England Journal of Medicine (2012), 367 (13), 1187-1197CODEN: NEJMAG; ISSN:0028-4793. (Massachusetts Medical Society)Background: Enzalutamide (formerly called MDV3100) targets multiple steps in the androgen-receptor-signaling pathway, the major driver of prostate-cancer growth. We aimed to evaluate whether enzalutamide prolongs survival in men with castration-resistant prostate cancer after chemotherapy. Methods: In our phase 3, double-blind, placebo-controlled trial, we stratified 1199 men with castration-resistant prostate cancer after chemotherapy according to the Eastern Cooperative Oncol. Group performance-status score and pain intensity. We randomly assigned them, in a 2:1 ratio, to receive oral enzalutamide at a dose of 160 mg per day (800 patients) or placebo (399 patients). The primary end point was overall survival. Results: The study was stopped after a planned interim anal. at the time of 520 deaths. The median overall survival was 18.4 mo (95% confidence interval [CI], 17.3 to not yet reached) in the enzalutamide group vs. 13.6 mo (95% CI, 11.3 to 15.8) in the placebo group (hazard ratio for death in the enzalutamide group, 0.63; 95% CI, 0.53 to 0.75; P<0.001). The superiority of enzalutamide over placebo was shown with respect to all secondary end points: the proportion of patients with a redn. in the prostate-specific antigen (PSA) level by 50% or more (54% vs. 2%, P<0.001), the soft-tissue response rate (29% vs. 4%, P<0.001), the quality-of-life response rate (43% vs. 18%, P<0.001), the time to PSA progression (8.3 vs. 3.0 mo; hazard ratio, 0.25; P<0.001), radiog. progression-free survival (8.3 vs. 2.9 mo; hazard ratio, 0.40; P<0.001), and the time to the first skeletal-related event (16.7 vs. 13.3 mo; hazard ratio, 0.69; P<0.001). Rates of fatigue, diarrhea, and hot flashes were higher in the enzalutamide group. Seizures were reported in five patients (0.6%) receiving enzalutamide. Conclusions: Enzalutamide significantly prolonged the survival of men with metastatic castration-resistant prostate cancer after chemotherapy.
- 37Loriot, Y.; Bianchini, D.; Ileana, E.; Sandhu, S.; Patrikidou, A.; Pezaro, C.; Albiges, L.; Attard, G.; Fizazi, K.; De Bono, J. S.; Massard, C. Antitumour activity of abiraterone acetate against metastatic castration-resistant prostate cancer progressing after docetaxel and enzalutamide (MDV3100). Ann. Oncol. 2013, 24 (7), 1807– 12, DOI: 10.1093/annonc/mdt136Google Scholar37https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC3srktF2muw%253D%253D&md5=c9a3e24cb46d2f5d99e1b52b3eed39a7Antitumour activity of abiraterone acetate against metastatic castration-resistant prostate cancer progressing after docetaxel and enzalutamide (MDV3100)Loriot Y; Bianchini D; Ileana E; Sandhu S; Patrikidou A; Pezaro C; Albiges L; Attard G; Fizazi K; De Bono J S; Massard CAnnals of oncology : official journal of the European Society for Medical Oncology (2013), 24 (7), 1807-12 ISSN:.BACKGROUND: Androgen receptor (AR) signalling remains critically important in metastatic castration-resistant prostate cancer (mCRPC) as confirmed by recent phase III trials, showing a survival advantage for abiraterone acetate and enzalutamide (MDV3100). The antitumour activity of abiraterone and prednisolone in patients pre-treated with enzalutamide is as yet unknown. PATIENTS AND METHODS: We investigated the antitumour activity of abiraterone and prednisolone in patients with mCRPC who had progressed following treatment with docetaxel (Taxotere) and enzalutamide. Clinical data were retrospectively analysed for prostate-specific antigen (PSA) and RECIST responses, clinical benefit and survival. RESULTS: Thirty-eight patients were included in the analysis. The median age was 71 years (range 52-84); metastatic sites included bone disease in 37 patients (97%), lymph nodes in 15 patients (39%) and visceral disease in 10 patients (26%). Abiraterone was well tolerated. Three patients (8%) attained a PSA response, defined as ≥50% decline in PSA confirmed after ≥4 weeks, while seven patients (18%) had a ≥30% PSA decline. The median progression-free survival (PFS) was 2.7 months (95% CI 2.3-4.1). Of the 12 patients assessable radiologically, only 1 (8%) attained a confirmed partial response. CONCLUSION: Abiraterone and prednisolone have modest antitumour activities in patients with mCRPC pretreated with docetaxel and enzalutamide.
