Review

published: 29 January 2016

doi: 10.3389/fonc.2016.00016

Genetics and epigenetics of

Glioblastoma: Applications and
Overall incidence of iDH1 Mutation
Aizhen Liu1, Chunfeng Hou2, Hongfang Chen1, Xuan Zong3* and Peijun Zong1
1
Department of Oncology, Yidu Central Hospital, Jinan, China, 2Department of Oncology Nursing, Yidu Central Hospital,
Jinan, China, 3Department of Oncology, Shandong University School of Medicine, Jinan, China

Glioblastoma is the most fatal brain cancer found in humans. Patients suffering from
glioblastoma have a dismal prognosis, with a median survival of 15months. The tumor
may develop rapidly de novo in older patients or through progression from anaplastic
astrocytomas in younger patients if glioblastoma is primary or secondary, respectively.
During the past decade, significant advances have been made in the understanding of
processes leading to glioblastoma, and several important genetic defects that appear
to be important for the development and progression of this tumor have been identified.
Particularly, the discovery of recurrent mutations in the isocitrate dehydrogenase 1
(IDH1) gene has shed new light on the molecular landscape in glioblastoma. Indeed,
Edited by:
Kerrie Leanne McDonald, emerging research on the consequences of mutant IDH1 protein expression suggests
UNSW Australia, Australia that its neomorphic enzymatic activity catalyzing the production of the oncometabolite
Reviewed by: 2-hydroxyglutarate influences a range of cellular programs that affect the epigenome
Keith Giles,
New York University School of
and contribute to glioblastoma development. One of the exciting observations is the
Medicine, USA presence of IDH1 mutation in the vast majority of secondary glioblastoma, while it is
Kamalakannan Palanichamy,
almost absent in primary glioblastoma. Growing data indicate that this particular muta-
The Ohio State University Medical
Center, USA tion has clinical and prognostic importance and will become a critical early distinction in
*Correspondence: diagnosis of glioblastoma.
Xuan Zong
[email protected] Keywords: glioblastoma, genetics, epigenetics, IDH1 mutation

Specialty section:
This article was submitted to INTRODUCTION
Neuro-Oncology,
a section of the journal Glioblastoma is the most common brain malignancy and one of the most aggressive cancers found
Frontiers in Oncology in humans. It is divided into primary and secondary types. Both are histologically identical, so clini-
Received: 14November2015 cal features are used to distinguish them. Primary glioblastoma manifests rapidly de novo without
Accepted: 16January2016 recognizable precursor lesions, and is by far the more common, accounting for 80% of cases. It
Published: 29January2016 predominates in elderly patients and is characterized by rapid progression and short survival time.
Citation: Secondary glioblastoma evolves from lower-grade gliomas, such as grade II diffuse astrocytoma or
LiuA, HouC, ChenH, ZongX and grade II anaplastic astrocytoma, and is typically seen in younger patients. It can be diagnosed with
ZongP (2016) Genetics and
clinical or histological evidence (1). Infiltrating glioblastomas are incurable with current treatment
Epigenetics of Glioblastoma:
Applications and Overall Incidence of
modalities that include surgery, radiation, and chemotherapy. For newly diagnosed patients, the
IDH1 Mutation. standard treatment is total removal, if possible, followed by the combination of chemotherapy and
Front. Oncol. 6:16. radiotherapy (2, 3). However, despite this maximum treatment, the prognosis of patients remains
doi: 10.3389/fonc.2016.00016 dismal with a median survival of 15months only (2). When compared with other malignancies, there

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Liu et al. Genetics and Epigenetics of Glioblastoma

