Editorial
Circulating Tumor Cells in Prostate Cancer: Does (Nuclear) Size Matter? Sumanta K. Pal, MD1; Neeraj Agarwal, MD2; and Jeremy O. Jones, PhD3
In the lay press, the terms “circulating tumor cell” (CTC) and “liquid biopsy” are often uttered in the same breath. Currently, however, the role of CTCs in clinical practice is solely as a prognostic tool as opposed to an aid in diagnosis or the personalization of therapy. In patients with prostate cancer, a correlation has been observed between baseline CTC enumeration and overall survival (OS) in a large cohort of men with castration-resistant prostate cancer (CRPC) who are receiving chemotherapy.1 More recently, a similar correlation with OS has been observed in men with metastatic CRPC treated on COU-AA-301, a pivotal phase 3 trial comparing abiraterone and prednisone in this population.2 As noted, these studies (both of which use the conventional CellSearch platform [Janssen Diagnostics, Raritan, NJ]) support the prognostic value of CTC enumeration; however, what remains elusive is a means of using this biomarker for treatment allocation. Although the study by Chen et al in this issue of Cancer does not definitively address this problem either, it does provide key insights into the role of CTCs in identifying unique subpopulations of prostate cancer.3 In their study, 57 men with varying stages of prostate cancer submitted a total of 148 blood samples. The NanoVelcro Chips system was used to isolate cells that were 4’,6-diamidino-2-phenylindole (DAPI) positive/cytokeratin (CK) positive/CD45 negative, and isolated cells were assessed at high magnification (3100 or 3400) for nuclear size. Visually, these cells appear to have greater clarity and resolution than those typically identified with the CellSearch platform. Nuclear size was measured and trichotomized into large nuclear CTC, small nuclear CTC (snCTC), and very small nuclear CTC (vsnCTC) categories. Two key observations were noted. First, snCTCs and vsnCTCs were more frequent in patients with metastatic disease compared with patients with localized disease. Second, vsnCTCs were more frequent in patients with visceral metastatic disease (eg, lesions in the lungs or liver) compared with those with bone-only or lymph node-only metastases. The methods used to characterize CTCs in this experience also warrant a second look. A recent report from Ferraldeschi et al used the Epic Sciences platform (Epic Sciences Inc, La Jolla, Calif) to compare samples derived from 39 patients with metastatic CRPC with those from 10 healthy volunteers.4 Of the 39 patients, 36 (92%) had CTCs detected that were CK negative, and among these, multiple patients had snCTCs identified. Thus, because the examination was limited to CK-positive events, the sensitivity of the current study by Chen et al could be limited. The findings of Chen et al’s study are tempered by the limited sample size, which was divided further into subcategories based on clinical status. For example, the comparison of visceral versus nonvisceral metastases was predicated on a comparison of 21 and 13 patients, respectively.3 Furthermore, little description was offered related to the extent of prior therapy. It was noted that patients with visceral metastases had received a multitude of prior therapies, including cytotoxic chemotherapy, novel endocrine therapies, and radioisotopes, yet it was unclear what treatment (if any) patients with nonvisceral metastases received. The possibility that vsnCTCs emerge as a function of prior systemic therapy cannot be excluded. Finally, many have drawn the association between visceral metastases and the presence of neuroendocrine prostate cancer.5 The authors reported anecdotal evidence that certain patients bore vsnCTCs expressing synaptophysin, one of several markers (such as vimentin, Aurora kinase A, etc) that could delineate neuroendocrine prostate cancer.6 More extensive assessment of these markers is necessary to determine whether they could serve as a better (and more biologically relevant) identifier of visceral disease compared with nuclear size.
