Charting differences between tumors and normal tissue is a mainstay of cancer research. However, clonal tumor expansion from complex normal tissue architectures potentially obscures cancer-specific ...events, including divergent epigenetic patterns. Using whole-genome bisulfite sequencing of normal B cell subsets, we observed broad epigenetic programming of selective transcription factor binding sites coincident with the degree of B cell maturation. By comparing normal B cells to malignant B cells from 268 patients with chronic lymphocytic leukemia (CLL), we showed that tumors derive largely from a continuum of maturation states reflected in normal developmental stages. Epigenetic maturation in CLL was associated with an indolent gene expression pattern and increasingly favorable clinical outcomes. We further uncovered that most previously reported tumor-specific methylation events are normally present in non-malignant B cells. Instead, we identified a potential pathogenic role for transcription factor dysregulation in CLL, where excess programming by EGR and NFAT with reduced EBF and AP-1 programming imbalances the normal B cell epigenetic program.
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IJS, NUK, SBMB, UL, UM, UPUK
Despite much evidence on epigenetic abnormalities in cancer, it is currently unclear to what extent epigenetic alterations can be associated with tumors’ clonal genetic origins. Here, we show that ...the prostate intratumor heterogeneity in DNA methylation and copy-number patterns can be explained by a unified evolutionary process. By assaying multiple topographically distinct tumor sites, premalignant lesions, and lymph node metastases within five cases of prostate cancer, we demonstrate that both DNA methylation and copy-number heterogeneity consistently reflect the life history of the tumors. Furthermore, we show cases of genetic or epigenetic convergent evolution and highlight the diversity in the evolutionary origins and aberration spectrum between tumor and metastatic subclones. Importantly, DNA methylation can complement genetic data by serving as a proxy for activity at regulatory domains, as we show through identification of high epigenetic heterogeneity at androgen-receptor-bound enhancers. Epigenome variation thereby expands on the current genome-centric view on tumor heterogeneity.
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•Epigenetic intratumor heterogeneity reflects prostate tumors’ clonal genetic origins•Tumor and metastatic subclones exhibit diverse evolutionary origins•Distinct tumor lineages show convergent evolution of methylation patterns•Degree of methylation variability at a locus depends on genetic and epigenetic context
Several recent studies have examined the clonal origins of tumor populations on the basis of genetic variability. Brocks et al. now show that subclonal lineages also differ in their DNA methylation patterns and that methylation heterogeneity is consistent with the tumors’ genetic history. Furthermore, high methylation variability at enhancers bound by androgen receptor, which are known to play a role in prostate tumorigenesis, suggests that DNA methylation data can serve as a proxy for the functional state of tumor subclones.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The molecular mechanisms underlying the genesis of cholangiocarcinomas (CCs) are poorly understood. Epigenetic changes such as aberrant hypermethylation and subsequent atypical gene expression are ...characteristic features of most human cancers. In CC, data regarding global methylation changes are lacking so far. We performed a genome‐wide analysis for aberrant promoter methylation in human CCs. We profiled 10 intrahepatic and 8 extrahepatic CCs in comparison to non‐neoplastic biliary tissue specimens, using methyl‐CpG immunoprecipitation (MCIp) combined with whole‐genome CpG island arrays. DNA methylation was confirmed by quantitative mass spectrometric analysis and functional relevance of promoter hypermethylation was shown in demethylation experiments of two CC cell lines using 5‐aza‐2′deoxycytidine (DAC) treatment. Immunohistochemical staining of tissue microarrays (TMAs) from 223 biliary tract cancers (BTCs) was used to analyze candidate gene expression at the protein level. Differentially methylated, promoter‐associated regions were nonrandomly distributed and enriched for genes involved in cancer‐related pathways including Wnt, transforming growth factor beta (TGF‐β), and PI3K signaling pathways. In CC cell lines, silencing of genes involved in Wnt signaling, such as SOX17, WNT3A, DKK2, SFRP1, SFRP2, and SFRP4 was reversed after DAC administration. Candidate protein SFRP2 was substantially down‐regulated in neoplastic tissues of all BTC subtypes as compared to normal tissues. A significant inverse correlation of SFRP2 protein expression and pT status was found in BTC patients. Conclusion: We provide a comprehensive analysis to define the genome‐wide methylation landscape of human CC. Several candidate genes of cancer‐relevant signaling pathways were identified, and closer analysis of selected Wnt pathway genes confirmed the relevance of this pathway in CC. The presented global methylation data are the basis for future studies on epigenetic changes in cholangiocarcinogenesis. (Hepatology 2014;59:544–554)
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Although clonal selection by genetic driver aberrations in cancer is well documented, the ability of epigenetic alterations to promote tumor evolution is undefined. We used 450k arrays and ...next-generation sequencing to evaluate intratumor heterogeneity and evolution of DNA methylation and genetic aberrations in chronic lymphocytic leukemia (CLL). CLL cases exhibit vast interpatient differences in intratumor methylation heterogeneity, with genetically clonal cases maintaining low methylation heterogeneity and up to 10% of total CpGs in a monoallelically methylated state. Increasing methylation heterogeneity correlates with advanced genetic subclonal complexity. Selection of novel DNA methylation patterns is observed only in cases that undergo genetic evolution, and independent genetic evolution is uncommon and is restricted to low-risk alterations. These results reveal that although evolution of DNA methylation occurs in high-risk, clinically progressive cases, positive selection of novel methylation patterns entails coevolution of genetic alteration(s) in CLL.