have only been small improvements in the prognosis of glioblas- TABLE 1 | Genes commonly mutated in glioblastoma.
toma patients over recent decades. Nevertheless, understanding Gene Gene name Function of encoded Point of
of the molecular pathogenesis of glioblastoma has greatly symbol protein mutation (%)
increased and is beginning to match that of other types of cancers.
EGFR Epidermal growth Regulator of cell signaling 1415
Several important genetic alterations have been known for some factor receptor involved in cell proliferation
time, but new technologies have allowed much deep genetic and and survival
epigenetic analysis of large numbers of glioblastoma samples, ERBB2 V-erb-b2-erythroblastic Regulator of cell signaling 07
leading to a number of novel discoveries. One of the most exciting leukemia viral involved in cell proliferation
and clinically relevant observations was the discovery that a high oncogene homolog 2 and survival
percentage of secondary glioblastomas and a very small percent- IDH1 Isocitrate NADPH production 1220
age of primary glioblastoma harbor mutations in the isocitrate dehydrogenase 1
dehydrogenase 1 (IDH1) gene. This stunning and unexpected NF1 Neurofibromin 1 Regulator of cell signaling 1517
discovery that holds clear clinical implications has led to new involved in cell proliferation
and survival
insights into glioblastoma biology. Indeed, IDH1 mutation results
in gain of function to catalyze the production of hydroxyglutarate PIK3CA Phosphoinositide-3- Regulator of cell signaling 710
kinase catalytic alpha involved in cell proliferation
(2-HG) (4), a possible oncometabolite that is thought to influence and survival
a range of cellular programs involved in epigenetic control and
PIK3R1 Phosphoinositide-3- Regulator of cell signaling 78
various processes leading to tumor development. Here, we review kinase regulatory 1 involved in cell proliferation
translational applications of this mutation as well as its incidence and survival
in glioblastoma and other cancers. In addition, we highlight PTEN Phosphatase and Regulator of cell signaling 2437
importance of epigenetic changes that may need to be considered tensin homolog involved in cell proliferation
in future diagnostics and therapy for glioblastoma. Initially, the and survival
review presents a brief overview on mutations commonly found PTPRD Protein tyrosine Regulator of cell signaling 06
in glioblastoma, and prunes the consequences of IDH1 mutation phosphatase receptor involved in cell proliferation
type D and survival
on glioblastoma biology to better understand the potential role of
RB1 Retinoblastoma 1 Regulator of cell cycle 813
this particular mutation in the development of this tumor.
TP53 Tumor protein p53 Apoptosis 3138

COMMON MUTATIONS IN
(5,6). They are found only sporadically in primary glioblastoma
GLIOBLASTOMA (5, 6). Because patients with IDH1-mutated primary glioblastoma
Although glioblastoma-specific mutations are seen, mutations are generally younger and have longer median survival and wild-
in common cancer genes, such as TP53 and PTEN, are very type EGFR, which are characteristics of secondary glioblastoma,
frequent in glioblastomas, but are not of prognostic importance it is hypothesized that these are in fact secondary glioblastomas
(Table1) (5, 6). EGFR point mutations have also been identified for which no histological evidence of evolution from a less malig-
in glioblastoma. The EGFRvIII mutant lacks 267 amino acids in nant glioma is found. Therefore, IDH1 mutation could be used to
the extracellular part, resulting in a constitutively activated recep- differentiate primary from secondary glioblastoma.
tor that no longer requires its ligand EGF to signal downstream
(5). Although mutations in certain cancer genes, such as BRAF BIOLOGICAL CONSEQUENCES OF IDH1
and the RAS genes, have rarely been observed in glioblastoma, MUTATION
inactivating mutations and deletions have been identified in Isocitrate dehydrogenase 1 is a metabolic enzyme that converts
their inhibitory tumor-suppressor gene NF1 (5). Mutations in isocitrate into -ketoglutarate (-KG) via oxidative decarboxyla-
PIK3CA and PIK3R1 genes, coding, respectively, for the PI3K tion using NADP+ as an electron acceptor, and producing NADPH
catalytic subunit p110 and regulatory subunit P85, have also (7). While NADPH is thought to be important for limiting cellular
been described (5, 6). oxidative damage and for lipid biosynthesis, -KG is considered
An interesting gene found to contain mutations in glioblas- as an essential intermediate in the Krebs cycle. Under hypoxia
toma is IDH1, which encodes IDH1 and is involved in energy conditions, IDH1 catalyzes the inverse reaction, converting -KG
metabolism. This gene shows differential expression between to isocitrate which can in turn be converted to acetyl-CoA for
primary and secondary glioblastoma, while PTEN loss, EGFR lipid metabolism and many biochemical reactions.
amplification, and loss of heterozygosity (LOH) of chromosome Since 2008, with the discovery of recurrent mutations in the
10 are associated with primary glioblastoma, and ATRX muta- IDH1 coding gene by the Vogelstein group analyzing the DNA
tion, loss of p53, and LOH of chromosome 19 are common in sequence of the glioblastoma genome (6), substantial progress has
secondary glioblastoma. The IDH1 mutation predicts secondary been made to understand how such genetic modification leads
glioblastoma better than these other mutations predict their IDH1 to play a role in the tumorigenesis. The major finding was
respective glioblastoma subtype. Indeed, IDH1 mutations have the discovery that the IDH1 mutation is a gain-of-function muta-
been predominantly identified in secondary glioblastoma and tion, conferring neomorphic activity to IDH1. Initially, a pivotal
low-grade gliomas, with mutations in more than 70% of cases study profiling IDH1 wild-type and mutant glioblastoma cells