- 38Brand, L. J.; Dehm, S. M. Androgen receptor gene rearrangements: new perspectives on prostate cancer progression. Curr. Drug Targets 2013, 14 (4), 441– 449, DOI: 10.2174/1389450111314040005Google Scholar38https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXotlyrsLs%253D&md5=e5d8a285f32674aaf05282a17f799ec7Androgen receptor gene rearrangements: new perspectives on prostate cancer progressionBrand, Lucas J.; Dehm, Scott M.Current Drug Targets (2013), 14 (4), 441-449CODEN: CDTUAU; ISSN:1389-4501. (Bentham Science Publishers Ltd.)A review. The androgen receptor (AR) is a master regulator transcription factor in normal and cancerous prostate cells. Canonical AR activation requires binding of androgen ligand to the AR ligand binding domain, translocation to the nucleus, and transcriptional activation of AR target genes. This regulatory axis is targeted for systemic therapy of advanced prostate cancer. However, a new paradigm for AR activation in castration-resistant prostate cancer (CRPC) has emerged wherein alternative splicing of AR mRNA promotes synthesis of constitutively active AR variants that lack the AR ligand binding domain (LBD). Recent work has indicated that structural alteration of the AR gene locus represents a key mechanism by which alterations in AR mRNA splicing arise. In this review, we examine the role of truncated AR variants (ARVs) and their corresponding genomic origins in models of prostate cancer progression, as well as the challenges they pose to the current std. of prostate cancer therapies targeting the AR ligand binding domain. Since ARVs lack the COOH-terminal LBD, the genesis of these AR gene rearrangements and their resulting ARVs provides strong rationale for the pursuit of new avenues of therapeutic intervention targeted at the AR NH2-terminal domain. We further suggest that genomic events leading to ARV expression could act as novel biomarkers of disease progression that may guide the optimal use of current and next-generation AR-targeted therapy.
- 39van der Steen, T.; Tindall, D. J.; Huang, H. Posttranslational modification of the androgen receptor in prostate cancer. Int. J. Mol. Sci. 2013, 14 (7), 14833– 59, DOI: 10.3390/ijms140714833Google Scholar39https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC3sXht1OntLzK&md5=f72323de1ac843834793fbf4d22c4a87Posttranslational modification of the androgen receptor in prostate cancervan der Steen, Travis; Tindall, Donald J.; Huang, HaojieInternational Journal of Molecular Sciences (2013), 14 (7), 14833-14859, 27 pp.CODEN: IJMCFK; ISSN:1422-0067. (MDPI AG)A review. The androgen receptor (AR) is important in the development of the prostate by regulating transcription, cellular proliferation, and apoptosis. AR undergoes posttranslational modifications that alter its transcription activity, translocation to the nucleus and stability. The posttranslational modifications that regulate these events are of utmost importance to understand the functional role of AR and its activity. The majority of these modifications occur in the activation function-1 (AF1) region of the AR, which contains the transcriptional activation unit 1 (TAU1) and 5 (TAU5). Identification of the modifications that occur to these regions may increase our understanding of AR activation in prostate cancer and the role of AR in the progression from androgen-dependent to castration-resistant prostate cancer (CRPC). Most of the posttranslational modifications identified to date have been detd. using the full-length AR in androgen dependent cells. Further investigations into the role of posttranslational modifications in androgen-independent activation of full-length AR and constitutively active splicing variants are warranted, findings from which may provide new therapeutic options for CRPC.
- 40Mohamadi, R. M.; Besant, J. D.; Mepham, A.; Green, B.; Mahmoudian, L.; Gibbs, T.; Ivanov, I.; Malvea, A.; Stojcic, J.; Allan, A. L.; Lowes, L. E.; Sargent, E. H.; Nam, R. K.; Kelley, S. O. Nanoparticle-mediated binning and profiling of heterogeneous circulating tumor cell subpopulations. Angew. Chem., Int. Ed. 2015, 54 (1), 139– 43, DOI: 10.1002/anie.201409376Google Scholar40https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhvVKlsLjI&md5=668e003fea2719f7bfbc02172f1a7150Nanoparticle-Mediated Binning and Profiling of Heterogeneous Circulating Tumor Cell SubpopulationsMohamadi, Reza M.; Besant, Justin D.; Mepham, Adam; Green, Brenda; Mahmoudian, Laili; Gibbs, Thaddeus; Ivanov, Ivaylo; Malvea, Anahita; Stojcic, Jessica; Allan, Alison L.; Lowes, Lori E.; Sargent, Edward H.; Nam, Robert K.; Kelley, Shana O.Angewandte Chemie, International Edition (2015), 54 (1), 139-143CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)The anal. of circulating tumor cells (CTCs) is an important capability that may lead to new approaches for cancer management. CTC capture devices developed to date isolate a bulk population of CTCs and do not differentiate subpopulations that may have varying phenotypes with different levels of clin. relevance. Here, the authors present a new device for CTC spatial sorting and profiling that sequesters blood-borne tumor cells with different phenotypes into discrete spatial bins. Validation data are presented showing that cancer cell lines with varying surface expression generate different binning profiles within the device. Working with patient blood samples, the authors obtain profiles that elucidate the heterogeneity of CTC populations present in cancer patients and also report on the status of CTCs within the epithelial-to-mesenchymal transition (EMT).
