Primary resistance to androgen receptor (AR) directed therapies in metastatic castrationresistant prostate cancer (mCRPC) is poorly understood. We randomized 202 treatment-naive mCRPC patients to abiraterone or enzalutamide, and performed whole exome and deep targeted 72-gene sequencing of plasma cell-free DNA prior to therapy. For these agents, which have never been directly compared, time to progression was similar. Defects in BRCA2 and ATM were strongly associated with poor clinical outcomes independently of clinical prognostic factors and circulating tumor DNA abundance. Somatic alterations in TP53, previously linked to reduced tumor dependency on AR signaling, were also independently associated with rapid resistance. Although detection of AR amplifications did not outperform standard prognostic biomarkers, AR gene structural rearrangements truncating the ligand binding domain were identified in several patients with primary resistance. These findings establish genomic drivers of resistance to first-line AR directed therapy in mCRPC and identify potential minimally-invasive biomarkers. Statement of SignificanceLeveraging plasma specimens collected in a large randomized phase II trial, we report the relative impact of common circulating tumor DNA alterations on patient response to the most widely-used therapies for advanced prostate cancer. Our findings suggest that liquid biopsy analysis can guide the use of AR-targeted therapy in general practice.

Background Abiraterone + prednisone (abiraterone) and enzalutamide are both indicated for the treatment of metastatic castration-resistant prostate cancer (mCRPC). We aimed to determine the best sequence in which to utilize both agents as well as their second-line efficacy. Methods In this multicentre, randomized, open-label phase II crossover trial conducted across 6 cancer centres in British Columbia, Canada, patients ≥ 18 years with newly-diagnosed mCRPC without neuroendocrine differentiation and ECOG performance status ≤ 2 were randomized 1:1 using simple randomization to receive abiraterone 1000 mg orally daily plus prednisone 5 mg orally twice daily followed by enzalutamide 160 mg orally daily (arm A), or the opposite sequence (arm B). Primary endpoints were time to second PSA progression and PSA response rate (≥ 30% decline) on second-line therapy, analyzed by intention-to-treat in randomized patients and patients that crossed over, respectively. The trial is registered with ClinicalTrials.gov, number NCT02125357. The trial is completed and final analyses are reported here. Findings 202 patients were randomized (101 to each arm) between October 21, 2014 and December 13, 2016. At the time of data cutoff 73 and 75 patients had crossed over in arm A and B, respectively. Time to second PSA progression was longer in arm A (median 19•3 vs 15•2 months, HR = 0•66, 95% CI 0•45-0•97, p = 0•036), at a median followup of 22•8 months (IQR 10•3-33•4). Second-line PSA response rates were 36% for enzalutamide and 4% for abiraterone (p < 0•0001). The most common grade 3-4 adverse events were hypertension (27 [27%] of 101 patients in arm A vs 18 [18%] of 101 in arm B) and fatigue (10 [10%] vs 4 [4%]). Serious adverse events were reported in 15 (15%) of 101 patients in arm A and 20 (20%) of 101 in arm B. There were no treatment related deaths.

Molecular stratification can improve the management of advanced cancers, but requires relevant tumor samples. Metastatic urothelial carcinoma (mUC) is poised to benefit given a recent expansion of treatment options and its high genomic heterogeneity. We profile minimally-invasive plasma circulating tumor DNA (ctDNA) samples from 104 mUC patients, and compare to same-patient tumor tissue obtained during invasive surgery. Patient ctDNA abundance is independently prognostic for overall survival in patients initiating first-line systemic therapy. Importantly, ctDNA analysis reproduces the somatic driver genome as described from tissue-based cohorts. Furthermore, mutation concordance between ctDNA and matched tumor tissue is 83.4%, enabling benchmarking of proposed clinical biomarkers. While 90% of mutations are identified across serial ctDNA samples, concordance for serial tumor tissue is significantly lower. Overall, our exploratory analysis demonstrates that genomic profiling of ctDNA in mUC is reliable and practical, and mitigates against disease undersampling inherent to studying archival primary tumor foci. We urge the incorporation of cell-free DNA profiling into molecularly-guided clinical trials for mUC.

