Genome-wide DNA Methylation Events in <i>TMPRSS2–ERG</i> Fusion-Negative Prostate Cancers Implicate an EZH2-Dependent Mechanism with <i>miR-26a</i> Hypermethylation

Börno, Stefan T.; Fischer, Axel; Kerick, Martin; Fälth, Maria; Laible, Mark; Brase, Jan C.; Kuner, Ruprecht; Dahl, Andreas; Grimm, Christina; Sayanjali, Behnam; Isau, Melanie; Röhr, Christina; Wunderlich, Andrea; Timmermann, Bernd; Claus, Rainer; Plass, Christoph; Graefen, Markus; Simon, Ronald; Demichelis, Francesca; Rubin, Mark A.; Sauter, Guido; Schlomm, Thorsten; Sültmann, Holger; Lehrach, Hans; Schweiger, Michal-Ruth

doi:10.1158/2159-8290.cd-12-0041

Cited by 127 publications

(108 citation statements)

References 55 publications

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“…6C; χ 2 =13.66; P=0.008). Thus, in corroboration to previous reports (4,14), our data suggests a more pronounced role of epigenetic alterations in ETS-fusion negative cases. Although, 6 of the 50 SPINK1+ cases failed to show any expression of EZH2, pointing that an alternative mechanism may be involved in SPINK1 regulation or possibly miRNA-338/-421 genomic deletion could be a cause in such cases.…”

supporting

confidence: 92%

“…Recently, a new subtype of ETS-fusion-negative tumors has been defined by frequent mutations in the epigenetic regulators and chromatin remodelers (55). Yet another study, using genome-wide methylated DNAimmunoprecipitation sequencing revealed higher number of methylation events in TMPRSS2-ERG fusion-negative as compared to normal and TMPRSS2-ERG fusion-positive PCa specimens (14), thus collectively, these independent findings reaffirm the critical role of epigenetic pathways engaged in the pathogenesis of SPINK1+ subtype.…”

Section: Discussionmentioning

confidence: 66%

“…Interestingly, increased methylated regions in the ETS-fusion negative patients have been attributed to hypermethylation of miR-26a, a post-transcriptional regulator of EZH2 (14).…”

Section: Discussionmentioning

confidence: 99%

“…Earlier studies have shown that aberrant expression of Enhancer of Zeste Homolog 2 (EZH2) owing to genomic loss of miRNA-101 (13) or hypermethylation of miR-26a (14) 7 respectively) show a significant reduction in SPINK1 expression at both transcript (~80-90%) and protein (Fig. 1G) levels.…”

Section: Introductionmentioning

confidence: 99%

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Epigenetic silencing of miRNA-338-5p and miRNA-421 drives SPINK1-positive prostate cancer