- 41Green, B. J.; Kermanshah, L.; Labib, M.; Ahmed, S. U.; Silva, P. N.; Mahmoudian, L.; Chang, I. H.; Mohamadi, R. M.; Rocheleau, J. V.; Kelley, S. O. Isolation of Phenotypically Distinct Cancer Cells Using Nanoparticle-Mediated Sorting. ACS Appl. Mater. Interfaces 2017, 9 (24), 20435– 20443, DOI: 10.1021/acsami.7b05253Google Scholar41https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2sXosVOktbw%253D&md5=869f093a722ed0245c2518483762ca0cIsolation of Phenotypically Distinct Cancer Cells Using Nanoparticle-Mediated SortingGreen, Brenda J.; Kermanshah, Leyla; Labib, Mahmoud; Ahmed, Sharif U.; Silva, Pamuditha N.; Mahmoudian, Laili; Chang, I-Hsin; Mohamadi, Reza M.; Rocheleau, Jonathan V.; Kelley, Shana O.ACS Applied Materials & Interfaces (2017), 9 (24), 20435-20443CODEN: AAMICK; ISSN:1944-8244. (American Chemical Society)Isolating subpopulations of heterogeneous cancer cells is an important capability for the meaningful characterization of circulating tumor cells at different stages of tumor progression and during the epithelial-to-mesenchymal transition. Here, the authors present a microfluidic device that can sep. phenotypically distinct subpopulations of cancer cells. Magnetic nanoparticles coated with antibodies against the epithelial cell adhesion mol. (EpCAM) were used to sep. breast cancer cells in the microfluidic platform. Cells are sorted into different zones on the basis of the levels of EpCAM expression, which enables the detection of cells that are losing epithelial character and becoming more mesenchymal. The phenotypic properties of the isolated cells with low and high EpCAM are then assessed using matrix-coated surfaces for collagen uptake anal. and an NAD(P)H assay that assesses metabolic activity. Low-EpCAM expressing cells have higher collagen uptake and higher folate-induced NAD(P)H responses compared to those of high-EpCAM expressing cells. In addn., the authors tested SKBR3 cancer cells undergoing chem. induced hypoxia. The induced cells have reduced expression of EpCAM, and these cells have higher collagen uptake and NAD(P)H metab. relative to noninduced cells. Nanoparticle-mediated binning facilitates the isolation of functionally distinct cell subpopulations and allows surface marker expression to be assocd. with invasiveness, including collagen uptake and metabolic activity.
- 42Poudineh, M.; Labib, M.; Ahmed, S.; Nguyen, L. N.; Kermanshah, L.; Mohamadi, R. M.; Sargent, E. H.; Kelley, S. O. Profiling Functional and Biochemical Phenotypes of Circulating Tumor Cells Using a Two-Dimensional Sorting Device. Angew. Chem., Int. Ed. 2017, 56 (1), 163– 168, DOI: 10.1002/anie.201608983Google Scholar42https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC28XhvFGqurbJ&md5=6b03fdf7980b9b6a31e6bd1334a4f1d5Profiling Functional and Biochemical Phenotypes of Circulating Tumor Cells Using a Two-Dimensional Sorting DevicePoudineh, Mahla; Labib, Mahmoud; Ahmed, Sharif; Nguyen, L. N. Matthew; Kermanshah, Leyla; Mohamadi, Reza M.; Sargent, Edward H.; Kelley, Shana O.Angewandte Chemie, International Edition (2017), 56 (1), 163-168CODEN: ACIEF5; ISSN:1433-7851. (Wiley-VCH Verlag GmbH & Co. KGaA)During cancer progression, tumors shed circulating tumor cells (CTCs) into the bloodstream. CTCs that originate from the same primary tumor can have heterogeneous phenotypes and, while some CTCs possess benign properties, others have high metastatic potential. Deconstructing the heterogeneity of CTCs is challenging and new methods are needed that can sort small nos. of cancer cells according to their phenotypic properties. Here the authors describe a new microfluidic approach that profiles, along two independent phenotypic axes, the behavior of heterogeneous cell subpopulations. Cancer cells are first profiled according to expression of a surface marker using a nanoparticle-enabled approach. Along the second dimension, these subsets are further sepd. into subpopulations corresponding to migration profiles generated in response to a chemotactic agent. The authors deploy this new technique and find a strong correlation between the surface expression and migration potential of CTCs present in blood from mice with xenografted tumors. This system provides an important new means to characterize functional diversity in circulating tumor cells.
- 43Xu, Y.; Zhao, H.; Hou, J. Correlation between overexpression of EpCAM in prostate tissues and genesis of androgen-dependent prostate cancer. Tumor Biol. 2014, 35 (7), 6695– 700, DOI: 10.1007/s13277-014-1892-2Google Scholar43https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXht1egu77F&md5=9f51be7711935b9164ac60da6b4d4980Correlation between overexpression of EpCAM in prostate tissues and genesis of androgen-dependent prostate cancerXu, Yuan; Zhao, Hu; Hou, JianquanTumor Biology (2014), 35 (7), 6695-6700CODEN: TUMBEA; ISSN:1010-4283. (Springer)The objective of this study was to investigate the role of epithelial cell adhesion mol. (EpCAM) in the genesis and the progress of prostate cancer, esp. of castration-resistant prostate cancer. Protein expression of EpCAM in ten pairs of prostate cancer tissues and normal adjacent tissues, plus three cell lines, was examd. Short hairpin RNA (shRNA) interference technique was employed to silence the expression of EpCAM in prostate cancer cell LNCaP and construct a stable transfected cell line. In vitro assay was conducted to analyze the effect of EpCAM expression on the expressions of Androgen receptor (AR), Prostate specific antigen (PSA), and cellular proliferation and invasion. EpCAM was found significantly expressed higher in prostate cancer tissues than in normal adjacent tissues. In three cell lines (DU-145, PC-3, and LNCaP), the expression of EpCAM in LNCaP, androgen-dependent prostate cancer cells, was significantly higher than that in the other two. As EpCAM was silenced in LNCaP, the expression levels of AR and PSA obviously descended, and cellular abilities of proliferation and invasion were obviously inhibited. The overexpression of EpCAM has correlation with the genesis of prostate cancer, esp. androgen-dependent prostate cancer. As the expression of AR is facilitated, prostate cancer cells' abilities to proliferate and invade are consequently enhanced.