Corresponding author: Sumanta K. Pal, MD, Department of Medical Oncology and Experimental Therapeutics, City of Hope Comprehensive Cancer Center, 1500 East Duarte Rd, Duarte, CA 91010; Fax: (626) 301-8233; [email protected] 1 Department of Medical Oncology and Experimental Therapeutics, City of Hope Comprehensive Cancer Center, Duarte, California; 2Division of Medical Oncology, Huntsman Cancer Institute, University of Utah, Salt Lake City, Utah; 3Department of Cancer Biology, City of Hope Comprehensive Cancer Center, Duarte, California
See referenced original article on pages 3240-51, this issue. DOI: 10.1002/cncr.29454, Received: March 30, 2015; Accepted: April 6, 2015, Published online May 14, 2015 in Wiley Online Library (wileyonlinelibrary.com)
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Cancer
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CTCs in Prostate Cancer/Pal et al
As Chen et al concede, their current study is exploratory; however, it a useful exercise in envisioning how the nuclear sizing of CTCs might be used in the clinic. The landscape of potential CTC-based diagnostics in prostate cancer is increasingly crowded. Furthest along are Antonarakis et al from Johns Hopkins University, who recently reported a prospective assessment of androgen receptor splice variant V7 in a series of patients receiving either novel endocrine therapies or taxanes.7,8 In patients receiving abiraterone or enzalutamide, the presence of androgen receptor splice variant V7 in CTCs predicted treatment resistance. The same predictive value was not observed within the context of taxane therapy. With further prospective validation, this biomarker may ultimately help to guide decisionmaking between cytotoxic and endocrine therapy. How might nuclear sizing be applied in practice? If the association between vsnCTCs and visceral disease holds true, there may be nuanced clinical settings in which the biomarker can be applied. For example, recent data from a phase 3 study of ipilimumab versus placebo in patients with metastatic CRPC who were treated with prior docetaxel therapy suggested a modest OS advantage with ipilimumab in patients with no visceral metastases (OS of 14.4 months vs 10.3 months; hazard ratio, 0.73 [95% confidence interval, 0.59-0.89]).9 In contrast, patients with visceral metastases appeared to not benefit from the drug (OS of 5.7 months vs 7.4 months; hazard ratio, 1.2 [95% confidence interval, 0.90-1.60]). With these data in mind, it would be interesting to learn whether patients with vsnCTCs (and likely visceral disease) could benefit from this immunotherapeutic approach. Furthermore, although there has been diminished enthusiasm for cabozantinib (a dual inhibitor of MET and vascular endothelial growth factor receptor 2) in the treatment of prostate cancer on the basis of recent phase 3 trial data, the investigators of the current study are leading an innovative effort exploring the agent in patients with metastatic CPRC with visceral metastases.10,11 Again, if vsnCTCs correlate with visceral disease, perhaps they could be explored as a surrogate for treatment response in this prospective trial. Offered here is conjecture regarding how the nuclear sizing of CTCs might be used in the clinic, but an important initial step will be to validate these results in larger series. Refining the patient population (eg, ensuring consistency in prior therapy) also will be essential, and the strategy of nuclear sizing must be compared with phenotypic characterization using cell surface markers. In addition, tools are now available that facilitate the comprehensive genomic profiling of single cells; it would Cancer
September 15, 2015
be of value to learn whether differences in nuclear size emanate from changes in genotype.12 The value of blood tests that do not rely on CTC capture, such as newer platforms assessing cell-free DNA and microRNA, also must be assessed.13,14 Ultimately, although Chen et al have done well to define a novel methodology for CTC assessment in patients with prostate cancer,3 the onus is now on them to generate a unique clinical niche among multiple competing diagnostics. FUNDING SUPPORT No specific funding was disclosed.
CONFLICT OF INTEREST DISCLOSURES Dr. Pal receives honoraria from Medivation and consulting fees from Astellas Pharma US for work performed outside of the current study. Dr. Jones is currently working on a collaborative research project involving circulating tumor cells with Janssen Diagnostics.