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Liu et al. Genetics and Epigenetics of Glioblastoma

with liquid chromatographymass spectrometry has reported 2.1% with glycine (R132G), 1.7% with serine (R132S), 0.8% with
high levels of the metabolite 2-HG in mutant cells (4). Then, it leucine (R132L), and 0.3% with glutamine (R132Q) (9).
has been demonstrated that mutant IDH1 catalyzes the reduction Following the first observation of recurrent IDH1 mutation in
of -KG to the R-enantiomer of the metabolite 2-HG (R-2-HG) glioblastoma, several groups have begun to clarify the frequency
(8). Thus, rather than catalyze the NADP+-dependent produc- and distribution of IDH1 mutation across all brain tumors,
tion of -KG, mutant IDH1 catalyzes the NADPH-dependent including gliomas and other subtypes. Data summarized from
reduction of -KG to produce only the R-enantiomer of 2-HG, many studies show that only approximately 5.6% of primary
indicating a gain of neomorphic function. Specifically, authors glioblastoma are IDH1 mutant, while more than 76% of second-
have demonstrated that the mutation reduces the affinity of the ary glioblastomas carry the IDH1 mutation. The reported rates
IDH1 active site for isocitrate while concomitantly increasing it of IDH1 mutation in lower-grade gliomas are comparable with
for NADPH and -KG (8). Reduced affinity for isocitrate occurs those of secondary glioblastoma (Table2) (10).
as a result of alterations to a binding site residue that forms Other brain tumors harboring IDH1 mutations with moderate
hydrogen bonds between the alpha and beta groups of isocitrate frequency include gangliogliomas, giant cell glioblastomas, and
(8). Consequently, the conversion of -KG is favorized, but rather primitive neuroectodermal tumors, although only small numbers
than isocitrate, the mutant enzyme converts -KG into R-2-HG, of these tumors have been studied (11).
an oncometabolite that promotes tumorigenesis through inhibi- Isocitrate dehydrogenase mutations are also present in some
tion of -KG-dependent (Figure1). tumors originating in cells outside of the central nervous system.
Indeed, about 7.7% of acute myeloid leukemia (AML) possessed
INCIDENCE OF IDH1 MUTATION IN the IDH1 mutation (12), but prevalence rates vary between 15
GLIOBLASTOMA AND OTHER CANCERS and 33% if IDH2 are also considered (13). Approximately 50% of
d-2-hydroxyglutaric aciduria (d-2-HGA), a rare inherited neuro-
Extensive genomic profiling has revealed that about 90% of IDH1 metabolic disorder, has also been found to display IDH1 mutations
mutations involve exon 4 at codon 132, replacing arginine with (14), as well as 10% of intrahepatic cholangiocarcinomas (15), 5%
histidine (R132H). Of the remaining 10% of IDH1 mutations, of myelodysplastic syndrome (MDS), 8.8% of myeloproliferative
4.7% are due to arginine being replaced with cysteine (R132C), neoplasms (MPN), and fewer than 10% of secondary AML (14).

FIGURE 1 | Potential mechanisms implicated in tumor formation induced by IDH1 mutation. A number of potential mechanisms have been proposed to
explain how R-2-HG produced by mutant IDH1 enzyme promotes tumor formation. Epigenetic modification, via inhibition of -KG-dependent dioxygenases leading
to DNA and histone hypermethylation, has been at the forefront of research efforts. Specifically, the inhibition of tet methylcytosine dioxygenase 2 (TET2) leads to
increased DNA hypermethylation and the inhibition of histone demethylases leads to increased histone tail methylation. Additional mechanisms include inhibition of
several groups of prolyl hydroxylases, leading to hypoxia-induced factor (HIF) activation and alterations in collagen formation.