- 44Wilbaux, M.; Tod, M.; De Bono, J.; Lorente, D.; Mateo, J.; Freyer, G.; You, B.; Henin, E. A Joint Model for the Kinetics of CTC Count and PSA Concentration During Treatment in Metastatic Castration-Resistant Prostate Cancer. CPT: Pharmacometrics Syst. Pharmacol. 2015, 4 (5), 277– 85, DOI: 10.1002/psp4.34Google Scholar44https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXnsVWrtrw%253D&md5=1094f9d797a067a8607f301896c25f90A Joint Model for the Kinetics of CTC Count and PSA Concentration During Treatment in Metastatic Castration-Resistant Prostate CancerWilbaux, M.; Tod, M.; De Bono, J.; Lorente, D.; Mateo, J.; Freyer, G.; You, B.; Henin, E.CPT: Pharmacometrics & Systems Pharmacology (2015), 4 (5), 277-285CODEN: CPSPBR; ISSN:2163-8306. (Wiley-Blackwell)Assessment of treatment efficacy in metastatic castration-resistant prostate cancer (mCRPC) is limited by frequent nonmeasurable bone metastases. The count of circulating tumor cells (CTCs) is a promising surrogate marker that may replace the widely used prostate-specific antigen (PSA). The purpose of this study was to quantify the dynamic relationships between the longitudinal kinetics of these markers during treatment in patients with mCRPC. Data from 223 patients with mCRPC treated by chemotherapy and/or hormonotherapy were analyzed for up to 6 mo of treatment. A semimechanistic model was built, combining the following several pharmacometric advanced features: (1) Kinetic-Pharmacodynamic (K-PD) compartments for treatments (chemotherapy and hormonotherapy); (2) a latent variable linking both marker kinetics; (3) modeling of CTC kinetics with a cell lifespan model; and (4) a neg. binomial distribution for the CTC random sampling. Linked with survival, this model would potentially be useful for predicting treatment efficacy during drug development or for therapeutic adjustment in treated patients.
- 45Kermanshah, L.; Poudineh, M.; Ahmed, S.; Nguyen, L. N. M.; Srikant, S.; Makonnen, R.; Pena Cantu, F.; Corrigan, M.; Kelley, S. O. Dynamic CTC phenotypes in metastatic prostate cancer models visualized using magnetic ranking cytometry. Lab Chip 2018, 18 (14), 2055– 2064, DOI: 10.1039/C8LC00310FGoogle Scholar45https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXhtFeiurvN&md5=452584c4e071201252af5367831a3fcbDynamic CTC phenotypes in metastatic prostate cancer models visualized using magnetic ranking cytometryKermanshah, Leyla; Poudineh, Mahla; Ahmed, Sharif; Nguyen, L. N. Matthew; Srikant, Sanjana; Makonnen, Rhema; Pena Cantu, Fernando; Corrigan, Michael; Kelley, Shana O.Lab on a Chip (2018), 18 (14), 2055-2064CODEN: LCAHAM; ISSN:1473-0189. (Royal Society of Chemistry)Tumors can shed thousands of cells into the circulation daily. These circulating tumor cells (CTCs) are heterogeneous, and their phenotypes change dynamically. Real-time monitoring of CTC phenotypes is crucial to elucidate the role of CTCs in the metastatic cascade. Here, we monitor phenotypic changes in CTCs in mice xenografted with tumors with varying aggressiveness during cancer progression and a course of chemotherapy to study the metastatic potential of CTCs and changes in the properties of these cells in response to treatment. A new device that enables magnetic ranking cytometry (MagRC) is employed to profile the phenotypic properties of CTCs. Overall, CTCs from metastatic xenografts in mice display dynamic and heterogeneous profiles while non-metastatic models had static profiles. Decreased heterogeneity followed by a redn. in metastasis incidence was obsd. after a course of chemotherapy administered to highly metastatic xenografts. Phenotypic profiling of CTCs could be employed to monitor disease progression and predict therapeutic responses.
- 46Muhanna, N.; Mepham, A.; Mohamadi, R. M.; Chan, H.; Khan, T.; Akens, M.; Besant, J. D.; Irish, J.; Kelley, S. O. Nanoparticle-based sorting of circulating tumor cells by epithelial antigen expression during disease progression in an animal model. Nanomedicine 2015, 11 (7), 1613– 20, DOI: 10.1016/j.nano.2015.04.017Google Scholar46https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXoslCmt7w%253D&md5=613449490a485edcfcbf3df0db75e004Nanoparticle-based sorting of circulating tumor cells by epithelial antigen expression during disease progression in an animal modelMuhanna, Nidal; Mepham, Adam; Mohamadi, Reza M.; Chan, Harley; Khan, Tahsin; Akens, Margarete; Besant, Justin D.; Irish, Jonathan; Kelley, Shana O.Nanomedicine (New York, NY, United States) (2015), 11 (7), 1613-1620CODEN: NANOBF; ISSN:1549-9634. (Elsevier)Circulating tumor cells (CTCs) can be used as markers for the detection, characterization, and targeted therapeutic management of cancer. We recently developed a nanoparticle-mediated approach for capture and sorting of CTCs based on their specific epithelial phenotype. In the current study, we investigate the phenotypic transition of tumor cells in an animal model and show the correlation of this transition with tumor progression. VX2 tumor cells were injected into rabbits, and CTCs were evaluated during tumor progression and correlated with computerized tomog. (CT) measurements of tumor vol. The results showed a dramatic increase of CTCs during the four weeks of tumor growth. Following resection, CTC levels dropped but then rebounded, likely due to lymph node metastases. Addnl., CTCs showed a marked loss of the epithelial cell adhesion mol. (EpCAM) relative to precursor cells. In conclusion, the device accurately traces disease progression and CTC phenotypic shift in an animal model.