REFERENCES 1. de Bono JS, Scher HI, Montgomery RB, et al. Circulating tumor cells predict survival benefit from treatment in metastatic castrationresistant prostate cancer. Clin Cancer Res. 2008;14:6302-6309. 2. Scher HI, Heller G, Molina A, et al. Evaluation of circulating tumor cell (CTC) enumeration as an efficacy response biomarker of overall survival (OS) in metastatic castration-resistant prostate cancer (mCRPC): planned final analysis (FA) of COU-AA-301, a randomized, double-blind, placebo-controlled, phase III study of abiraterone acetate (AA) plus low-dose prednisone (P) post docetaxel [abstract]. J Clin Oncol. 2011;29:LBA4517. 3. Chen JF, Ho H, Lichterman J, et al. Subclassification of prostate cancer circulating tumor cells by nuclear size reveals very small nuclear circulating tumor cells in patients with visceral metastases. Cancer. 2015;121:3240-3251. 4. Ferraldeschi R, McDaniel A, Krupa R, et al. CK- and small nuclear size circulating tumor cell (CTCs) phenotypes in metastatic castration-resistant prostate cancer (mCRPC) [abstract]. J Clin Oncol. 2014;32:209. 5. Beltran H, Tomlins S, Aparicio A, et al. Aggressive variants of castration-resistant prostate cancer. Clin Cancer Res. 2014;20: 2846-2850. 6. Beltran H, Jendrisak A, Landers M, et al. Phenotypic characterization of circulating tumor cells (CTCs) from neuroendocrine prostate cancer (NEPC) and metastatic castration-resistant prostate cancer (mCRPC) patients to identify a novel diagnostic algorithm for the presence of NEPC [abstract]. J Clin Oncol. 2015;33:197. 7. Antonarakis ES, Lu C, Wang H, et al. Androgen receptor splice variant, AR-V7, and resistance to enzalutamide and abiraterone in men with metastatic castration-resistant prostate cancer (mCRPC) [abstract]. J Clin Oncol. 2014;32:5001. 8. Antonarakis ES, Lu C, Wang H, et al. AR-V7 and resistance to enzalutamide and abiraterone in prostate cancer. N Engl J Med. 2014;371:1028-1038. 9. Drake CG, Kwon ED, Fizazi K, et al. Results of subset analyses on overall survival (OS) from study CA184-043: ipilimumab (Ipi) versus placebo (Pbo) in post-docetaxel metastatic castration-resistant prostate cancer (mCRPC) [abstract]. J Clin Oncol. 2014;32:2. 10. ClinicalTrials.gov. NCT01834651: a phase II study of cabozantinib (XL184) therapy in castrate resistant prostate cancer (CRPC) with visceral metastases. Available at http://www.clinicaltrials.gov. Accessed March 17, 2015. 11. Smith MR, De Bono JS, Sternberg CN, et al. Final analysis of COMET-1: cabozantinib (Cabo) versus prednisone (Pred) in
3191
Editorial metastatic castration-resistant prostate cancer (mCRPC) patients (pts) previously treated with docetaxel (D) and abiraterone (A) and/ or enzalutamide (E) [abstract]. J Clin Oncol. 2015;33:139. 12. Peeters DJ, De Laere B, Van den Eynden GG, et al. Semiautomated isolation and molecular characterisation of single or highly purified tumour cells from CellSearch enriched blood samples using dielectrophoretic cell sorting. Br J Cancer. 2013;108:1358-1367.
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13. Azad AA, Volik SV, Wyatt AW, et al. Androgen receptor gene aberrations in circulating cell-free DNA: biomarkers of therapeutic resistance in castration-resistant prostate cancer [published online ahead of print February 23, 2015]. Clin Cancer Res. 14. Lin HM, Castillo L, Mahon KL, et al. Circulating microRNAs are associated with docetaxel chemotherapy outcome in castrationresistant prostate cancer. Br J Cancer. 2014;110:2462-2471.
