Molecular Diagnostics of Gliomas
Marina N. Nikiforova, MD; Ronald L. Hamilton, MD
NContext.—Gliomas are the most common primary brain
tumors of adults and include a variety of histologic types
and morphologies. Histologic evaluation remains the gold
standard for glioma diagnosis; however, diagnostic diffi-
culty may arise from tumor heterogeneity, overlapping
morphologic features, and tumor sampling. Recently, our
knowledge about the genetics of these tumors has
expanded, and new molecular markers have been devel-
oped. Some of these markers have shown diagnostic value,
whereas others are useful prognosticators for patient
survival and therapeutic response.
Objective.—To review the most clinically useful molec-
ular markers and their detection techniques in gliomas.
Data Sources.—Review of the pertinent literature and
personal experience with the molecular testing in gliomas.
Conclusions.—This article provides an overview of the
most common molecular markers in neurooncology,
including 1p/19q codeletion in oligodendroglial tumors,
mutations in the isocitrate dehydrogenase 1 and 2 genes in
diffuse gliomas, hypermethylation of the O6-methylgua-
nine-DNA methyltransferase gene promoter in glioblasto-
mas and anaplastic gliomas, alterations in the epidermal
growth factor receptor and phosphatase and tensin
homolog genes in high-grade gliomas, as well as BRAF
alterations in pilocytic astrocytomas. Molecular testing of
gliomas is increasingly used in routine clinical practice and
requires that neuropathologists be familiar with these
genetic markers and the molecular diagnostic techniques
for their detection.
(Arch Pathol Lab Med. 2011;135:558–568)
Currently, the histologic identification is based on the
morphologic resemblance of the tumor cells to nonneo-
plastic glial cells (ie, astrocytes, oligodendrocytes, and
ependymal cells), and, therefore, most malignant gliomas
of adults are classified into astrocytic, oligodendroglial,
mixed oligoastrocytic, and ependymal tumors, according
to the 2007 World Health Organization (WHO) classifica-
tion.1Molecular testing of ependymomas is of limited
value and will not be discussed in this review. In adults,
the most important types of gliomas are those that
diffusely infiltrate the surrounding brain tissue, making
these ‘‘diffuse gliomas’’ resistant to surgical resection. An
exception is the pilocytic astrocytoma (WHO grade I), the
most common pediatric glioma, which is relatively well
demarcated from the surrounding tissues and can be
resected. Diffuse gliomas are categorized into low-grade
gliomas (WHO grade II), which usually demonstrate
relatively slow growth, and high-grade gliomas (WHO
grades III and IV), which grow more rapidly. The most
malignant (and most common) of the high-grade gliomas
is glioblastoma multiforme (GBM). Primary GBMs arise
de novo in older patients and have a short duration of
clinical symptoms (,3 months). On the other hand,
secondary GBMs develop from preceding grade II or III
liomas in adults are the most common primary brain
tumors and include a variety of histologic types.
gliomas, have a longer duration of symptoms, and
frequently develop in patients younger than 40 years.
During the past decade, understanding of gliomagenesis
has expanded significantly. Current evidence suggests
that the initiation and progression of gliomas may involve
the accumulation of multiple genetic alterations. For
example, isocitrate dehydrogenase (IDH) 1 and IDH2
mutations are identified in most low-grade gliomas,
suggesting that IDH mutations are an early event in
gliomagenesis. Other genetic abnormalities may accumu-
late during tumor progression and include 1p/19q
codeletion in oligodendroglial tumors and TP53 mutation
or 17p13 loss in astrocytic tumors. Oligoastrocytomas may
exhibit any of these genetic abnormalities (Figure 1).2
Anaplastic gliomas are characterized by loss of 9p and
homozygous deletion of the CDKN2A/B gene (p16/p15),
and the progression of an anaplastic astrocytoma to a
secondary GBM is frequently associated with 10q loss.