- 47Scher, H. I.; Lu, D.; Schreiber, N. A.; Louw, J.; Graf, R. P.; Vargas, H. A.; Johnson, A.; Jendrisak, A.; Bambury, R.; Danila, D.; McLaughlin, B.; Wahl, J.; Greene, S. B.; Heller, G.; Marrinucci, D.; Fleisher, M.; 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 Oncol 2016, 2 (11), 1441– 1449, DOI: 10.1001/jamaoncol.2016.1828Google Scholar47https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC2s%252Fms1Kitw%253D%253D&md5=92be71b45282c41a743d5afc03813167Association of AR-V7 on Circulating Tumor Cells as a Treatment-Specific Biomarker With Outcomes and Survival in Castration-Resistant Prostate CancerScher Howard I; Danila Daniel; Lu David; Louw Jessica; Graf Ryon P; Johnson Ann; Jendrisak Adam; Wahl Justin; Greene Stephanie B; Marrinucci Dena; Dittamore Ryan; Schreiber Nicole A; McLaughlin Brigit; Vargas Hebert A; Bambury Richard; Heller Glenn; Fleisher MartinJAMA oncology (2016), 2 (11), 1441-1449 ISSN:.IMPORTANCE: A critical decision in the management of metastatic castration-resistant prostate cancer (mCRPC) is when to administer an androgen receptor signaling (ARS) inhibitor or a taxane. OBJECTIVE: To determine if pretherapy nuclear androgen-receptor splice variant 7 (AR-V7) protein expression and localization on circulating tumor cells (CTCs) is a treatment-specific marker for response and outcomes between ARS inhibitors and taxanes. DESIGN, SETTING, AND PARTICIPANTS: For this cross-sectional cohort study at Memorial Sloan Kettering Cancer Center, 265 men with progressive mCRPC undergoing a change in treatment were considered; 86 were excluded because they were not initiating ARS or taxane therapy; and 18 were excluded for processing time constraints, leaving 161 patients for analysis. Between December 2012 and March 2015, blood was collected and processed from patients with progressive mCRPC immediately prior to new line of systemic therapy. Patients were followed up to 3 years. MAIN OUTCOMES AND MEASURES: Prostate-specific antigen (PSA) response, time receiving therapy, radiographic progression-free survival (rPFS), and overall survival (OS). RESULTS: Overall, of 193 prospectively collected blood samples from 161 men with mCRPC, 191 were evaluable (128 pre-ARS inhibitor and 63 pretaxane). AR-V7-positive CTCs were found in 34 samples (18%), including 3% of first-line, 18% of second-line, and 31% of third- or greater line samples. Patients whose samples had AR-V7-positive CTCs before ARS inhibition had resistant posttherapy PSA changes (PTPC), shorter rPFS, shorter time on therapy, and shorter OS than those without AR-V7-positive CTCs. Overall, resistant PTPC were seen in 65 of 112 samples (58%) without detectable AR-V7-positive CTCs prior to ARS inhibition. There were statistically significant differences in OS but not in PTPC, time on therapy, or rPFS for patients with or without pretherapy AR-V7-positive CTCs treated with a taxane. A multivariable model adjusting for baseline factors associated with survival showed superior OS with taxanes relative to ARS inhibitors when AR-V7-positive CTCs were detected pretherapy (hazard ratio, 0.24; 95% CI, 0.10-0.57; P = .035). CONCLUSIONS AND RELEVANCE: The results validate CTC nuclear expression of AR-V7 protein in men with mCRPC as a treatment-specific biomarker that is associated with superior survival on taxane therapy over ARS-directed therapy in a clinical practice setting. Continued examination of this biomarker in prospective studies will further aid clinical utility.