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Liu et al. Genetics and Epigenetics of Glioblastoma

TABLE 2 | Frequency of IDH1 mutation in various tumors. mutations being queried. Other PCR-based techniques include
Tumor types IDH1 mutation frequency (%) coamplification at lower temperature (COLD) PCR with high-
resolution melting (HRM) and real-time PCR, and post-PCR
Astrocytoma pilocytic (WHO Grade I) 0.01 fluorescent melting curve analysis (FMCA). Through COLD
Diffuse glioma (WHO Grade II) 76
PCR combined with HRM, Boisselier and colleagues were able to
Astrocytoma anaplastic (WHO Grade III) 62.2
detect mutant allele concentrations of 0.25% in a span of only 3h
Primary glioblastoma (WHO Grade IV) 5.6
Secondary glioblastoma (WHO Grade IV) 76.4
(21). However, because the technique requires the new mutation
to have a melting temperature (Tm) that is lower than IDH1 wild
Oligodendroglioma
type, it theoretically may not be able to detect R132G mutation.
WHO Grade II 78.8
Regarding real-time PCR with post-PCR FMCA, this technique
WHO Grade III 67.5
was shown to be more sensitive than Sanger sequencing with
Oligoastrocytoma detection rate of as little as 10% mutant DNA and a processing
WHO Grade II 79.8
time of 80min.
WHO Grade III 69.7
Other technologies currently in use to detect mutations in the
CNS, central nervous system; WHO, World Health Organisation; AML, acute myeloid IDH1 gene include SNaPshot (22) and Oncomap (23), both of
leukemia; d-2-HGA, d-2-hydroxyglutaric aciduria; MDS, myelodysplastic syndrome;
which can be used with paraffin-embedded tissue, using base pair
MPN, myeloproliferative neoplasms.
extension that results in an allele-specific probe that is read out by
either fluorescence detection (SNaPshot) or mass spectrometry
In their investigation, Amary and colleagues found that nearly (Oncomap). In addition to these approaches, Boisselier and
60% of central and periosteal cartilaginous tumors displayed colleagues demonstrated evidence of principle in detection of
DH1mutations (16). The same group has also identified IDH1 IDH1 mutations from the plasma of patients with mutant glioma
mutation to occur in patients with Ollier disease and Maffucci (24). Although the potential application for monitoring disease
syndrome (17). The majority of patients exhibited the R132C non-invasively is compelling, the sensitivity attained using this
IDH1 mutation, in contrast to most secondary glioblastoma, assay was 60%. Further work on the nature of circulating tumor
which harbor the R132H mutation. material will be necessary to determine whether it will be possible
to monitor the IDH1 mutation status in the peripheral blood of
TRANSLATIONAL APPLICATIONS OF all patients with mutant gliomas.
MUTANT IDH1 Recently, a new technique called amplification-refractory
mutation system (ARMS) has been developed (25) to identify
In a relatively short time after the discovery of IDH1 mutation in R140Q mutations in IDH2 and such novel methods can be of
glioblastoma, a tremendous amount of work has been performed great help in detecting IDH1 mutations. Recently, a novel strategy
on the clinical relevance of this mutation regarding particularly of PCR clamping was employed for qualitative detection of seven
its applications in the diagnosis, prognosis, and treatment of different mutations in IDH1 and five mutations in IDH2 in a
patients suffering from glioblastoma. single PCR assay (26).
A monoclonal antibody specific for the IDH-R132H mutation
Diagnostic Applications (mIDH1R132H), developed by Cappers group, recognizes the
The determination of IDH1 mutation status is highly relevant mutant protein with a high degree of sensitivity and specific-
for the diagnosis of primary brain tumors, and strongly supports ity (27). Using this antibody, 90% of IDH1 mutation can be
the differential diagnosis between an anaplastic glioma and a detected on paraffin sections, and it is recommended to test
glioblastoma. the remaining 10% by sequencing. Nevertheless, proponents of
Traditionally, IDH1 mutation status was detected through immunohistochemistry-based antibody staining argue that the
classic Sanger sequencing and polymerase chain reaction (PCR). use of mIDH1R132H antibody to identify IDH1 mutation may
Although this has the clear advantage of being able to detect non- even be superior to direct sequencing because there are reported
R132H mutation, it is time consuming and requires there to be cases in which this antibody detects mutations missed by direct
at least 20% mutant allele frequency within the tissue specimen sequencing (28).
for reliable detection (18). Pyrosequencing is an alternative that Beyond determination of IDH1 mutation status, the histo-
allows for rapid high-throughput analysis of IDH1 mutation, and pathological utility of the mIDH1R132H antibody has extended to
has demonstrated an advantage over classic Sanger sequencing additional clinical scenarios such as the discrimination between
in that it can detect mutated allele frequencies down to 5% (19). diffuse astrocytoma and reactive astrocytosis when combined
Derived cleaved amplified polymorphic sequence analysis is with a panel of key molecular feature (29). Indeed, seeing that
another alternative to DNA sequencing that uses mismatched IDH1 mutations are specific to gliomas, this antibody can be
primers for specific mutations, which, following PCR amplifica- used to help differentiate between diffuse gliomas and areas of
tion will create differing restriction endonuclease sites depend- reactive gliosis. Accordingly, the detection of positive cells by
ent on the presence of the mutation (20). The advantage of this IDH1-R132H immounohistochemistry allows a clear and safe
technique is that it uses supplies commonly found in most labo- separation between low-grade glioma and reactive gliosis, and
ratories, obviating expensive sequencing equipment. However, clearly supports the diagnosis of a diffusely infiltrating glioma
unlike sequencing, the method is limited in that it can only detect as well as the differential diagnosis between an anaplastic glioma