Cancer
September 15, 2015
Circulating Tumor Cells in Prostate Cancer: Does (Nuclear) Size Matter? Sumanta K. Pal, MD1; Neeraj Agarwal, MD2; and Jeremy O. Jones, PhD3
In the lay press, the terms “circulating tumor cell” (CTC) and “liquid biopsy” are often uttered in the same breath. Currently, however, the role of CTCs in clinical practice is solely as a prognostic tool as opposed to an aid in diagnosis or the personalization of therapy. In patients with prostate cancer, a correlation has been observed between baseline CTC enumeration and overall survival (OS) in a large cohort of men with castration-resistant prostate cancer (CRPC) who are receiving chemotherapy.1 More recently, a similar correlation with OS has been observed in men with metastatic CRPC treated on COU-AA-301, a pivotal phase 3 trial comparing abiraterone and prednisone in this population.2 As noted, these studies (both of which use the conventional CellSearch platform [Janssen Diagnostics, Raritan, NJ]) support the prognostic value of CTC enumeration; however, what remains elusive is a means of using this biomarker for treatment allocation. Although the study by Chen et al in this issue of Cancer does not definitively address this problem either, it does provide key insights into the role of CTCs in identifying unique subpopulations of prostate cancer.3 In their study, 57 men with varying stages of prostate cancer submitted a total of 148 blood samples. The NanoVelcro Chips system was used to isolate cells that were 4’,6-diamidino-2-phenylindole (DAPI) positive/cytokeratin (CK) positive/CD45 negative, and isolated cells were assessed at high magnification (3100 or 3400) for nuclear size. Visually, these cells appear to have greater clarity and resolution than those typically identified with the CellSearch platform. Nuclear size was measured and trichotomized into large nuclear CTC, small nuclear CTC (snCTC), and very small nuclear CTC (vsnCTC) categories. Two key observations were noted. First, snCTCs and vsnCTCs were more frequent in patients with metastatic disease compared with patients with localized disease. Second, vsnCTCs were more frequent in patients with visceral metastatic disease (eg, lesions in the lungs or liver) compared with those with bone-only or lymph node-only metastases. The methods used to characterize CTCs in this experience also warrant a second look. A recent report from Ferraldeschi et al used the Epic Sciences platform (Epic Sciences Inc, La Jolla, Calif) to compare samples derived from 39 patients with metastatic CRPC with those from 10 healthy volunteers.4 Of the 39 patients, 36 (92%) had CTCs detected that were CK negative, and among these, multiple patients had snCTCs identified. Thus, because the examination was limited to CK-positive events, the sensitivity of the current study by Chen et al could be limited. The findings of Chen et al’s study are tempered by the limited sample size, which was divided further into subcategories based on clinical status. For example, the comparison of visceral versus nonvisceral metastases was predicated on a comparison of 21 and 13 patients, respectively.3 Furthermore, little description was offered related to the extent of prior therapy. It was noted that patients with visceral metastases had received a multitude of prior therapies, including cytotoxic chemotherapy, novel endocrine therapies, and radioisotopes, yet it was unclear what treatment (if any) patients with nonvisceral metastases received. The possibility that vsnCTCs emerge as a function of prior systemic therapy cannot be excluded. Finally, many have drawn the association between visceral metastases and the presence of neuroendocrine prostate cancer.5 The authors reported anecdotal evidence that certain patients bore vsnCTCs expressing synaptophysin, one of several markers (such as vimentin, Aurora kinase A, etc) that could delineate neuroendocrine prostate cancer.6 More extensive assessment of these markers is necessary to determine whether they could serve as a better (and more biologically relevant) identifier of visceral disease compared with nuclear size.