- 48Antonarakis, E. S.; Lu, C.; Wang, H.; Luber, B.; Nakazawa, M.; Roeser, J. C.; Chen, Y.; Mohammad, T. A.; Chen, Y.; Fedor, H. L.; Lotan, T. L.; Zheng, Q.; De Marzo, A. M.; Isaacs, J. T.; Isaacs, W. B.; Nadal, R.; Paller, C. J.; Denmeade, S. R.; Carducci, M. A.; Eisenberger, M. A.; Luo, J. AR-V7 and resistance to enzalutamide and abiraterone in prostate cancer. N. Engl. J. Med. 2014, 371 (11), 1028– 38, DOI: 10.1056/NEJMoa1315815Google Scholar48https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2cXhs1Olt73L&md5=c9e06bd306abae0616d963ed82c443eeAR-V7 and resistance to enzalutamide and abiraterone in prostate cancerAntonarakis, Emmanuel S.; Lu, Changxue; Wang, Hao; Luber, Brandon; Nakazawa, Mary; Roeser, Jeffrey C.; Chen, Yan; Mohammad, Tabrez A.; Chen, Yidong; Fedor, Helen L.; Lotan, Tamara L.; Zheng, Qizhi; De Marzo, Angelo M.; Isaacs, John T.; Isaacs, William B.; Nadal, Rosa; Paller, Channing J.; Denmeade, Samuel R.; Carducci, Michael A.; Eisenberger, Mario A.; Luo, JunNew England Journal of Medicine (2014), 371 (11), 1028-1038, 11 pp.CODEN: NEJMAG; ISSN:1533-4406. (Massachusetts Medical Society)Background The androgen-receptor isoform encoded by splice variant 7 lacks the ligand-binding domain, which is the target of enzalutamide and abiraterone, but remains constitutively active as a transcription factor. We hypothesized that detection of androgen-receptor splice variant 7 mRNA (AR-V7) in circulating tumor cells from men with advanced prostate cancer would be assocd. with resistance to enzalutamide and abiraterone. Methods We used a quant. reverse-transcriptase-polymerase-chain-reaction assay to evaluate AR-V7 in circulating tumor cells from prospectively enrolled patients with metastatic castration-resistant prostate cancer who were initiating treatment with either enzalutamide or abiraterone. We examd. assocns. between AR-V7 status (pos. vs. neg.) and prostate-specific antigen (PSA) response rates (the primary end point), freedom from PSA progression (PSA progression-free survival), clin. or radiog. progression-free survival, and overall survival. Results A total of 31 enzalutamide-treated patients and 31 abiraterone-treated patients were enrolled, of whom 39% and 19%, resp., had detectable AR-V7 in circulating tumor cells. Among men receiving enzalutamide, AR-V7-pos. patients had lower PSA response rates than AR-V7-neg. patients (0% vs. 53%, P = 0.004) and shorter PSA progression-free survival (median, 1.4 mo vs. 6.0 mo; P<0.001), clin. or radiog. progression-free survival (median, 2.1 mo vs. 6.1 mo; P<0.001), and overall survival (median, 5.5 mo vs. not reached; P = 0.002). Similarly, among men receiving abiraterone, AR-V7-pos. patients had lower PSA response rates than AR-V7-neg. patients (0% vs. 68%, P = 0.004) and shorter PSA progression-free survival (median, 1.3 mo vs. not reached; P<0.001), clin. or radiog. progression-free survival (median, 2.3 mo vs. not reached; P<0.001), and overall survival (median, 10.6 mo vs. not reached, P = 0.006). The assocn. between AR-V7 detection and therapeutic resistance was maintained after adjustment for expression of full-length androgen receptor mRNA. Conclusions Detection of AR-V7 in circulating tumor cells from patients with castration-resistant prostate cancer may be assocd. with resistance to enzalutamide and abiraterone. These findings require large-scale prospective validation.
- 49Antonarakis, E. S.; Lu, C.; Luber, B.; Wang, H.; Chen, Y.; Nakazawa, M.; Nadal, R.; Paller, C. J.; Denmeade, S. R.; Carducci, M. A.; Eisenberger, M. A.; Luo, J. Androgen Receptor Splice Variant 7 and Efficacy of Taxane Chemotherapy in Patients With Metastatic Castration-Resistant Prostate Cancer. JAMA Oncol 2015, 1 (5), 582– 91, DOI: 10.1001/jamaoncol.2015.1341Google Scholar49https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A280%3ADC%252BC28%252FjsVGquw%253D%253D&md5=445419defe501788b64fea6426b58231Androgen Receptor Splice Variant 7 and Efficacy of Taxane Chemotherapy in Patients With Metastatic Castration-Resistant Prostate CancerAntonarakis Emmanuel S; Luber Brandon; Wang Hao; Nadal Rosa; Paller Channing J; Denmeade Samuel R; Carducci Michael A; Eisenberger Mario A; Lu Changxue; Chen Yan; Nakazawa Mary; Luo JunJAMA oncology (2015), 1 (5), 582-91 ISSN:.IMPORTANCE: We previously showed that detection of androgen receptor splice variant 7 (AR-V7) in circulating tumor cells (CTCs) from men with castration-resistant prostate cancer (CRPC) was associated with primary resistance to enzalutamide and abiraterone therapy, but the relevance of AR-V7 status in the context of chemotherapy is unknown. OBJECTIVE: To investigate whether AR-V7-positive patients would retain sensitivity to taxane chemotherapy and whether AR-V7 status would have a differential impact on taxane-treated men compared with enzalutamide- or abiraterone-treated men. DESIGN, SETTING, AND PARTICIPANTS: We examined CTCs for AR-V7 mRNA using a reverse-transcription polymerase chain