Diagenode® Premium RRBS technology: cost-effective DNA methylation mapping with superior coverage

Veillard, Anne-Clémence; Datlinger, Paul; Laczik, Miklos; Squazzo, Sharon L.; Bock, Christoph

doi:10.1038/nmeth.f.391

Cited by 28 publications

(24 citation statements)

References 7 publications

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“…RRBS was performed as described previously51,52, starting with 100 ng of genomic DNA per sample. Custom-designed methylated and unmethylated oligonucleotides were added at a concentration of 0.1% to serve as spike-in controls for monitoring bisulfite conversion efficiency.…”

Section: Methodsmentioning

confidence: 99%

DNA methylation heterogeneity defines a disease spectrum in Ewing sarcoma

Sheffield

Pierron

Klughammer

et al. 2017

Nat Med

Self Cite

210

216

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Developmental tumors in children and young adults carry few genetic alterations, yet they have diverse clinical presentation. Focusing on Ewing sarcoma, we sought to establish the prevalence and characteristics of epigenetic heterogeneity in genetically homogeneous cancers. We performed genome-scale DNA methylation sequencing for a large cohort of Ewing sarcoma tumors and analyzed epigenetic heterogeneity on three levels: between cancers, between tumors, and within tumors. We observed consistent DNA hypomethylation at enhancers regulated by the disease-defining EWS-FLI1 fusion protein, thus establishing epigenomic enhancer reprogramming as a ubiquitous and characteristic feature of Ewing sarcoma. DNA methylation differences between tumors identified a continuous disease spectrum underlying Ewing sarcoma, which reflected the strength of an EWS-FLI1 regulatory signature and a continuum between mesenchymal and stem cell signatures. There was substantial epigenetic heterogeneity within tumors, particularly in patients with metastatic disease. In summary, our study provides a comprehensive assessment of epigenetic heterogeneity in Ewing sarcoma and thereby highlights the importance of considering nongenetic aspects of tumor heterogeneity in the context of cancer biology and personalized medicine.

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Section: Methodsmentioning

confidence: 99%

DNA methylation heterogeneity defines a disease spectrum in Ewing sarcoma

Sheffield

Pierron

Klughammer

et al. 2017

Nat Med

Self Cite

210

216

View full text Add to dashboard Cite

show abstract

“…RRBS was performed by Diagenode, Inc. as described before [ 64 ]. Sequencing was carried out on an Illumina HiSeq 3000.…”

Section: Methodsmentioning

confidence: 99%

DNMT3B overexpression contributes to aberrant DNA methylation and MYC-driven tumor maintenance in T-ALL and Burkitt’s lymphoma

et al. 2017

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Aberrant DNA methylation is a hallmark of cancer. However, our understanding of how tumor cell-specific DNA methylation patterns are established and maintained is limited. Here, we report that in T-cell acute lymphoblastic leukemia (T-ALL) and Burkitt’s lymphoma the MYC oncogene causes overexpression of DNA methyltransferase (DNMT) 1 and 3B, which contributes to tumor maintenance. By utilizing a tetracycline-regulated MYC transgene in a mouse T-ALL (EμSRα-tTA;tet-o-MYC) and human Burkitt’s lymphoma (P493-6) model, we demonstrated that DNMT1 and DNMT3B expression depend on high MYC levels, and that their transcription decreased upon MYC-inactivation. Chromatin immunoprecipitation indicated that MYC binds to the DNMT1 and DNMT3B promoters, implicating a direct transcriptional regulation. Hence, shRNA-mediated knock-down of endogenous MYC in human T-ALL and Burkitt’s lymphoma cell lines downregulated DNMT3B expression. Knock-down and pharmacologic inhibition of DNMT3B in T-ALL reduced cell proliferation associated with genome-wide changes in DNA methylation, indicating a tumor promoter function during tumor maintenance. We provide novel evidence that MYC directly deregulates the expression of both de novo and maintenance DNMTs, showing that MYC controls DNA methylation in a genome-wide fashion. Our finding that a coordinated interplay between the components of the DNA methylating machinery contributes to MYC-driven tumor maintenance highlights the potential of specific DNMTs for targeted therapies.

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“…DNA methylation profiling was performed on archival formalin-fixed paraffin-embedded (FFPE) samples using reduced representation bisulfite sequencing (RRBS)28–30. Compared to Infinium microarrays, our optimized RRBS protocol yields higher genomic coverage31 and works well on degraded DNA from FFPE samples of variable quality32.…”

Section: Introductionmentioning

confidence: 99%

The DNA methylation landscape of glioblastoma disease progression shows extensive heterogeneity in time and space

Klughammer

Kiesel

Roetzer

et al. 2018

Nat Med

Self Cite

239

199

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Glioblastoma is characterized by widespread genetic and transcriptional heterogeneity, yet little is known about the role of the epigenome in glioblastoma disease progression. Here, we present genome-scale maps of DNA methylation in matched primary and recurring glioblastoma tumors, using data from a highly annotated clinical cohort that was selected through a national patient registry. We demonstrate the feasibility of DNA methylation mapping in a large set of routinely collected FFPE samples, and we validate bisulfite sequencing as a multipurpose assay that allowed us to infer a range of different genetic, epigenetic, and transcriptional characteristics of the profiled tumor samples. On the basis of these data, we identified subtle differences between primary and recurring tumors, links between DNA methylation and the tumor microenvironment, and an association of epigenetic tumor heterogeneity with patient survival. In summary, this study establishes an open resource for dissecting DNA methylation heterogeneity in a genetically diverse and heterogeneous cancer, and it demonstrates the feasibility of integrating epigenomics, radiology, and digital pathology for a national cohort, thereby leveraging existing samples and data collected as part of routine clinical practice.

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Diagenode® Premium RRBS technology: cost-effective DNA methylation mapping with superior coverage

Cited by 28 publications

References 7 publications

DNA methylation heterogeneity defines a disease spectrum in Ewing sarcoma

DNA methylation heterogeneity defines a disease spectrum in Ewing sarcoma

DNMT3B overexpression contributes to aberrant DNA methylation and MYC-driven tumor maintenance in T-ALL and Burkitt’s lymphoma

The DNA methylation landscape of glioblastoma disease progression shows extensive heterogeneity in time and space

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