Development of an immunofluorescent AR-V7 circulating tumor cell assay – A blood-based test for men with metastatic prostate cancer
Introduction: Here we describe the development of a protein immunofluorescent assay for the detection of nuclear-localized androgen receptor variant 7 (AR-V7) protein within circulating tumor cells (CTCs) identified in patient blood samples. Used in the clinic, the test result serves as a validated biomarker of futility for patients with progressing metastatic castration-resistant prostate cancer (mCRPC) who are treated with androgen receptor targeted therapies (AATT) in whom nuclear-localized AR-V7 CTCs are identified and have received level 2A evidence in the 2019 National Cancer Center Network (NCCN) guidelines (v1.0).
Methods: Assay development was completed on the Epic Sciences rare cell detection platform using control cell lines of known AR-V7 status and clinical testing of mCRPC patient samples obtained at the decision point in management.
Results and conclusions: Using these samples, all assay parameters, scoring criteria, and clinical cutoffs for positivity were prospectively selected and locked. After assay lock, blinded clinical validation testing was initiated on multiple, independent, clinical cohorts as reported by Scher et al (JAMA Oncol. 2016;2:1441-1449; JAMA Oncol. 2018;4:1179-1186) and Armstrong et al (J Clin Oncol. 2019;37:1120-1129).
Beer TM, Armstrong AJ, Rathkopf DE, et al. Enzalutamide in metastatic prostate cancer before chemotherapy. N Engl J Med. 2014;371:424-433.
Scher HI, Fizazi K, Saad F, et al. Increased survival with enzalutamide in prostate cancer after chemotherapy. N Engl J Med. 2012;367:1187-1197.
Ryan CJ, Smith MR, Fizazi K, et al. Abiraterone acetate plus prednisone versus placebo plus prednisone in chemotherapy-naive men with metastatic castration-resistant prostate cancer (COU-AA-302): final overall survival analysis of a randomised, double-blind, placebo-controlled phase 3 study. Lancet Oncol. 2015;16:152-160.
de Bono JS, Logothetis CJ, Molina A, et al. Abiraterone and increased survival in metastatic prostate cancer. N Engl J Med. 2011;364:1995-2005.
Ryan CJ, Smith MR, de Bono JS, et al. Abiraterone in metastatic prostate cancer without previous chemotherapy. N Engl J Med. 2013;368:138-148.
Smith MR, Saad F, Chowdhury S, et al. Apalutamide treatment and metastasis-free survival in prostate cancer. N Engl J Med. 2018;378:1408-1418.
Fizazi K, Shore N, Tammela TL, et al. Darolutamide in nonmetastatic, castration-resistant prostate cancer. N Engl J Med. 2019;380:1235-1246.
George DJ, Sartor O, Miller K, et al. Treatment patterns and outcomes in patients with metastatic castration-resistant prostate cancer in a real-world clinical practice setting in the United States. Clin Genitourin Cancer. 2020;18:284-94.
Scher HI, Lu D, Schreiber NA, et al. 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;2:1441-1449.
Scher HI, Graf RP, Schreiber NA, et al. Assessment of the validity of nuclear-localized androgen receptor splice variant 7 in circulating tumor cells as a predictive biomarker for castration-resistant prostate cancer. JAMA Oncol. 2018;4:1179-1186.
Armstrong AJ, Halabi S, Luo J, et al. Prospective multicenter validation of androgen receptor splice variant 7 and hormone therapy resistance in high-risk castration-resistant prostate cancer: The PROPHECY Study. J Clin Oncol. 2019;37:1120-1129.
Werner SL, Graf RP, Landers M, et al. Analytical validation and capabilities of the epic CTC platform: enrichment-free circulating tumour cell detection and characterization. J Circ Biomark. 2015;4:3.
Scher HI, Graf RP, Schreiber NA, et al. Nuclear-specific AR-V7 protein localization is necessary to guide treatment selection in metastatic castration-resistant prostate cancer. Eur Urol. 2017;71:874-882.
Hu R, Dunn TA, Wei S, et al. Ligand-independent androgen receptor variants derived from splicing of cryptic exons signify hormone-refractory prostate cancer. Cancer Res. 2009;69:16-22.
Efstathiou E, Titus M, Wen S, et al. Molecular characterization of enzalutamide-treated bone metastatic castration-resistant prostate cancer. Eur Urol. 2015;67:53-60.
Qu Y, Dai B, Ye D, et al. Constitutively active AR-V7 plays an essential role in the development and progression of castration-resistant prostate cancer. Sci Rep. 2015;5:7654.
Welti J, Rodrigues DN, Sharp A, et al. Analytical validation and clinical qualification of a new immunohistochemical assay for androgen receptor splice variant-7 protein expression in metastatic castration-resistant prostate cancer. Eur Urol. 2016;70:599-608.
Antonarakis ES, Nakazawa M, Luo J. Resistance to androgen-pathway drugs in prostate cancer. N Engl J Med. 2014;371:2234.
Ballman KV. Biomarker: predictive or prognostic? J Clin Oncol. 2015;33:3968-3971.
Watson PA, Arora VK, Sawyers CL. Emerging mechanisms of resistance to androgen receptor inhibitors in prostate cancer. Nat Rev Cancer. 2015;15:701-711.
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