Combined genomic profiling of cell-free DNA (cfDNA) and circulating tumor cell DNA (ctcDNA) in S1802, a prospective phase 3 trial for metastatic prostate cancer (mHSPC).

Abstract

e17111 Background: Parallel analysis of cfDNA and ctcDNA can yield expanded and complementary molecular insights. We developed HERCULES, a prostate cancer specific targeted amplicon sequencing panel capable of assessing single nucleotide variants (SNVs) and copy number variants (CNVs) in cfDNA and single CTCs obtained concurrently from a single tube of blood. Here we present results from the first 35 patients analyzed at multiple time points in S1802, an NCTN/SWOG randomized prospective phase 3 trial of standard systemic therapy +/- definitive treatment of the primary tumor in newly diagnosed mHSPC. Methods: Blood was collected in Streck DNA preservative tubes at 4 prespecified time points: registration, randomization, post-definitive therapy, and progression. Plasma and single CTCs were isolated using the RareCyte platform, and DNA was extracted using the Apostle MiniMax cfDNA extraction kit (Beckman) or Single Cell Lysis Kit (Thermo Fisher). The HERCULES AmpliSeq-HD panel includes 35 prostate cancer -relevant genes. SNVs and CNVs were called using Ion Reporter Software (Thermo Fisher) and filtered for variants associated with clonal hematopoiesis of indeterminate significance (CHIP). Results: A total of 85 samples were obtained from 35 patients at up to 4 different time points. CTCs were detected in 22(61%) patients, with median 3 CTCs/7.5ml (range 1-986). SNV limit of detection varied with input DNA, from 0.16% at >20 ng input to 0.91% with 1-5 ng input. Fewer than 5% of all SNVs were censored due to potential CHIP. Across all times points, SNVs and CNVs were detected in 32 and 20 patients, respectively. SNVs were detected both in cfDNA and in ctcDNA. Known prostate cancer driver variants were observed in AR, CTNNB1, FOXA1, SF3B1, TP53 and others. Recurrent CNVs were observed in AR, AR enhancer, FOXA1, CDK4, MYC and CHD1. In general, the 4 th time point, representing transition to mCRPC, had twice the number of alterations as earlier mHSPC time points. Conclusions: Analysis of the first 35 patients sequenced at multiple time points in S1802 demonstrates the feasibility of concurrent, same-sample genomic profiling of cfDNA and ctcDNA in a multi-center prospective phase 3 NCTN setting. The HERCULES sequencing workflow enables parallel processing of cfDNA and single-cell ctcDNA, with LOD that exceeds that of hybrid capture panels at these input levels. Presence of concordant and distinct alterations in cfDNA and CTCs is consistent with findings from prior small studies and underscores the potential value of analyzing both of these liquid biopsy components.