Corresponding author: Sumanta K. Pal, MD, Department of Medical Oncology and Experimental Therapeutics, City of Hope Comprehensive Cancer Center, 1500 East Duarte Rd, Duarte, CA 91010; Fax: (626) 301-8233; [email protected] 1 Department of Medical Oncology and Experimental Therapeutics, City of Hope Comprehensive Cancer Center, Duarte, California; 2Division of Medical Oncology, Huntsman Cancer Institute, University of Utah, Salt Lake City, Utah; 3Department of Cancer Biology, City of Hope Comprehensive Cancer Center, Duarte, California
See referenced original article on pages 3240-51, this issue. DOI: 10.1002/cncr.29454, Received: March 30, 2015; Accepted: April 6, 2015, Published online May 14, 2015 in Wiley Online Library (wileyonlinelibrary.com)
3190
Cancer
September 15, 2015
CTCs in Prostate Cancer/Pal et al
As Chen et al concede, their current study is exploratory; however, it a useful exercise in envisioning how the nuclear sizing of CTCs might be used in the clinic. The landscape of potential CTC-based diagnostics in prostate cancer is increasingly crowded. Furthest along are Antonarakis et al from Johns Hopkins University, who recently reported a prospective assessment of androgen receptor splice variant V7 in a series of patients receiving either novel endocrine therapies or taxanes.7,8 In patients receiving abiraterone or enzalutamide, the presence of androgen receptor splice variant V7 in CTCs predicted treatment resistance. The same predictive value was not observed within the context of taxane therapy. With further prospective validation, this biomarker may ultimately help to guide decisionmaking between cytotoxic and endocrine therapy. How might nuclear sizing be applied in practice? If the association between vsnCTCs and visceral disease holds true, there may be nuanced clinical settings in which the biomarker can be applied. For example, recent data from a phase 3 study of ipilimumab versus placebo in patients with metastatic CRPC who were treated with prior docetaxel therapy suggested a modest OS advantage with ipilimumab in patients with no visceral metastases (OS of 14.4 months vs 10.3 months; hazard ratio, 0.73 [95% confidence interval, 0.59-0.89]).9 In contrast, patients with visceral metastases appeared to not benefit from the drug (OS of 5.7 months vs 7.4 months; hazard ratio, 1.2 [95% confidence interval, 0.90-1.60]). With these data in mind, it would be interesting to learn whether patients with vsnCTCs (and likely visceral disease) could benefit from this immunotherapeutic approach. Furthermore, although there has been diminished enthusiasm for cabozantinib (a dual inhibitor of MET and vascular endothelial growth factor receptor 2) in the treatment of prostate cancer on the basis of recent phase 3 trial data, the investigators of the current study are leading an innovative effort exploring the agent in patients with metastatic CPRC with visceral metastases.10,11 Again, if vsnCTCs correlate with visceral disease, perhaps they could be explored as a surrogate for treatment response in this prospective trial. Offered here is conjecture regarding how the nuclear sizing of CTCs might be used in the clinic, but an important initial step will be to validate these results in larger series. Refining the patient population (eg, ensuring consistency in prior therapy) also will be essential, and the strategy of nuclear sizing must be compared with phenotypic characterization using cell surface markers. In addition, tools are now available that facilitate the comprehensive genomic profiling of single cells; it would Cancer
September 15, 2015
be of value to learn whether differences in nuclear size emanate from changes in genotype.12 The value of blood tests that do not rely on CTC capture, such as newer platforms assessing cell-free DNA and microRNA, also must be assessed.13,14 Ultimately, although Chen et al have done well to define a novel methodology for CTC assessment in patients with prostate cancer,3 the onus is now on them to generate a unique clinical niche among multiple competing diagnostics. FUNDING SUPPORT No specific funding was disclosed.
CONFLICT OF INTEREST DISCLOSURES Dr. Pal receives honoraria from Medivation and consulting fees from Astellas Pharma US for work performed outside of the current study. Dr. Jones is currently working on a collaborative research project involving circulating tumor cells with Janssen Diagnostics.
