Genomic Analysis of Circulating Tumor DNA Identifies Recurrent Molecular Features with Clinical Significance in Advanced Prostate Cancer

Abstract

<h3>Purpose/Objective(s)</h3> Circulating tumor DNA (ctDNA) is a promising approach for noninvasive molecular profiling. However, the feasibility of ctDNA analysis remains to be established in scenarios where the circulating tumor fraction (TF) is low, as can be the case with prostate adenocarcinoma. Ultra-low-pass whole genome sequencing (ULP-WGS) is a cost-effective method for estimating the TF and may identify patients with a sufficiently high TF to allow for ctDNA to be used in comprehensive molecular profiling. We examined whether targeted panel sequencing (TPS) with a custom panel which includes intergenic regions is feasible for identifying clinically relevant molecular alterations and novel biomarkers in patients with metastatic castrate-resistant prostate cancer (mCRPC) and a sufficiently high TF (as estimated from ULP-WGS) prior to starting docetaxel. <h3>Materials/Methods</h3> Patients with mCRPC who received docetaxel at our institution and with available pre-treatment plasma specimens were retrospectively identified, and isolated ctDNA underwent ULP-WGS. The ichorCNA algorithm was used to estimate TF and assess large-scale copy number alterations. TPS using an institutional prostate cancer-specific panel of 319 genes with duplex sequencing (utilizing unique molecular identifiers) for error suppression was performed on samples with TF ≥3-7% and used to call single nucleotide variants, insertions/deletions, and <i>ETS</i> fusions in a tumor-naïve manner. The association of clinicogenomic features with response to docetaxel, as determined by a prostate-specific antigen (PSA) decline ≥50% (PSA50) within 16 week of treatment initiation, was assessed using logistic regression. <h3>Results</h3> Banked specimens from 144 patients who initiated docetaxel between July 2001 and May 2016 underwent ULP-WGS. Median PSA was 99 ng/mL (interquartile range [IQR] 25-290 ng/mL). 81 patients with TF ≥3-7% (median 12%, IQR 5-29%) underwent TPS. Mean duplex target coverage was 403x. TPS identified evidence of <i>ETS</i> fusions in 56% (n=45) of samples as well as mutations in genes known to be recurrently mutated in prostate cancer, including <i>TP53</i> (44%, n=34), <i>AR</i> (14%, n=11), <i>BRCA2</i> (6%, n=5, with 4 patients having pathogenic germline variants), and <i>CHEK2</i> (4%, n=3). ichorCNA additionally identified recurrent <i>AR</i> amplifications (26%, n=21). PSA50 was observed in 47% (n=38) of the cohort. On univariable analysis, ploidy as determined by ichorCNA was significantly associated with likelihood of PSA50 (21% vs 52% for ploidy ≥3 vs <3), and this association persisted on multivariable analysis (adjusted odds ratio 0.21, 95% confidence interval 0.04-0.77, <i>P</i>=0.029). <h3>Conclusion</h3> Screening of ctDNA with ULP-WGS is feasible for identifying patients with advanced prostate cancer who are suitable for comprehensive molecular profiling. In addition to detecting clinically relevant molecular alterations, ctDNA analysis nominated ploidy as a potential novel biomarker in the setting of docetaxel, which warrants further investigation.