Primary and secondary GBMs have a different subset of
genetic abnormalities. In particular, primary glioblasto-
mas demonstrate frequent epidermal growth factor
receptor (EGFR) amplification, 10q loss, and mutations
in the phosphatase and tensin homolog (PTEN) gene, but
rarely have IDH mutations. On the other hand, secondary
glioblastomas lack EGFR amplification but show muta-
tions in the TP53 and IDH genes (Figure 1). Most of
pilocytic astrocytomas are characterized by BRAF fusion
(BRAF/KIAA1549) but rarely have other genetic abnor-
malities (Figure 1).
Although histologic evaluation remains the gold stan-
dard for glioma diagnosis, diagnostic difficulty may arise
from tumor heterogeneity, overlapping morphologic
features, and tumor sampling. Malignant gliomas are
incurable, and most patients die despite aggressive
Accepted for publication January 11, 2011.
From the Department of Pathology, University of Pittsburgh,
The authors have no relevant financial interest in the products or
companies described in this article.
Reprints: Marina N. Nikiforova, MD, Department of Pathology,
University of Pittsburgh, 200 Lothrop Ave, Pittsburgh, PA 15213 (e-mail:
558Arch Pathol Lab Med—Vol 135, May 2011Molecular Diagnostics of Gliomas—Nikiforova & Hamilton
therapy. Advances in neurosurgery, radiation, and che-
motherapy during the past decade have provided only
small improvements in clinical outcome. As with many
other tumors, the development of new molecular markers
is expected to lead to improved diagnosis and prognosis
and to aid in the clinical management of gliomas.
Currently, the most common molecular markers in
neurooncology are 1p/19q codeletion in oligodendroglial
tumors, mutations in the IDH1/2 genes in diffuse gliomas,
hypermethylation of the O6-methylguanine-DNA methyl-
transferase (MGMT) gene promoter in glioblastomas and
anaplastic gliomas, alterations in the EGFR and PTEN
genes, and 10q deletions in GBMs, as well as BRAF
alterations in pilocytic astrocytomas. Some of these
markers can be used diagnostically to help the neuropa-
thologist in glioma classification and grading, especially
for tumors with ambiguous histologic features. Alterna-
tively, some of them can be used to estimate prognosis for
patients and to predict response to certain therapies.
1p AND 19q CODELETION
Loss of the short arm of chromosome 1 (1p), along with
the long arm of chromosome 19 (19q), is an established
genetic marker of oligodendroglial tumors.3The frequen-
cy of 1p and 19q codeletion has been reported to be 80% to
90% in oligodendrogliomas (WHO grade II), 60% in
anaplastic oligodendrogliomas (WHO grade III), and
30% to 50% in oligoastrocytomas (Table 1).3,4There is a
strong association between 1p/19q codeletion and the
classic histologic features of an oligodendroglioma (eg,
round, uniform nuclei with perinuclear halos and a
‘‘chicken-wire’’ vascular pattern). However, not all oligo-
dendroglial tumors reveal such correlation, and morphol-
ogy alone cannot predict the 1p/19q status.5Most 1p and
19q deletions appear to involve the loss of the entire 1p
and 19q chromosomal arms. The mechanism of this
codeletion was recently explained by the unbalanced
centromeric translocation t(1;19)(q10;p10) found in these
tumors, which mediates the loss of a 1p/19q derivative
chromosome and the preservation of a 1q/19p derivative
chromosome.6,7Almost all oligodendrogliomas with a 1p/
19q codeletion are positive for IDH1 or IDH2 mutations8
but rarely have the TP53 mutation, a 10q deletion, or
amplification of the EGFR gene.9,10
The 1pand 19q codeletion isfound to bea useful marker
for prognostication about patient survival and chemosen-
ampl., amplification; BRAF, v-raf murine sarcoma viral oncogene homolog B1 gene; CDKN2A/B, cyclin-dependent kinase inhibitors 2A and 2B
genes; EGFR, epidermal growth factor receptor gene; GBM, glioblastoma multiforme; IDH, isocitrate dehydrogenase gene; mut., mutation; PTEN,
phosphatase and tensin homolog gene; TP53, tumor protein p53 gene; WHO, World Health Organization.