REFERENCES 1. de Bono JS, Scher HI, Montgomery RB, et al. Circulating tumor cells predict survival benefit from treatment in metastatic castrationresistant prostate cancer. Clin Cancer Res. 2008;14:6302-6309. 2. Scher HI, Heller G, Molina A, et al. Evaluation of circulating tumor cell (CTC) enumeration as an efficacy response biomarker of overall survival (OS) in metastatic castration-resistant prostate cancer (mCRPC): planned final analysis (FA) of COU-AA-301, a randomized, double-blind, placebo-controlled, phase III study of abiraterone acetate (AA) plus low-dose prednisone (P) post docetaxel [abstract]. J Clin Oncol. 2011;29:LBA4517. 3. Chen JF, Ho H, Lichterman J, et al. Subclassification of prostate cancer circulating tumor cells by nuclear size reveals very small nuclear circulating tumor cells in patients with visceral metastases. Cancer. 2015;121:3240-3251. 4. Ferraldeschi R, McDaniel A, Krupa R, et al. CK- and small nuclear size circulating tumor cell (CTCs) phenotypes in metastatic castration-resistant prostate cancer (mCRPC) [abstract]. J Clin Oncol. 2014;32:209. 5. Beltran H, Tomlins S, Aparicio A, et al. Aggressive variants of castration-resistant prostate cancer. Clin Cancer Res. 2014;20: 2846-2850. 6. Beltran H, Jendrisak A, Landers M, et al. Phenotypic characterization of circulating tumor cells (CTCs) from neuroendocrine prostate cancer (NEPC) and metastatic castration-resistant prostate cancer (mCRPC) patients to identify a novel diagnostic algorithm for the presence of NEPC [abstract]. J Clin Oncol. 2015;33:197. 7. Antonarakis ES, Lu C, Wang H, et al. Androgen receptor splice variant, AR-V7, and resistance to enzalutamide and abiraterone in men with metastatic castration-resistant prostate cancer (mCRPC) [abstract]. J Clin Oncol. 2014;32:5001. 8. Antonarakis ES, Lu C, Wang H, et al. AR-V7 and resistance to enzalutamide and abiraterone in prostate cancer. N Engl J Med. 2014;371:1028-1038. 9. Drake CG, Kwon ED, Fizazi K, et al. Results of subset analyses on overall survival (OS) from study CA184-043: ipilimumab (Ipi) versus placebo (Pbo) in post-docetaxel metastatic castration-resistant prostate cancer (mCRPC) [abstract]. J Clin Oncol. 2014;32:2. 10. ClinicalTrials.gov. NCT01834651: a phase II study of cabozantinib (XL184) therapy in castrate resistant prostate cancer (CRPC) with visceral metastases. Available at http://www.clinicaltrials.gov. Accessed March 17, 2015. 11. Smith MR, De Bono JS, Sternberg CN, et al. Final analysis of COMET-1: cabozantinib (Cabo) versus prednisone (Pred) in
3191
Editorial metastatic castration-resistant prostate cancer (mCRPC) patients (pts) previously treated with docetaxel (D) and abiraterone (A) and/ or enzalutamide (E) [abstract]. J Clin Oncol. 2015;33:139. 12. Peeters DJ, De Laere B, Van den Eynden GG, et al. Semiautomated isolation and molecular characterisation of single or highly purified tumour cells from CellSearch enriched blood samples using dielectrophoretic cell sorting. Br J Cancer. 2013;108:1358-1367.
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13. Azad AA, Volik SV, Wyatt AW, et al. Androgen receptor gene aberrations in circulating cell-free DNA: biomarkers of therapeutic resistance in castration-resistant prostate cancer [published online ahead of print February 23, 2015]. Clin Cancer Res. 14. Lin HM, Castillo L, Mahon KL, et al. Circulating microRNAs are associated with docetaxel chemotherapy outcome in castrationresistant prostate cancer. Br J Cancer. 2014;110:2462-2471.
Cancer
September 15, 2015