reaction assay. From January 2013 to July 2014, we prospectively enrolled patients with metastatic CRPC initiating taxane chemotherapy (docetaxel or cabazitaxel) at a single academic institution (Johns Hopkins). Our prespecified statistical plan required a sample size of 36 taxane-treated men. MAIN OUTCOMES AND MEASURES: We evaluated associations between AR-V7 status and prostate-specific antigen (PSA) response rates, PSA progression-free survival (PSA PFS), and clinical and/or radiographic progression-free survival (PFS). After incorporating updated data from our prior study of 62 patients treated with enzalutamide or abiraterone, we also investigated the interaction between AR-V7 status (positive or negative) and treatment type (taxane vs enzalutamide or abiraterone). RESULTS: Of 37 taxane-treated patients enrolled, 17 (46%) had detectable AR-V7 in CTCs. Prostate-specific antigen responses were achieved in both AR-V7-positive and AR-V7-negative men (41% vs 65%; P = .19). Similarly, PSA PFS (hazard ratio [HR], 1.7, 95% CI, 0.6-5.0; P = .32) and PFS (HR, 2.7, 95% CI, 0.8-8.8; P = .11) were comparable in AR-V7-positive and AR-V7-negative patients. A significant interaction was observed between AR-V7 status and treatment type (P < .001). Clinical outcomes were superior with taxanes compared with enzalutamide or abiraterone therapy in AR-V7-positive men, whereas outcomes did not differ by treatment type in AR-V7-negative men. In AR-V7-positive patients, PSA responses were higher in taxane-treated vs enzalutamide- or abiraterone-treated men (41% vs 0%; P < .001), and PSA PFS and PFS were significantly longer in taxane-treated men (HR, 0.19 [95% CI, 0.07-0.52] for PSA PFS, P = .001; HR, 0.21 [95% CI, 0.07-0.59] for PFS, P = .003). CONCLUSIONS AND RELEVANCE: Detection of AR-V7 in CTCs from men with metastatic CRPC is not associated with primary resistance to taxane chemotherapy. In AR-V7-positive men, taxanes appear to be more efficacious than enzalutamide or abiraterone therapy, whereas in AR-V7-negative men, taxanes and enzalutamide or abiraterone may have comparable efficacy. Circulating tumor cell-based AR-V7 detection may serve as a treatment selection biomarker in CRPC.
- 50Crespo, M.; van Dalum, G.; Ferraldeschi, R.; Zafeiriou, Z.; Sideris, S.; Lorente, D.; Bianchini, D.; Rodrigues, D. N.; Riisnaes, R.; Miranda, S.; Figueiredo, I.; Flohr, P.; Nowakowska, K.; de Bono, J. S.; Terstappen, L. W.; Attard, G. Androgen receptor expression in circulating tumour cells from castration-resistant prostate cancer patients treated with novel endocrine agents. Br. J. Cancer 2015, 112 (7), 1166– 74, DOI: 10.1038/bjc.2015.63Google Scholar50https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC2MXls1artrk%253D&md5=95c7a159574eb10d17eec49acbd8c7d8Androgen receptor expression in circulating tumour cells from castration-resistant prostate cancer patients treated with novel endocrine agentsCrespo, M.; van Dalum, G.; Ferraldeschi, R.; Zafeiriou, Z.; Sideris, S.; Lorente, D.; Bianchini, D.; Rodrigues, D. N.; Riisnaes, R.; Miranda, S.; Figueiredo, I.; Flohr, P.; Nowakowska, K.; de Bono, J. S.; Terstappen, L. W. M. M.; Attard, G.British Journal of Cancer (2015), 112 (7), 1166-1174CODEN: BJCAAI; ISSN:0007-0920. (Nature Publishing Group)Background: Abiraterone and enzalutamide are novel endocrine treatments that abrogate androgen receptor (AR) signalling in castration-resistant prostate cancer (CRPC). Here, we developed a circulating tumor cells (CTCs)-based assay to evaluate AR expression in real-time in CRPC and investigated nuclear AR expression in CTCs in patients treated with enzalutamide and abiraterone. Methods: CTCs were captured and characterized using the CellSearch system. An automated algorithm to identify CTCs and quantify AR expression was employed. The primary aim was to evaluate the assocn. between CTC AR expression and prior treatment with abiraterone or enzalutamide. Results: AR expression in CTCs was evaluated in 94 samples from 48 metastatic CRPC patients. We obsd. large intra-patient heterogeneity of AR expression in CTCs. Prior exposure to abiraterone or enzalutamide was not assocd. with a change in CTCs AR expression (median intensity and distribution of AR-pos. classes). In support of this, we also confirmed maintained nuclear AR expression in tissue samples collected after progression on abiraterone. AR staining also identified addnl. AR-pos. CD45-neg. circulating cells that were CK-neg./weak and therefore missed using std. protocols. The no. of these events correlated with traditional CTCs and was assocd. with worse outcome on univariate anal. Conclusions: We developed a non-invasive method to monitor AR nuclear expression in CTCs. Our studies confirm nuclear AR expression in CRPC patients progressing on novel endocrine treatments. Owing to the significant heterogeneity of AR expression in CTCs, studies in larger cohorts of patients are required to identify assocns. with outcome.