Molecular pathways and common genetic alterations in astrocytic, oligodendroglial, and oligoastrocytic neoplasms. Abbreviations:
Average Prevalence of Genetic Abnormalities in Gliomas and Their Clinical Utility
Clinical utility of molecular testDiagnosticDiagnostic
Abbreviations: AA, anaplastic astrocytoma; AO, anaplastic oligodendroglioma; BRAF, v-raf murine sarcoma viral oncogene homolog B1 gene; DA,
diffuse astrocytoma; EGFR, epidermal growth factor receptor gene; GBM, glioblastoma multiforme; IDH, isocitrate dehydrogenase gene; LOH, loss
of heterozygosity; MGMT, O6-methylguanine-DNA methyltransferase gene; O, oligodendroglioma; PA, pilocytic astrocytoma; PTEN, phosphatase
and tensin homolog gene; WHO, World Health Organization; ?, clinical utility is not defined yet.
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75. Korshunov A, Meyer J, Capper D, et al. Combined molecular analysis of
BRAF and IDH1 distinguishes pilocytic astrocytoma from diffuse astrocytoma.
Acta Neuropathol. 2009;118(3):401–405.
76. Burger PC, Minn AY, Smith JS, et al. Losses of chromosomal arms 1p and
19q in the diagnosis of oligodendroglioma: a study of paraffin-embedded
sections. Mod Pathol. 2001;14(9):842–853.
77. Gan HK, Kaye AH, Luwor RB. The EGFRvIII variant in glioblastoma
multiforme. J Clin Neurosci. 2009;16(6):748–754.
78. Hatanpaa KJ, Burma S, Zhao D, Habib AA. Epidermal growth factor
receptor in glioma: signal transduction, neuropathology, imaging, and radiore-
sistance. Neoplasia. 2010;12(9):675–684.
79. Smith JS, Tachibana I, Passe SM, et al. PTEN mutation, EGFR
amplification, and outcome in patients with anaplastic astrocytoma and
glioblastoma multiforme. J Natl Cancer Inst. 2001;93(16):1246–1256.
80. Halatsch ME, Schmidt U, Behnke-Mursch J, Unterberg A, Wirtz CR.
Epidermal growth factor receptor inhibition for the treatment of glioblastoma
multiforme and other malignant brain tumours. Cancer Treat Rev. 2006;32(2):
81. Mellinghoff IK, Wang MY, Vivanco I, et al. Molecular determinants of the
response of glioblastomas to EGFR kinase inhibitors. N Engl J Med. 2005;353(19):
82. Haas-Kogan DA, Prados MD, Tihan T, et al. Epidermal growth factor
receptor, protein kinase B/Akt, and glioma response to erlotinib. J Natl Cancer
83. Heimberger AB, Sampson JH. The PEPvIII-KLH (CDX-110) vaccine in
glioblastoma multiforme patients. Expert Opin Biol Ther. 2009;9(8):1087–1098.
84. Yoshimoto K, Dang J, Zhu S, et al. Development of a real-time RT-PCR
assay for detecting EGFRvIII in glioblastoma samples. Clin Cancer Res. 2008;
85. Ohgaki H, Kleihues P. Genetic alterations and signaling pathways in the
evolution of gliomas. Cancer Sci. 2009;100(12):2235–2241.
86. Koul D. PTEN signaling pathways in glioblastoma. Cancer Biol Ther. 2008;
87. Hill C, Hunter SB, Brat DJ. Genetic markers in glioblastoma: prognostic
significance and future therapeutic implications. Adv Anat Pathol. 2003;10(4):
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