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Liu et al. Genetics and Epigenetics of Glioblastoma

and a glioblastoma. This approach allows narrowing down the improved prognosis with respect to both overall survival and
possible diagnosis to the group of diffusely infiltrating of WHO progression-free survival for the rare glioblastoma patients
grade II and III and secondary glioblastoma and to a certain who express this mutation. Indeed, since the publication of the
extent to primary glioblastoma. IDH1-R132H immounohisto- first report on improved survival in glioblastoma patients with
chemistry has also been shown to be effective in separating out mutation by Parsons and colleagues, indicating 45.6 months
oligodendrogliomas from several other similar entities, such as overall survival in IDH1-mutant versus 13.2 months overall
central neurocytomas, tanycytic ependymomas, and pilocytic survival in IDH1 wild type (6), numerous groups have been able
astrocytomas (27). to replicate similar findings (3641). Besides improved overall
Another area of investigation has focused on the detection of survival, Sanson et al. (37) were able to demonstrate improved
the 2-HG metabolite rather than on the specific sequence of the progressive-free survival (PFS) as well in their set of patients
IDH1 gene or protein product. In theory, sensitive and specific with glioblastoma, with 55 months PFS in patients with IDH1
detection of 2-HG is sequence independent in that 2-HG should mutation versus 8.8months PFS in those without this mutation.
be present regardless of the type of mutation in IDH1 or IDH2. The analysis was extended to anaplastic (WHO grade III) tumors
High levels of 2-HG have been detected in ex vivo tissue samples because many groups were readily able to show an improved
by using two approaches. In the first approach, combination overall survival in grade III tumors that harbored the IDH1 muta-
gas or liquid chromatography/mass spectrometry was used to tion compared with those that did not in both univariate (36)
identify 2-HG in frozen or paraffin-embedded glioma tissue. and multivariate analyses (37, 40). With anaplastic astrocytomas,
However, these extraction-based approaches do not preserve the patients harboring IDH1 mutation had an overall survival of
integrity of the sampled tissue. In the second approach, proton 65 months compared to 20 months for patients without IDH1
high-resolution magic angle spinning (HRMAS) MRS is used to mutation (38). The survival benefit also extended to grade II glio-
determine the metabolic profiles in ex vivo tissue. This technique mas, showing a median overall survival of 12.6years in patients
does not require alteration of tissue samples, and identified 2-HG with IDH1 mutation versus 5.5years in patients devoid of this
in ex vivo specimens with high sensitivity and specificity. Unlike mutation. In a prospective analysis, Wick and colleagues found
the case with AML, wherein 2-HG can be detected in the blood that grade III astrocytomas that possessed the IDH1 mutation
of patients with IDH-mutant AML (30), its presence in peripheral were associated with greater PFS regardless of the treatment (42).
blood is similar between patients with IDH1-mutant and wild- In studies pooling low-grade astrocytomas and oligodendro-
type tumors (31). gliomas, the IDH1 mutation status was prognostic for overall and
Detection of 2-HG by MRS represents a completed non- PFS. In primary glioblastoma, IDH1 mutational status has been
invasive method with which to determine the presence of IDH reported to be the only factor that showed significant association
mutations in gliomas, irrespective of the sequence of the muta- with patient survival times. However, the evidence for low-grade
tion or the mutation maps to IDH1 or IDH2. Importantly, this gliomas and the prognostic value of IDH1 mutation is slightly
approach represents the only example in human cancer in which more controversial. Two independent groups found that IDH1
a genomic feature can be identified specifically by using imaging- mutations in low-grade gliomas were associated with significant
based metabolic profiling. Prior work completed by MRS-based improved overall survival (43, 44), whereas others could not find
approach has demonstrated that IDH-mutant tumors display any significant association (36, 45). Nevertheless, the consistent
characteristic imaging findings. The IDH1-mutant tumors dis- finding of a more favorable outcome of diffuse gliomas patients
play reduced contrast enhancement, less surrounding edema, with IDH1 mutation implies that IDH1 testing might be useful
cystic components, and often found in the frontal lobe compared for prognostic considerations in the clinical setting.
with IDH-wild-type tumors (32). Because it is recognized that the
spectrum of 2-HG has some overlap with other commonly found Predictive Implications
metabolites, such as glutamate and glutamine, the methodology Isocitrate dehydrogenase 1 mutation was correlated with a higher
for obtaining and analyzing in vivo MRS is critical. One study rate of response to up-front temozolomide in low-grade glioma
to demonstrate 2-HG detection in glioma by MRS used the patients. Furthermore, evidence for differential responsiveness to
acquisition of a point-resolved spectroscopy (PRESS) sequence genotoxic therapy of IDH1 mutant versus IDH1 wild-type low-
(33). There was good concordance between sequence-based grade gliomas has been provided. Indeed, the presence of IDH1
mutation status and 2-HG detection, although several false mutation was associated with favorable progression-free and
positives and false negatives were reported. Additional studies in overall survival in WHO grade II gliomas who received radio- or
which spectral-editing analysis (34) or 2D-correlation MRS with chemotherapy. Current studies aim to validate and clarify the
spectral editing (35) were used demonstrated that IDH mutation predictive value of IDH1 mutation in the different glioma types.
status can be identified non-invasively by MRS techniques with a It is still unclear if IDH1 mutational status is a prognostic
high levels of sensitivity and specificity. indicator or predictive measure of response to treatment.
Houillier and colleagues stratified a cohort of low-grade
Prognostic Implications gliomas into three groups based on prognostic factors
Extensive research has been performed to determine the prog- according to the presence of 1p19q deletion, IDH1 mutation,
nostic value of IDH1 mutations, and a better prognosis has been or both together (46). They found that each of these factors
generally reported in glioblastoma patients carrying an IDH1 was an independent predictor of improved clinical outcome
mutation. Studies have shown that IDH1 mutation convey an in response to treatment with the chemotherapeutic agent

