Techniques used for detection of the 5-hmC
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5397694/pdf/13072_2017_Article_123.pdf
very imp. : Regulation and Functional Significance of 5-Hydroxymethylcytosine in Cancer
Interplay Between the Cancer Genome and Epigenome
Disruption of Epigenetic Control in Cancer Most studies of cancer epigenetics have focused on DNA methylation, as the epigenetic mark that most easily survives various forms of sample processing, including DNA extraction, and even formalin fixation and paraffin embedding (Laird, 2010). However, other epigenetic marks also undergo broad changes, including long non-coding RNAs and miRNAs (Baer et al., 2013; Baylin and Jones, 2011; Dawson and Kouzarides, 2012; Sandoval and Esteller, 2012), and loss of K16 acetylation and K20 trimethylation at histone H4 (Fraga et al., 2005; Hon et al., 2012; Kondo et al., 2008; Seligson et al., 2005; Yamazaki et al., 2013). Loss of 5-methylcytosine in cancer cells was discussed more than three decades ago (Ehrlich and Wang, 1981), with global DNA hypomethylation reported in cancer cell lines (Diala and Hoffman, 1982; Ehrlich et al., 1982) and reduced levels of DNA methylation found at selected genes in primary human tumors compared to normal tissues (Feinberg and Vogelstein, 1983). The widespread loss of DNA methylation contrasted starkly with the subsequent finding of hypermethylation of CpG islands in cancer (Baylin et al., 1986), including of promoter CpG islands of tumor-suppressor genes (Jones and Baylin, 2002). These seemingly contradictory findings have been widely reported for many types of cancer (Baylin and Jones, 2011). The causal relevance of epigenetic changes in cancer was initially questioned but this concern has now largely been laid to rest. First, many known tumor-suppressor genes have been shown to be silenced by promoter CpG island hypermethylation (Jones and Baylin, 2002). Importantly, the finding that these silencing events are mutually exclusive with structural or mutational inactivation of the same gene, such as the case for BRCA1 in ovarian cancer (TCGA, 2011) and for CDKN2A in squamous cell lung cancer (TCGA, 2012a), reinforces the concept that epigenetic silencing can serve as an alternative mechanism in Knudson's two-hit hypothesis (Jones and Laird, 1999). Second, mouse models of cancer have been shown to require epigenetic writers and readers for tumor development (Laird et al., 1995; Prokhortchouk et al., 2006; Sansom et al., 2003). Third, some DNA methylation changes appear to be essential for cancer cell survival, suggesting an acquired addiction to epigenetic alterations (De Carvalho et al., 2012). Finally, a plethora of significantly mutated epigenetic regulators has now been reported for many types of human cancer, as discussed further below.
5-Hydroxymethylcytosine (5hmC), or How to Identify Your Favorite Cell
Recently described as the sixth base of the DNA macromolecule, the precise role of5-hydroxymethylcytosine (5hmC) is the subject of debate. Early studies indicate that it is functionally distinct from cytosine DNA methylation (5mC), and there is evidence for 5hmC being a stable derivate of 5mC, rather than just an intermediate of demethylation. Because of the growing understanding of the role of progenitor cells in disease origin, we attempted to provide a detailed summary on the currently available literature supporting 5hmC as a key player in adult progenitor cell differentiation.

5-hydroxymethylcytosine and its potential roles in development and cancer

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3645968/
Only a few years ago it was demonstrated that mammalian DNA contains oxidized forms of 5-methylcytosine (5mC). The base 5-hydroxymethylcytosine (5hmC) is the most abundant of these oxidation products and is referred to as the sixth DNA base. 5hmC is produced from 5mC in an enzymatic pathway involving three 5mC oxidases, Ten-eleven translocation (TET)1, TET2, and TET3. The biological role of 5hmC is still unclear. Current models propose that 5hmC is an intermediate base in an active or passive DNA demethylation process that operates during important reprogramming phases of mammalian development. Tumors originating in various human tissues have strongly depleted levels of 5hmC. Apparently, 5hmC cannot be maintained in proliferating cells. Furthermore, mutations in the TET2 gene are commonly observed in human myeloid malignancies. Since TET proteins and many lysine demethylases require 2-oxoglutarate as a cofactor, aberrations in cofactor biochemical pathways, including mutations in isocitrate dehydrogenase (IDH), may affect levels of 5hmC and 5mC in certain types of tumors, either directly or indirectly. We discuss current data and models of the function of 5hmC in general, with special emphasis on its role in mechanisms of development and cancer.
5-methylcytosine (5mC) is created in a postreplicative enzymatic reaction in which a DNA methyltransferase enzyme transfers a methyl group from S-adenosylmethionine onto the 5-carbon of cytosine, mostly within the CpG sequence context [1]. Presence of 5mC at gene promoters is most often linked to transcriptional repression [2]. It was long thought that 5mC was the only modified base in animal DNA. 5-hydroxymethylcytosine (5hmC) was initially found in the DNA of certain bacteriophages [3] and was reported in mammalian tissues as early as 1972 [4]. However, the levels reported by Penn et al. [4] seemed too high and could not be confirmed in subsequent studies [5]. The earlier report by Penn et al. [4] had put the levels of 5hmC in brain and liver DNA at 15% of the level of cytosine, which is at least an order of magnitude higher than currently established levels for brain and around two orders of magnitude higher than levels found in liver DNA [6]. Also, in the same study, 5mC was not detected casting doubt on these earlier results.