Purpose: DNA mismatch repair defects (MMRd) and tumor hypermutation are rare and under-characterized in metastatic prostate cancer (mPC). Furthermore, because hypermutated MMRd prostate cancers can respond to immune checkpoint inhibitors, there is an urgent need for practical detection tools.Experimental Design: We analyzed plasma cell-free DNAtargeted sequencing data from 433 patients with mPC with circulating tumor DNA (ctDNA) purity !2%. Samples with somatic hypermutation were subjected to 185 Â whole-exome sequencing and capture of mismatch repair gene introns. Archival tissue was analyzed with targeted sequencing and IHC.Results: Sixteen patients (3.7%) had somatic hypermutation with MMRd etiology, evidenced by deleterious alterations in MSH2, MSH6, or MLH1, microsatellite instability, and characteristic trinucleotide signatures. ctDNA was concordant with mismatch repair protein IHC and DNA sequencing of tumor tissue. Tumor suppressors such as PTEN, RB1, and TP53 were inactivated by mutation rather than copy-number loss. Hotspot mutations in oncogenes such as AKT1, PIK3CA, and CTNNB1 were common, and the androgen receptor (AR)-ligand binding domain was mutated in 9 of 16 patients. We observed high intrapatient clonal diversity, evidenced by subclonal driver mutations and shifts in mutation allele frequency over time. Patients with hypermutation and MMRd etiology in ctDNA had a poor response to AR inhibition and inferior survival compared with a control cohort.Conclusions: Hypermutated MMRd mPC is associated with oncogene activation and subclonal diversity, which may contribute to a clinically aggressive disposition in selected patients. In patients with detectable ctDNA, cell-free DNA sequencing is a practical tool to prioritize this subtype for immunotherapy.See related commentary by Schweizer and Yu, p. 981

Purpose: Cross-resistance renders multiple lines of androgen receptor (AR) signaling inhibitors increasingly futile in metastatic castration-resistant prostate cancer (mCRPC). We sought to determine acquired genomic contributors to cross-resistance. Experimental Design: We collected 458 serial plasma cell-free DNA samples at baseline and progression timepoints from 202 patients with mCRPC receiving sequential AR signaling inhibitors (abiraterone and enzalutamide) in a randomized phase II clinical trial (NCT02125357). We utilized deep targeted and whole-exome sequencing to compare baseline and posttreatment somatic genomic profiles in circulating tumor DNA (ctDNA). Results: Patient ctDNA abundance was correlated across plasma collections and independently prognostic for sequential therapy response and overall survival. Most driver alterations in established prostate cancer genes were consistently detected in ctDNA over time. However, shifts in somatic populations after treatment were identified in 53% of patients, particularly after strong treatment responses. Treatment-associated changes converged upon the AR gene, with an average 50% increase in AR copy number, changes in AR mutation frequencies, and a 2.5-fold increase in the proportion of patients carrying AR ligand binding domain truncating rearrangements. Conclusions: Our data show that the dominant AR genotype continues to evolve during sequential lines of AR inhibition and drives acquired resistance in patients with mCRPC.

Prostate cancer is heterogeneous and patients would benefit from methods that stratify those who are likely to respond to systemic therapy. Here, we employ single-cell assays for transposase-accessible chromatin (ATAC) and RNA sequencing in models of early treatment response and resistance to enzalutamide. In doing so, we identify pre-existing and treatment-persistent cell subpopulations that possess regenerative potential when subjected to treatment. We find distinct chromatin landscapes associated with enzalutamide treatment and resistance that are linked to alternative transcriptional programs. Transcriptional profiles characteristic of persistent cells are able to stratify the treatment response of patients. Ultimately, we show that defining changes in chromatin and gene expression in single-cell populations from pre-clinical models can reveal as yet unrecognized molecular predictors of treatment response. This suggests that the application of single-cell methods with high analytical resolution in pre-clinical models may powerfully inform clinical decision-making.

The evolutionary progression from primary to metastatic prostate cancer is largely uncharted, and the implications for liquid biopsy are unexplored. We infer detailed reconstructions of tumor phylogenies in ten prostate cancer patients with fatal disease, and investigate them in conjunction with histopathology and tumor DNA extracted from blood and cerebrospinal fluid. Substantial evolution occurs within the prostate, resulting in branching into multiple spatially intermixed lineages. One dominant lineage emerges that initiates and drives systemic metastasis, where polyclonal seeding between sites is common. Routes to metastasis differ between patients, and likely genetic drivers of metastasis distinguish the metastatic lineage from the lineage that remains confined to the prostate within each patient. Body fluids capture features of the dominant lineage, and subclonal expansions that occur in the metastatic phase are non-uniformly represented. Cerebrospinal fluid analysis reveals lineages not detected in blood-borne DNA, suggesting possible clinical utility.

No abstract