Technical advances in global DNA methylation analysis in human cancers

Abstract

Prototypical abnormalities of genome-wide DNA methylation constitute the most widely investigated epigenetic mechanism in human cancers. Errors in the cellular machinery to faithfully replicate the global 5-methylcytosine (5mC) patterns, commonly observed during tumorigenesis, give rise to misregulated biological pathways beneficial to the rapidly propagating tumor mass but deleterious to the healthy tissues of the affected individual. A growing body of evidence suggests that the global DNA methylation levels could serve as utilitarian biomarkers in certain cancer types. Important breakthroughs in the recent years have uncovered further oxidized derivatives of 5mC - 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC), thereby expanding our understanding of the DNA methylation dynamics. While the biological roles of these epigenetic derivatives are being extensively characterized, this review presents a perspective on the opportunity of innovation in the global methylation analysis platforms. While multiple methods for global analysis of 5mC in clinical samples exist and have been reviewed elsewhere, two of the established methods - Liquid Chromatography coupled with mass spectrometry (LC-MS/MS) and Immunoquantification have successfully evolved to include the quantitation of 5hmC, 5fC and 5caC. Although the analytical performance of LC-MS/MS is superior, the simplicity afforded by the experimental procedure of immunoquantitation ensures it’s near ubiquity in clinical applications. Recent developments in spectroscopy, nanotechnology and sequencing also provide immense promise for future evaluations and are discussed briefly. Finally, we provide a perspective on the current scenario of global DNA methylation analysis tools and present suggestions to develop the next generation toolset.

The Warburg Effect in Diabetic Kidney Disease

“…Mouse models of DKD and in vitro studies also are warranted to confirm and validate the human data. Furthermore, with the development of high sensitive technologies such as global DNA methylations, 80 singular-cell RNA sequencing, 81 RNA bar-coding based on single-molecule fluorescence in situ hybridization, 82 MALDI-MSI, 33 laser capture micro-dissection -omics, 83 and cytometry by time of flight, 84 it becomes feasible to measure multiomics (ie, genomics, transcriptomics, proteomics, and metabolomics/lipidomics) data in a single-cell or cell type. In the near future, we could link pathologic changes with multi-omics in glomeruli, podocytes, and tubules of DKD, which will allow in-depth understanding of DKD and pave the path for personalized kidney precision medicine.…”

The clinical values of dysregulated DNA methylation and demethylation intermediates in acute lymphoblastic leukemia

“…This method is highly sensitive for quantitative analysis, but it is very complicated to perform and requires specific expertise and equipment. Besides HPLC-MS/MS, antibody-based immunochemical analysis, such as ELISA, immunofluorescence and immunohistochemistry (IHC), has been proved as a powerful tool for studying global DNA methylation and demethylation intermediates [31]. In this study, we used ELISA assay, which has been described before, to detect global DNA methylation and demethylation intermediates, and the detection accuracy was acceptable for the quantification of 5-mC, 5-hmC and 5-fC.…”

Clinical Significance of Measuring Global Hydroxymethylation of White Blood Cell DNA in Prostate Cancer: Comparison to PSA in a Pilot Exploratory Study

“…The magnitude of these changes is in contrast with the relatively small differences in median 5hmc levels of WBC DNA that are observed in the present study between PCa and controls. These discrepancies are highly unlikely to originate only from methodological issues [ 46 , 47 ] and inter-tissue differences in total 5hmC content [ 38 , 48 ]; rather, these differences may reflect the fact that the applicability of global DNA hydroxymethylation for PCa diagnosis is tissue-specific.…”