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Androgen receptor (AR) drives prostate cancer (PCa) development and progression. AR chromatin binding profiles are highly plastic and form recurrent programmatic changes that differentiate disease stages, subtypes and patient outcomes. While prior studies focused on concordance between patient subgroups, inter-tumor heterogeneity of AR enhancer selectivity remains unexplored. Here we report high levels of AR chromatin binding heterogeneity in human primary prostate tumors, that overlap with heterogeneity observed in healthy prostate epithelium. Such heterogeneity has functional consequences, as somatic mutations converge on commonly-shared AR sites in primary over metastatic tissues. In contrast, less-frequently shared AR sites associate strongly with AR-driven gene expression, while such heterogeneous AR enhancer usage also distinguishes patients’ outcome. These findings indicate that epigenetic heterogeneity in primary disease is directly informative for risk of biochemical relapse. Cumulatively, our results illustrate a high level of AR enhancer heterogeneity in primary PCa driving differential expression and clinical impact., We would like to acknowledge the NKI Genomics Core Facility for Illumina sequencing and bioinformatics support and the NKI Research High-Performance Computing (RHPC) facility for computational infrastructure. We express gratitude to all members of the Zwart and Bergman lab, and members of the NKI Oncogenomics division for helpful scientific discussion. This work was supported by the Prostate Cancer Foundation (21CHAL04), Department of Defense (W81XWH-21-1-0234, W81XWH-19-1-0565), Oncode Institute and Alpe d’HuZes/KWF Dutch Cancer Society (10084).

Muscle-invasive bladder cancer (MIBC) is a heterogeneous disease that often recurs despite aggressive treatment with neoadjuvant chemotherapy and (radical) cystectomy. Basal and luminal molecular subtypes have been identified that are linked to clinical characteristics and have differential sensitivities to chemotherapy. While it has been suggested that epigenetic mechanisms play a role in defining these subtypes, a thorough understanding of the biological mechanisms is lacking. This report details the first genome-wide analysis of histone methylation patterns of human primary bladder tumours by chromatin immunoprecipitations and next-generation sequencing (ChIP-seq). We profiled multiple histone marks: H3K27me3, a marker for repressed genes, and H3K4me1 and H3K4me3, which are indicators of active enhancers and active promoters. Integrated analysis of ChIP-seq data and RNA sequencing revealed that H3K4 mono-methylation demarcates MIBC subtypes, while no association was found for the other two histone modifications in relation to basal and luminal subtypes. Additionally, we identified differentially methylated H3K4me1 peaks in basal and luminal tumour samples, suggesting that active enhancers play a role in defining subtypes. Our study is the first analysis of histone modifications in primary bladder cancer tissue and provides an important resource for the bladder cancer community.

The Androgen Receptor (AR) is the key-driving transcription factor in prostate cancer, tightly controlled by epigenetic regulation. To date, most epigenetic profiling has been performed in cell lines or limited tissue samples. Here, to comprehensively study the epigenetic landscape, we perform RNA-seq with ChIP-seq for AR and histone modification marks (H3K27ac, H3K4me3, H3K27me3) in 100 primary prostate carcinomas. Integrative molecular subtyping of the five data streams revealed three major subtypes of which two were clearly TMPRSS2-ERG dictated. Importantly, we identify a third subtype with low chromatin binding and activity of AR, but with high activity of FGF and WNT signaling. While positive for neuroendocrine-hallmark genes, these tumors were copy number-neutral with low mutational burden, significantly depleted for genes characteristic of poor-outcome associated luminal B-subtype. We present a unique resource on transcriptional and epigenetic control in prostate cancer, revealing tight control of gene regulation differentially dictated by AR over three subtypes., The Androgen Receptor (AR) is the main driver of prostate cancer and functions in conjunction with chromatin modifications. Here, the authors comprehensively profile 100 primary prostate carcinomas by sequencing RNA transcripts in combination with ChIP-sequencing for AR and the active histone marks H3K27ac, H3K4me3 and repressive mark H3K27me3.

Although ductal carcinoma in situ (DCIS) is a non-obligate precursor to ipsilateral invasive breast cancer (iIBC), most DCIS lesions remain indolent. Hence, overdiagnosis and overtreatment of DCIS is a major concern. There is an urgent need for prognostic markers that can distinguish harmless from potentially hazardous DCIS. We hypothesised that features of the breast adipose tissue may be associated with risk of subsequent iIBC. We performed a case–control study nested in a population-based DCIS cohort, consisting of 2658 women diagnosed with primary DCIS between 1989 and 2005, uniformly treated with breast conserving surgery (BCS) alone. We assessed breast adipose features with digital pathology (HALO®, Indica Labs) and related these to iIBC risk in 108 women that developed subsequent iIBC (cases) and 168 women who did not (controls) by conditional logistic regression, accounting for clinicopathological and immunohistochemistry variables. Large breast adipocyte size was significantly associated with iIBC risk (odds ratio (OR) 2.75, 95% confidence interval (95% CI) = 1.25–6.05). High cyclooxygenase (COX)-2 protein expression in the DCIS cells was also associated with subsequent iIBC (OR 3.70 (95% CI = 1.59–8.64). DCIS with both high COX-2 expression and large breast adipocytes was associated with a 12-fold higher risk (OR 12.0, 95% CI = 3.10–46.3, P

Systematic identification of noncoding regulatory elements has, to date, mainly relied on large-scale reporter assays that do not reproduce endogenous conditions. We present two distinct CRISPR-Cas9 genetic screens to identify and characterize functional enhancers in their native context. Our strategy is to target Cas9 to transcription factor binding sites in enhancer regions. We identified several functional enhancer elements and characterized the role of two of them in mediating p53 (TP53) and ERα (ESR1) gene regulation. Moreover, we show that a genomic CRISPR-Cas9 tiling screen can precisely map functional domains within enhancer elements. Our approach expands the utility of CRISPR-Cas9 to elucidate the functions of the noncoding genome.

The complexity of oestrogen receptor α (ERα)-mediated transcription is becoming apparent, but global insight into the co-regulatory proteins that assist ERα transcription is incomplete. Here, we present the most comprehensive chromatin-binding landscape of ERα co-regulatory proteins to date. We map by ChIP-seq the essential p160 co-regulators (SRC1, SRC2 and SRC3), and the histone acetyl transferases p300 and CBP in MCF-7 breast cancer cells. We find a complex network of co-regulator binding, with preferential binding sites for each co-regulator. Unlike previous suggestions, we find SRC recruitment almost exclusively following ligand treatment. Interestingly, we find specific subsets of genes regulated by ligand-dependent and -independent co-regulator recruitment. Co-factor-binding profiles were integrated with expression data from cell lines and primary tumour cohorts, to reveal specific transcriptional networks that influence clinical outcome. Genes that are bound by SRC3, but not other p160 proteins, have predictive value in cohorts of breast cancer patients. By generating a robust and global view of co-factor-binding properties, we discover new levels of co-regulator complexity, but also reveal specific gene networks that may influence endocrine response.