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Liu et al. Genetics and Epigenetics of Glioblastoma

temozolomide and that the group of patients with both muta- silencing of the -KG-dependent DNA modifying enzyme, Tet
tions had the best treatment response. Objective response was methylcytosine dioxygenase 2 (TET2) (Figure1).
in 80% with both mutations, 61% of IDH1-mutants without Since the first report demonstrating that a subset of glioblas-
1p19q deletion, and 17% without either mutation. In a similar toma exhibits a global decrease in 5-methylcytosine, subsequent
fashion, Hartman and colleagues found that, in their cohort follow-up studies have revealed not only that genome-wide or
of patients receiving adjuvant therapy, IDH1 mutation status global hypomethylation occurs at a frequency of 80% in primary
was the single most important predictor of PFS and overall glioblastoma, but also that the level of hypomethylation varies
survival. This was not seen in their cohort of patients that did between glioblastomas, ranging from near normal levels to
not receive adjuvant therapy (47). These findings support the approximately 50% of normal in about 20% of cases (51). The
notion that IDH1 mutation may be an important predictor to most severe globally hypomethylated primary glioblastomas are
treatment response. also the most proliferative and display dramatic hypomethyla-
tion (2250% of normal brain) of the tandem repeat satellite 2
Treatment Implications (Sat2) located at the juxtacentromeric region of chromosomes 1,
The identification of IDH1 mutation and the rapid characteriza- 9, and 16, and moderate hypomethylation (7180%) of the D4Z4
tion of its protein products present a therapeutic opportunity located at the subtelomeric regions of chromosomes 4q35 and
to treat the IDH1-mutant tumors. From a medical standpoint, 10q26 (51). Moreover, glioblastoma with hypomethylated Sat2
it will be important to determine whether these mutant tumors also harbored copy number alterations of adjacent euchromatic
would be sensitive to molecules that inhibit the mutant enzyme. sequences, specifically near the pericentromeric region of chro-
Although there are no published studies to date addressing this mosome 1. These data suggest that one consequence of hypo-
possibility, one study performed by Jin and colleagues suggested methylated repetitive sequences in glioblastoma is predisposition
that mutant tumor cells may depend on the continued expression to chromosomal breakage and copy number alteration. Although
of the mutant enzyme and/or its resulting 2-HG metabolite. In the full consequences of genomic hypomethylation are unknown,
their investigation, the authors showed that a cell line expressing murine models of defective imprinting provide evidence for a
endogenous mutant IDH1 required its expression for survival causal role of DNA methylation alteration in tumorigenesis.
and anchorage-independent growth (48), suggesting that phar- DNA methylation has been shown to play critical roles in the
macological inhibition of mutant IDH1 may recapitulate this control of gene activity and the architecture of the nucleus of the
result. With respect to mutant IDH1-mediated biology, this result cell. In humans, DNA methylation occurs in cytosines that pre-