Synovial sarcoma is a rare and highly aggressive subtype of soft tissue sarcoma. The clinical challenge posed by advanced or metastatic synovial sarcoma, marked by limited treatment options and ...suboptimal outcomes, necessitates innovative approaches. The topoisomerase II (Topo II) inhibitor doxorubicin has remained the cornerstone systemic treatment for decades, and there is pressing need for improved therapeutic strategies for these patients. This study highlights the potential to enhance the cytotoxic effects of doxorubicin within well-characterized synovial sarcoma cell lines using the potent and selective DNA-PK inhibitor, peposertib. In vitro investigations unveil a p53-mediated synergistic anti-tumor effect when combining doxorubicin with peposertib. The in vitro findings were substantiated by pronounced anti-tumor effects in mice bearing subcutaneously implanted tumors. A well-tolerated regimen for the combined application was established using both pegylated liposomal doxorubicin (PLD) and unmodified doxorubicin. Notably, the combination of PLD and peposertib displayed enhanced anti-tumor efficacy compared to unmodified doxorubicin at equivalent doses, suggesting an improved therapeutic window-a critical consideration for clinical translation. Efficacy studies in two patient-derived xenograft models of synovial sarcoma, accurately reflecting human metastatic disease, further validate the potential of this combined therapy. These findings align with previous evidence showcasing the synergy between DNA-PK inhibition and Topo II inhibitors in diverse tumor models, including breast and ovarian cancers. Our study extends the potential utility of combined therapy to synovial sarcoma.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Changes in DNA methylation identified by epigenome-wide association studies (EWAS) have been recently linked to increased lung cancer risk. However, the cellular effects of these differentially ...methylated positions (DMPs) are often unclear. Therefore, we investigated top differentially methylated positions identified from an EWAS study. This included a putative regulatory region of NHLRC1. Hypomethylation of this gene was recently linked with decreased survival rates in lung cancer patients. HumanMethylation450 BeadChip array (450K) analysis was performed on 66 lung cancer case-control pairs from the European Prospective Investigation into Cancer and Nutrition Heidelberg lung cancer EWAS (EPIC HD) cohort. DMPs identified in these pre-diagnostic blood samples were then investigated for differential DNA methylation in lung tumor versus adjacent normal lung tissue from The Cancer Genome Atlas (TCGA) and replicated in two independent lung tumor versus adjacent normal tissue replication sets with MassARRAY. The EPIC HD top hypermethylated DMP cg06646708 was found to be a hypomethylated region in multiple data sets of lung tumor versus adjacent normal tissue. Hypomethylation within this region caused increased mRNA transcription of the closest gene NHLRC1 in lung tumors. In functional assays, we demonstrate attenuated proliferation, viability, migration, and invasion upon NHLRC1 knock-down in lung cancer cells. Furthermore, diminished AKT phosphorylation at serine 473 causing expression of pro-apoptotic AKT-repressed genes was detected in these knock-down experiments. In conclusion, this study demonstrates the powerful potential for discovery of novel functional mechanisms in oncogenesis based on EWAS DNA methylation data. NHLRC1 holds promise as a new prognostic biomarker for lung cancer survival and prognosis, as well as a target for novel treatment strategies in lung cancer patients.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Genome-wide association studies (GWAS) have identified SNPs linked with lung cancer risk. Our aim was to discover the genes, non-coding RNAs, and regulatory elements within GWAS-identified risk ...regions that are deregulated in non-small cell lung carcinoma (NSCLC) to identify novel, clinically targetable genes and mechanisms in carcinogenesis. A targeted bisulphite-sequencing approach was used to comprehensively investigate DNA methylation changes occurring within lung cancer risk regions in 17 NSCLC and adjacent normal tissue pairs. We report differences in differentially methylated regions between adenocarcinoma and squamous cell carcinoma. Among the minimal regions found to be differentially methylated in at least 50% of the patients, 7 candidates were replicated in 2 independent cohorts (n = 27 and n = 87) and the potential of 6 as methylation-dependent regulatory elements was confirmed by functional assays. This study contributes to understanding the pathways implicated in lung cancer initiation and progression, and provides new potential targets for cancer treatment.
