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Page 316 Meyer et al. Cancer Drug Resist 2019;2:313-25 I http://dx.doi.org/10.20517/cdr.2019.11
[9]
hypermethylation are shared across diverse B-ALLs , analyses of ETV6/RUNX1 and high hyperdiploid
B-ALLs demonstrated widespread hypermethylation of promoter CGIs with significant overlap between
the corresponding methylation profiles. However, despite these commonalities, unsupervised clustering of
these methylome data successfully distinguished ETV6/RUNX1 B-ALLs from high hyperdiploid B-ALLs,
suggesting that, in addition to distinct chromosomal alterations, methylation patterns are defining
features of these molecular subtypes. Interestingly, this study demonstrated that regions associated
with hypermethylation in ETV6/RUNX1 ALLs were relatively less methylated on the tri- and tetrasomic
chromosomes in the high hyperdiploid ALL samples, leading to relative overexpression at the gene level
of the genes at these loci. While the functional significance of these findings remains to be elucidated, the
authors propose that the altered methylation pattern and gene expression profile may play a critical role
[10]
in maintaining cellular fitness during the process of leukemogenesis . In an even more diverse cohort
[11]
encompassing all major B-ALL subtypes as well as T-ALLs, Figueroa et al. performed an integrated
analysis of copy number alterations, gene expression patterns, and methylation changes. Once again, this
analysis demonstrated commonalities between all ALL subtypes, which included epigenetic dysregulation
of genes involved in cell cycle progression and transcriptional regulation, further supporting the idea that
epigenetic changes may be required for leukemogenesis. However, these DNA methylation profiles were
also sufficient to distinguish T-ALL from B-ALL and to reliably subdivide B-ALLs according to molecular
subtype. Furthermore, across these subtypes, the DNA methylation profiles were found to tightly correlate
with gene expression patterns, suggesting that these patterns of aberrant methylation may facilitate the
unique gene expression signatures associated with different ALL subtypes.

Despite the frequent hypermethylation of CGIs at gene promoters in cancer cells, global hypomethylation of
[7]
CpG sites outside of promoter sequences is a common feature of cancer . Consistent with this, comparative
genome-wide methylation profiling of primary B-ALLs and of normal B-cell controls revealed promoter
hypermethylation in the ALL samples but demonstrated that the vast majority of differentially methylated
regions occurred outside of promoter sequences in intergenic and intronic regions, where hypomethylation
was more common. Furthermore, many of these regions were found to correspond to putative enhancer
elements. Correlating the methylation status of these enhancer elements with expression of the associated
genes revealed that enhancer hypomethylation resulted in increased gene expression and that this most
commonly involved genes associated with processes such as lymphocyte activation, cell migration,
[12]
and apoptosis . Taken together, these data suggest that aberrant methylation may play a causal role in
leukemogenesis.

PROGNOSTIC SIGNIFICANCE OF METHYLATION CHANGES
The evidence that aberrant methylation underlies some of the key biological features of ALL subtypes has
stimulated interest in understanding how these methylation patterns are associated with clinical outcomes,
which could, in turn, inform efforts to pharmacologically target aberrant epigenetic processes. In an analysis
[13]
of over 1000 CpG sites in a large number of ALL samples, Milani et al. identified 300 CpG sites with
highly variable degrees of methylation across ALL samples. Differential methylation of these sites was not
only sufficient to distinguish different molecular subtypes of ALL, but also predicted relapse risk in ETV6/
RUNX1 and high hyperdiploid ALLs, two subtypes that are otherwise associated with a good prognosis.
Furthermore, the authors performed univariate regression analysis to identify 22 CpG sites for which
methylation status was tightly correlated with relapse risk, providing support for the use of epigenetic
information for risk stratification. Consistent with these findings, methylation profiling of 29 B-ALL samples
and normal B-cell controls revealed that a subset of B-ALL methylomes clustered with those of normal
B-cells while the remainder clustered separately from the normal controls and were associated with highly
aberrant methylomes. When stratified by outcome, diagnostic leukemias with methylomes that deviated
[14]
more significantly from healthy controls were associated with increased rates of relapse .