would also suggest that 2-HG-induced changes, such as global cede the guanines to create 5-methylcytosine and these are com-
DNA hypermethylation, are either potentially reversible or, if monly called dinucleotide CpGs (52). These dinucleotides are not
not, are insufficient for tumor maintenance. It may be anticipated randomly distributed in the genome, but instead are present as
that inhibition of this pathway would increase patient survival. CpG-rich regions referred to as CpG islands. Hypermethylation of
Although there has been some discussion of whether it is prudent CpG islands in the promoter regions of tumor-suppressor genes is
to inhibit mutant IDH1 because patients with mutant tumors have a major event in the origin of many cancers. In glioblastoma, CpG
a better survival than patients with wild-type tumors, it is not promoter hypermethylation occurs at genes with diverse func-
expected that inhibiting the mutant enzyme would make mutant tions related to tumorigenesis and tumor progression, including
tumors behave like their more aggressive wild-type counterparts. cell cycle regulation (CDK2A-p16INK4a and CDK2B-p15INK4b)
The differences in survival most likely stem from the fact that tumor suppression (RB1, VHL, EMP3, RASSF1A, and BLU), DNA
IDH1-mutant and wild-type tumors arise from distinct lineages repair [methylguanine DNA methyltransferase (MGMT) and
and ontogenies and, thus, represent entirely different neoplastic MLH1], inhibition of apoptosis (DAPK1, TIMP3, CDH1), and
disease processes. genes associated with angiogenesis, regulation of tumor invasion,
and drug resistance (53). Many new tumor-suppressor candidates
have been identified, including the cell motility regulator testis-
EPIGENETIC CHANGES IN derived transcript (TES) as well as many polycomb repressor
GLIOBLASTOMA complex 2 (PRC2) target genes (54), and epithelial membrane
protein 3 (EMP3), a myelin-related gene involved in cell prolifera-
Epigenetic is the mitotically heritable changes in gene expression tion and cellcell interaction that is silenced by hypermethylation
that are not due to changes in the DNA sequence, and has emerged in glioblastoma (55).
as hallmark of human cancers. Indeed, aberrant epigenetic Promoter hypermethylation has been demonstrated to regu-
mechanisms, such as DNA methylation, histone modifications, late the oncogenic and proliferation-promoting transforming
chromatin remodeling, or altered non-coding RNA expression, growth factor (TGF)-beta signaling pathway in aggressive, highly
are currently recognized as relevant events in tumor formation proliferative glioblastomas. High levels of TGF-beta signaling are
(49). Until now, most studies about the epigenetic changes in glio- normally associated with poor prognosis. TGF-beta signaling
blastoma have focused on DNA methylation, including hyper- promotes proliferation through the induction of platelet-derived
methylation, gene-specific hypomethylation, and genome-wide growth factor-beta (PDGF-B). However, epigenetic silencing of
hypomethylation (50). The leading mechanism attributed to the PDGF-B can override the increased proliferative effects of TGF-
observed hypermethylation phenotype in IDH1 mutant involves beta signaling. Specifically, PDGF-B promoter hypermethylation

