Abstract LB-269: Detection of cell-free DNA alterations in AR and its enhancer significantly outperformed CTC AR-V7 detection for identifying treatment resistance in patients with metastatic prostate cancer
Authors: Chauhan Pradeep S. , Dang Ha X. , Ellis Haley , Feng Wenjia , Harris Peter K. , Smith Grace , Qiao Mark , Dienstbach Katherine , Beck Rachel , Atkocius Andrew , Qaium Faridi , Luo Jingqin , Michalski Jeff M. , Picus Joel , Pachynski Russell K. , Maher Christopher A. , Chaudhuri Aadel A. 
Published: September 2020

Abstract
Background: Liquid biopsy analysis of cell-free DNA (cfDNA) and circulating tumor cells (CTCs) have emerged as potential tools to predict response to androgen receptor (AR)-directed therapy in metastatic prostate cancer (mPCa). However, current clinically available assays (e.g. CTC AR-V7) focus on one AR variant, demonstrate low sensitivity, and remain underutilized. The recent discovery of AR enhancer amplification in 81% of metastatic patients and its association with disease resistance opens opportunities to improve upon current assays. We hypothesized that tracking AR/enhancer genomic alterations in plasma cfDNA would more sensitively detect resistance with higher accuracy than the CTC AR-V7 assay used in clinical practice. Methods: We developed a targeted sequencing assay and analysis method called Enhancer and neighboring loci of Androgen Receptor Sequencing (EnhanceAR-Seq). We applied EnhanceAR-Seq to plasma collected from 40 patients with mPCa treated with AR-directed therapy to monitor AR/enhancer genomic alterations and correlated these events with therapy resistance, progression-free survival (PFS), and overall survival (OS). In parallel, the Oncotype DX AR-V7 Nucleus Detect CTC assay was performed in peripheral blood from a subset of patients. Results: The most frequent genomic events detected were AR/enhancer alterations (copy number variation, tandem duplication or missense mutations) in 18 patients (45%), of which 16 patients (40%) had AR enhancer copy number gain or tandem duplication. Three patients (8%) were found to have independent AR enhancer amplification without AR gene body amplification. TP53 (8 patients, 20%) and PTEN (6 patients, 15%) were the other most frequently altered genes, subject to nonsynonymous mutations and copy number alterations. We also detected the TMPRSS2-ERG gene fusion in 5 cases (13%). cfDNA-detected alterations in the full AR locus including the AR enhancer were highly significant for overall survival (P=0.0015; HR=11.5) by Kaplan-Meier analysis, as well as progression-free survival by Kaplan-Meier analysis (P=0.0002; HR=6.8) and multivariate Cox proportional hazards regression (P=0.001; HR=12.7). While AR/enhancer alterations in plasma cfDNA identified 18 out of 23 treatment-resistant cases with 100% positive predictive value and 78% sensitivity (Fisher's P&lt;0.0001), the Oncotype AR-V7 CTC assay in 25 patients performed poorly with 50% positive predictive value and 6% sensitivity (Fisher's P&gt;0.99). Conclusions: cfDNA based AR locus alterations including the enhancer are strongly associated with resistance to AR-directed therapy and significantly worse survival outcomes. EnhanceAR-Seq significantly outperformed the CTC AR-V7 assay used in standard clinical practice.
Citation Format: Pradeep S. Chauhan, Ha X. Dang, Haley Ellis, Wenjia Feng, Peter K. Harris, Grace Smith, Mark Qiao, Katherine Dienstbach, Rachel Beck, Andrew Atkocius, Faridi Qaium, Jingqin Luo, Jeff M. Michalski, Joel Picus, Russell K. Pachynski, Christopher A. Maher, Aadel A. Chaudhuri. Detection of cell-free DNA alterations in AR and its enhancer significantly outperformed CTC AR-V7 detection for identifying treatment resistance in patients with metastatic prostate cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-269.