Bhatia

Yadav

Tiwari

et al. 2018

Preprint

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PurposeSerine Peptidase Inhibitor, Kazal type-1 (SPINK1) overexpression defines the second most recurrent and aggressive prostate cancer (PCa) subtype. However, the underlying molecular mechanism and pathobiology of SPINK1 in PCa remains largely unknown.Experimental DesignMicroRNA-prediction tools were employed to examine the SPINK1-3’UTR for miRNAs binding. Luciferase reporter assays were performed to confirm the SPINK1-3’UTR binding of shortlisted miR-338-5p/miR-421. Further, miR-338-5p/-421 overexpressing cancer cells (SPINK1-positive) were evaluated for oncogenic properties using cell-based functional assays and mice xenograft model. Global gene expression profiling was performed to unravel the biological pathways altered by miR-338-5p/-421. Immunohistochemistry and RNA in-situ hybridization was carried-out on PCa patients’ tissue microarray for SPINK1 and EZH2 expression respectively. Chromatin immunoprecipitation assay was performed to examine EZH2 occupancy on the miR-338-5p/-421 regulatory regions. Bisulfite sequencing and methylated DNA-immunoprecipitation was performed on PCa cell lines and patients’ specimens.ResultsWe established a critical role of miRNA-338-5p/-421 in post-transcriptional regulation of SPINK1. Ectopic expression of miRNA-338-5p/-421 in SPINK1-positive PCa cells abrogate oncogenic properties including cell-cycle progression, stemness and drug resistance, and show reduced tumor burden and distant metastases in mice model. Importantly, we show SPINK1-positive PCa patients exhibit increased EZH2 expression, suggesting its role in miRNA-338-5p/-421 epigenetic silencing. Furthermore, presence of CpG dinucleotide DNA methylation marks on the regulatory regions of miR-338-5p/-421 in SPINK1-positive PCa cells and patients’ specimens confirms epigenetic silencing.ConclusionOur findings revealed that miRNA-338-5p/-421 are epigenetically silenced in SPINK1-positive PCa, while restoring the expression of these miRNAs using epigenetic drugs or synthetic mimics could abrogate SPINK1-mediated oncogenesis.TRANSLATIONAL IMPACTWe establish a regulatory model involving the functional interplay between SPINK1, miRNA-338-5p/miRNA-421 and EZH2, thereby, revealing hitherto unknown mechanism of SPINK1 up-regulation in SPINK1-positive subtype. Our findings provide a strong rationale for the development of potential therapeutic strategies for SPINK1-positive malignancies. We demonstrate that restoring miRNA-338-5p/miRNA-421 expression using epigenetic drugs including DNMTs inhibitors in combination with HDACs or HKMTs inhibitors or miRNA synthetic mimics in SPINK1-positive prostate cancer abrogate SPINK1-mediated oncogenicity. The major findings of this study will not only advance the prostate cancer field, but will also be valuable for treatment and disease management of other SPINK1-positive malignancies.

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supporting

confidence: 92%

Section: Discussionmentioning

confidence: 66%

“…Interestingly, increased methylated regions in the ETS-fusion negative patients have been attributed to hypermethylation of miR-26a, a post-transcriptional regulator of EZH2 (14).…”

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Epigenetic silencing of miRNA-338-5p and miRNA-421 drives SPINK1-positive prostate cancer

Bhatia

Yadav

Tiwari

et al. 2018

Preprint

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“…Using high-throughput genome sequencing and DNA methylation analyses, .147 000 cancer-associated epigenetic alterations were observed in 51 tumour and 53 benign prostate samples; the specific nature of these alterations were dependent on the presence of absence of a TMPRSS2-ERG rearrangement. 52 This observation of differential methylation events in fusion-negative tumours (based on enhancer of zeste homologue 2 (EZH2) gene activation) explains CaP carcinogenesis in the 50% of patients who are TMPRSS2-ERG fusion-negative. Systematic overviews of studies for the prognostic role of specific gene methylation (e.g.…”

Section: Genetic Instability and Genomic Assays In Localized Prostatementioning

confidence: 99%

An arranged marriage for precision medicine: hypoxia and genomic assays in localized prostate cancer radiotherapy

Bristow

Berlin²,

Pra

2014

BJR

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Prostate cancer (CaP) is the most commonly diagnosed malignancy in males in the Western world with one in six males diagnosed in their lifetime. Current clinical prognostication groupings use pathologic Gleason score, pre-treatment prostatic-specific antigen and Union for International Cancer Control-TNM staging to place patients with localized CaP into low-, intermediate-and high-risk categories. These categories represent an increasing risk of biochemical failure and CaP-specific mortality rates, they also reflect the need for increasing treatment intensity and justification for increased side effects. In this article, we point out that 30-50% of patients will still fail image-guided radiotherapy or surgery despite the judicious use of clinical risk categories owing to interpatient heterogeneity in treatment response. To improve treatment individualization, better predictors of prognosis and radiotherapy treatment response are needed to triage patients to bespoke and intensified CaP treatment protocols. These should include the use of pre-treatment genomic tests based on DNA or RNA indices and/or assays that reflect cancer metabolism, such as hypoxia assays, to define patient-specific CaP progression and aggression. More importantly, it is argued that these novel prognostic assays could be even more useful if combined together to drive forward precision cancer medicine for localized CaP.