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Liu et al. Genetics and Epigenetics of Glioblastoma

prevents PDGF-B transcriptional activation by TGF-beta-induced in glioblastoma compared to low-grade astrocytomas and normal
Smad proteins (56). The oncogenic effect of the TGF-beta pathway brain, and overall histone H3 was more acetylated in glioblastoma
is, therefore, blocked by epigenetic alteration of one of its targets. (60). Large-scale sequencing of protein-coding genes in glioblas-
Genes involved in invasion and metastasis can also be affected toma revealed mutations in many genes involved in epigenetic
by promoter hypermethylation in glioblastoma. Approximately regulation, including HDACs, HDAC2 and HDAC9, histone dem-
87% of glioblastoma exhibit CpG hypermethylation of the ethylases, JMD1A and JMD1B, histone methyltransferases, SET7,
protocadherin-gamma subfamily A11 (PCDH-gamma-A11) SETD7, MLL, MLL4, and methyl-CpG binding domain protein1
gene, which is thought to be important in invasion of cancer cells (MBD1) (6). These intriguing initial studies suggest that alteration
into normal brain parenchyma (57). in epigenetic mechanisms could be a major defect in glioblastoma.
The hypermethylation of promoter can also modulate sensitiv-
ity to drugs and radiotherapy in glioblastoma. The best known CONCLUSION
example is the O6-MGMT promoter methylation and response to
alkylating agents, but there is also the gene suppressor of cytokine Following the discovery of IDH1 mutation, our understanding of
signaling 1 (SOCS1), which in some glioblastomas enhanced the biochemistry, genetics, and epigenetics as well as the preva-
resistance to ionizing radiation and decreased activation of lence and pathogenic role of this mutation has grown at a rapid
MAPKs associated with the ERK pathway after transcriptional rate, and a tremendous amount of work has been performed on
silencing by hypermethylation (58). This suggests that epigenetic its translational relevance in a relative short time. It is, henceforth,
profiling might be one way to categorize glioblastomas and to clear that IDH1 status is a major determinant of survival, and
rationally apply patient-specific therapy. many ways have been developed to identify the IDH mutation
For certain sets of genes, promoter hypermethylation might not as well as the oncometabolite 2-HG from clinical samples, using
be causal or may not be required for gene silencing in glioblastoma. non-invasive procedures. The determination of IDH1 status in
There is some evidence for deregulation of gene controlling histone glioblastoma will likely be an early step in treatment algorithms
modifications in this tumor. The gene encoding BMI-1, a member for patients suffering from this tumor.
of the polycomb group complex that regulates histone H3K27
methylation, is subject to frequent copy number alterations in both AUTHOR CONTRIBUTIONS
low- and high-grade gliomas (59). Expression of some histone dea-
cetylase (HDAC) proteins is reported to be altered in glioblastoma. The authors collected and analyzed the information and wrote
Class II and IV HDACs displayed decreased mRNA expression the manuscript.

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cytomas. Neoplasia (2005) 7:1939. doi:10.1593/neo.04490 Conflict of Interest Statement: The authors declare that the research was con-
58. Zhou H, Miki R, Eeva M, Fike FM, Seligson D, Yang L, et al. Reciprocal ducted in the absence of any commercial or financial relationships that could be
regulation of SOCS 1 and SOCS3 enhances resistance to ionizing radi- construed as a potential conflict of interest.
ation in glioblastoma multiforme. Clin Cancer Res (2007) 13:234453.
doi:10.1158/1078-0432.CCR-06-2303 Copyright 2016 Liu, Hou, Chen, Zong and Zong. This is an open-access article
59. Hayry V, Tanner M, Blom T, Tynninen O, Roselli A, Ollikainen M, etal. Copy distributed under the terms of the Creative Commons Attribution License (CC BY).
number alterations of the polycomb gene BMI1 in gliomas. Acta Neuropathol The use, distribution or reproduction in other forums is permitted, provided the
(2008) 116:97102. doi:10.1007/s00401-008-0376-0 original author(s) or licensor are credited and that the original publication in this
60. Lucio-Eterovic AK, Cortez MA, Valera ET, Motta FJ, Queiroz RG, journal is cited, in accordance with accepted academic practice. No use, distribution
Machado HR, et al. Differential expression of 12 histone deacetylase or reproduction is permitted which does not comply with these terms.

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