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BFBNIB, GIS, IJS, KISLJ, NUK, PNG, UL, UM, UPUK
Genetic aberrations in DNA repair genes are linked to cancer, but less is reported about epigenetic regulation of DNA repair and functional consequences. We investigated the intragenic methylation ...loss at the three prime repair exonuclease 2 (TREX2) locus in laryngeal (n = 256) and colorectal cancer cases (n = 95) and in pan-cancer data from The Cancer Genome Atlas (TCGA).
Significant methylation loss at an intragenic site of TREX2 was a frequent trait in both patient cohorts (p = 0.016 and < 0.001, respectively) and in 15 out of 22 TCGA studies. Methylation loss correlated with immunohistochemically staining for TREX2 (p < 0.0001) in laryngeal tumors and improved overall survival of laryngeal cancer patients (p = 0.045). Chromatin immunoprecipitation, demethylation experiments, and reporter gene assays revealed that the region of methylation loss can function as a CCAAT/enhancer binding protein alpha (CEBPA)-responsive enhancer element regulating TREX2 expression.
The data highlight a regulatory role of TREX2 DNA methylation for gene expression which might affect incidence and survival of laryngeal cancer. Altered TREX2 protein levels in tumors may affect drug-induced DNA damage repair and provide new tailored therapies.
Malignancies are characterized by extensive global reprogramming of epigenetic patterns, including gains or losses in DNA methylation and changes to histone marks. Furthermore, high-resolution ...genome-sequencing efforts have discovered a wealth of mutations in genes encoding epigenetic regulators that have roles as 'writers', 'readers' or 'editors' of DNA methylation and/or chromatin states. In this Review, we discuss how these mutations have the potential to deregulate hundreds of targeted genes genome wide. Elucidating these networks of epigenetic factors will provide mechanistic understanding of the interplay between genetic and epigenetic alterations, and will inform novel therapeutic strategies.
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DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Identifying the earliest somatic changes in prostate cancer can give important insights into tumor evolution and aids in stratifying high- from low-risk disease. We integrated whole genome, ...transcriptome and methylome analysis of early-onset prostate cancers (diagnosis ≤55 years). Characterization across 292 prostate cancer genomes revealed age-related genomic alterations and a clock-like enzymatic-driven mutational process contributing to the earliest mutations in prostate cancer patients. Our integrative analysis identified four molecular subgroups, including a particularly aggressive subgroup with recurrent duplications associated with increased expression of ESRP1, which we validate in 12,000 tissue microarray tumors. Finally, we combined the patterns of molecular co-occurrence and risk-based subgroup information to deconvolve the molecular and clinical trajectories of prostate cancer from single patient samples.
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•Clock-like mutation process attributed to APOBEC3 mediates earliest mutations in PC•Identification of four molecular subgroups that stratifies intermediate-risk disease•Rearrangements at the ESRP1 locus associated with aggressive and proliferative cancer•Development of method to predict clinical trajectories of PC from DNA sequencing data
Gerhauser et al. molecularly characterize prostate cancers diagnosed before 56 years old, which reveals an APOBEC-driven mutational process and identifies an aggressive subgroup with increased expression of ESRP1. They develop a framework to predict the order of somatic alterations and clinical outcome.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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