- 51Zeinali, M.; Murlidhar, V.; Fouladdel, S.; Shao, S.; Zhao, L.; Cameron, H.; Bankhead, A., III; Shi, J.; Cuneo, K. C.; Sahai, V.; Azizi, E.; Wicha, M. S.; Hafner, M.; Simeone, D. M.; Nagrath, S. Profiling heterogeneous circulating tumor cells (CTC) populations in pancreatic cancer using a serial microfluidic CTC carpet chip. Adv. Biosystems 2018, 2 (12), 1800228, DOI: 10.1002/adbi.201800228Google Scholar51https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXisFSnu7bE&md5=56c8b468d9e306f32d1d240b5d9cc7b1Profiling Heterogeneous Circulating Tumor Cells (CTC) Populations in Pancreatic Cancer Using a Serial Microfluidic CTC Carpet ChipZeinali, Mina; Murlidhar, Vasudha; Fouladdel, Shamileh; Shao, Shimeng; Zhao, Lili; Cameron, Heather; Bankhead, Armand, III; Shi, Jiaqi; Cuneo, Kyle C.; Sahai, Vaibhav; Azizi, Ebrahim; Wicha, Max S.; Hafner, Mathias; Simeone, Diane M.; Nagrath, SunithaAdvanced Biosystems (2018), 2 (12), 1800228/1-1800228/13CODEN: ABDIHL; ISSN:2366-7478. (Wiley-VCH Verlag GmbH & Co. KGaA)Although isolation of circulating tumor cells (CTCs) from pancreatic adenocarcinoma patients is feasible, investigating their clin. utility has proven less successful than other cancers due to the limitations of epithelial cellular adhesion mol. (EpCAM)-only based CTC assays. An integrated technol.- and biol.-based approach using a microfluidic "Carpet Chip" is presented to study the biol. relevance of heterogeneous CTC populations. Both epithelial CTCs (EpCs) and epithelial-to-mesenchymal transition (EMT)-like CTCs (EMTCs) are isolated simultaneously from the whole blood of pancreatic cancer (PaCa) patients (n = 35) by sep. targeting two surface markers: EpCAM and CD133. Recovery of cancer cell lines spiked into whole blood is ≥97% with >76% purity. Thirty-four patients had ≥5 EpCs mL-1 and 35 patients had ≥15 EMTCs mL-1. Overall, significantly higher nos. of EMTCs than EpCs are recovered, reflecting the aggressive nature of PaCa. Furthermore, higher nos. of EMTCs are obsd. in patients with lymph node involvement compared to patients without. Gene expression profiling of CTCs from 17 patients reveals that CXCR1 is significantly upregulated in EpCs, while known stem cell markers POU5F1/Oct-4 and MYC are upregulated in EMTCs. In conclusion, successful isolation and genomic profiling of heterogeneous CTC populations are demonstrated, revealing genetic signatures relevant to patient outcomes.
- 52Kang, Y.-T.; Kim, Y. J.; Bu, J.; Chen, S.; Cho, Y.-H.; Lee, H. M.; Ryu, C. J.; Lim, Y.; Han, S.-W. Epithelial and mesenchymal circulating tumor cell isolation and discrimination using dual-immunopatterned device with newly-developed anti-63B6 and anti-EpCAM. Sens. Actuators, B 2018, 260, 320– 330, DOI: 10.1016/j.snb.2017.12.157Google Scholar52https://chemport.cas.org/services/resolver?origin=ACS&resolution=options&coi=1%3ACAS%3A528%3ADC%252BC1cXmt1Oisg%253D%253D&md5=cd74b3e899b6fc273801b147fa8899b8Epithelial and mesenchymal circulating tumor cell isolation and discrimination using dual-immunopatterned device with newly-developed anti-63B6 and anti-EpCAMKang, Yoon-Tae; Kim, Young Jun; Bu, Jiyoon; Chen, Shilin; Cho, Young-Ho; Lee, Hyun Min; Ryu, Chun Jeih; Lim, Yoojoo; Han, Sae-WonSensors and Actuators, B: Chemical (2018), 260 (), 320-330CODEN: SABCEB; ISSN:0925-4005. (Elsevier B.V.)We report a dual-immunopatterned microfluidic device for capturing both epithelial and mesenchymal circulating tumor cells (CTCs) simultaneously in a single device using the two antibodies of anti-epithelial cell adhesion mol. (EpCAM) antibody and newly developed anti-63B6 antibody. In addn. to the conventional epithelial antibody of anti-EpCAM, our inhouse produced anti-63B6 antibody targets mesenchymal stem cell-like cancer cells and intermediate cancer cells, thus overcoming limitation of the conventional single anti-EpCAM based CTC isolations. These two antibodies are immobilized to each top and bottom layer of the present device and the combination of two layers forms a circular chamber with the center inlet. The mutual complementary cell isolation through competing reaction in different velocity zones between mesenchymal and epithelial antibodies enables the precise screening and profiling of CTCs depending on their positivity and degree of epithelial/mesenchymal surface antigen expression, which may differ in compliance with disease status. From the expts. using epithelial and mesenchymal-like cancer cells, the present device captures 94.47% of cancer cells, which is substantially higher than the EpCAM-only-based method in terms of heterogeneous cancer cell isolation including mesenchymal-like cancer cells. Patient blood samples were employed to assess the clin. application of the present device for examg. the patient status based on their CTC heterogeneity. The CTCs captured by both antibodies exhibit considerably varied expression profiles of epithelial protein including cytokeratin and EpCAM. The present device facilitates the isolation of heterogeneous subtypes of CTCs. These undiscovered heterogeneities would be helpful for precise anal. of CTCs to find their underlying meaning in cancer progression.
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