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Determination of ERG(+), EZH2, NKX3.1, and SPINK‐1 subtypes to evaluate their association with clonal origin and disease progression in multifocal prostate cancer

et al. 2022

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cause death. The criteria to establish PCa prognosis include PSA level, Gleason score, and clinical TNM stage, but these criteria still do not establish the prognosis with su cient certainty (2).Many studies have established different classi cations of PCa in molecular subtypes that can contribute to determining the prognosis, among them ETS(+) subtypes, characterized by chromosomal fusions, which cause the overexpression of ETS transcription factors. The most prevalent ETS fusion in PCa is TMPRSS2-ERG, which leads to ERG overexpression (3-5). ERG overexpression leads to an increase in the expression of EZH2, which acts as a histone methyltransferase enzyme, as well as to a decrease in the expression of the tumor suppressor gene NKX3.1, which is an androgen-regulated prostate-speci c homeobox gene and a transcription factor. NKX3.1 also binds to TMPRSS2 upstream sequences, and in TMPRSS2-ERG cases, it negatively regulates ERG expression and constitutes a negative feedback control since ERG in turn directly represses NKX3.1 (6,7). EZH2 and NKX3.1 had also been described as PCa subtypes. Another subtype reported in PCa is characterized by SPINK-1 overexpression, which is speci cally overexpressed in a subset of ETS(-) and is associated with another subtype, the SPOP subtype, characterized by a mutation in this gene (3). SPINK-1 protein is a trypsin inhibitor, which can function as an autocrine growth factor (8,9), and SPOP is an E3 ubiquitin ligase substrate-binding subunit of the proteasome complex that mediates the ubiquitination of target proteins, leading more frequently to their proteasomal degradation (10).The prognostic relevance of PCa molecular subtypes remains controversial (11)(12)(13)(14)(15)(16)(17)(18)(19)(20). The presence of multiple foci -with the possibility of different subtypes-in the same patient with PCa could contribute to the di culty of predicting the clinical behavior of a speci c PCa subtype (21-23). It is not well known whether these foci have a monoclonal origin that expands through the prostate, giving rise to new foci, and share some molecular characteristics by origin, which begins to acquire changes in subclonal events; or whether each focus appears independently, so they have a multiclonal origin and they are expected to be very heterogeneous at the molecular level (22,24,25).To help elucidate the origin of heterogeneity in PCa and its association with disease progression, ERG(+), SPOP, EZH2, SPINK-1, and NKX3.1 subtypes were analyzed in 83 samples from prostatic foci and regional metastasis to lymph nodes (LN) from 20 PCa patients with poor prognosis. We also established molecular patterns in samples and patients, combining the presence of all subtypes analyzed. Methods SamplesSamples from formalin-xed para n-embedded (FFPE) tissues obtained from radical prostatectomy (RP) of 20 PCa patients with poor prognosis (regional metastases and/or biochemical recurrence (BCR)) and 5-years of follow-up were included. Cases were selected from the project "Exploration of potential predictor b...

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Genome-wide DNA Methylation Events in TMPRSS2–ERG Fusion-Negative Prostate Cancers Implicate an EZH2-Dependent Mechanism with miR-26a Hypermethylation

Cited by 127 publications

References 55 publications

Epigenetic silencing of miRNA-338-5p and miRNA-421 drives SPINK1-positive prostate cancer

Epigenetic silencing of miRNA-338-5p and miRNA-421 drives SPINK1-positive prostate cancer

An arranged marriage for precision medicine: hypoxia and genomic assays in localized prostate cancer radiotherapy

Determination of ERG(+), EZH2, NKX3.1, and SPINK‐1 subtypes to evaluate their association with clonal origin and disease progression in multifocal prostate cancer

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