Surgical Neurolog
Genetics
Recent advances in the molecular genetics of malignant gliomas
disclose targets for antitumor agent perillyl alcohol
Clovis Orlando da Fonseca, MDa,T, Jose Alberto Landeiro, PhDa, Steven S. Clark, PhDb,
Thereza Quirico-Santos, PhDb,Maria da Gloria da Costa Carvalho, PhDc, Cerli Rocha Gattass, PhDd
aServico de Neurocirurgia, Hospital Universitario Antonio Pedro, UFF, Niteroi, RJ, BrazilbDepartamento de Biologia Celular and Molecular, Instituto de Biologia, UFF, Niteroi, RJ, Brazil
cLaboratorio de Controle da Expressao Genica and dLaboratorio de Imunologia Celular, Instituto de Biofısica Carlos Chagas Filho,
UFRJ, Rio de Janeiro, RJ, Brazil
Received 17 January 2005; accepted 9 June 2005
Abstract Tumors of glial origin such as glioblastoma multiforme (GBM) comprise the majority of human brain
www.surgicalneurology-online.com
0090-3019/$ – see fro
doi:10.1016/j.surneu.2
T Corresponding
CEP 22631-050, Rio
E-mail address: c
tumors. Despite advances in surgery, radiation, and chemotherapy, the prognosis for patients with
malignant glioma has not improved, emphasizing the need for a search for new chemotherapeutic
drugs. Deregulated p21-Ras function, as a result of mutation, overexpression, or growth factor-
induced overactivation, contributes to the growth of GBM. The monoterpene perillyl alcohol (POH)
has preventive and therapeutic effects in a wide variety of preclinical tumor models and is currently
under phase I and phase II clinical trials. As inhibition of posttranslational isoprenylation of Ras, a
family of proteins that are involved in signal transduction is among the drug-related activities
observed in this compound; POH may be a potential chemotherapeutic agent for GBM. Intranasal
delivery is a practical and noninvasive approach that allows therapeutic agents that do not cross the
blood-brain barrier to enter the central nervous system, reducing unwanted systemic side effects. This
article describes the effect of intranasal delivery of POH in a patient with relapsed GBM.
D 2006 Elsevier Inc. All rights reserved.
Keywords: Intranasal delivery; Perillyl alcohol; Glioblastoma multiforme therapy; Brain tumor
1. Introduction
For some types of tumors, the conventional treatment
approach, radiation and chemotherapy, has translated into
increased survival. Unfortunately, the effect of this approach
in patients with high-grade gliomas has been modest, and the
median survival of patients remains less than 1 year from the
time of diagnosis. These tumors rarelymetastasize outside the
central nervous system (CNS), but they diffusely invade the
host brain, and the lack of clear tumor margins makes it
impossible to remove the entire tumor [24]. Among astrocytic
gliomas, glioblastoma multiforme (GBM) is the most com-
mon and most aggressive type. In recent years, understanding
of glioma tumorigenesis, proliferation, and invasion had a
great advance as researchers gained a better understanding of
nt matter D 2006 Elsevier Inc. All rights reserved.
005.06.030
author. Avenida Canal de Marapendi no. 1600/1303,
de Janeiro, Brazil.
[email protected] (C.O. da Fonseca).
the molecular biology of these tumors [6]. Abnormal growth
factor signaling is implicated in the pathogenesis of gliomas.
Glioblastoma multiforme commonly overexpresses the
oncogenes EGFR and PDGFR, and contains mutations and
deletions of tumor suppressor genes TP53 and PTEN. Some
of these alterations lead to activation of Ras/MAPK pathways
[11] and PI3K/Akt [23], providing targets for therapy. This
study summarizes the current knowledge on the signaling
pathways contributing to the pathogenesis of malignant
gliomas, describes strategies to inhibit constitutively activat-
ed Ras/MAPK and PI3K/Akt signaling pathways, and
suggests perillyl alcohol (POH) as a potential agent for the
treatment of GBM.
2. Glioblastoma multiforme
Glioblastoma multiforme is by far the most common and
most malignant of the glial tumors. Composed of poorly
y 65 (2006) S1:2–S1:9
C.O. da Fonseca et al. / Surgical Neurology 65 (2006) S1:2–S1:9 S1:3
differentiated neoplastic astrocytes, glioblastomas primarily
affect adults, and they are located preferentially in the
cerebral hemispheres. Rarely, GBM can affect the brain stem
in children and the spinal cord. These tumors may develop
from lower-grade astrocytomas (World Health Organization
grade II) or anaplastic astrocytomas (World Health Organi-
zation grade III), but, more frequently, they manifest de novo,
without any evidence of a less malignant precursor lesion. In
addition to cellular atypia, increased mitotic index, and
infiltrative growth into adjacent normal brain, GBM shows
intratumoral necrosis and vascular endothelial proliferation.
The study of gliomagenesis is an attempt to assign specific
genetic abnormalities to distinct phases or grades of
astrocytomas. The aim is to understand how particular
signaling pathways might cause the progression of astrocy-
tomas from lower to higher grades of malignancy or how they
might trigger the generation of highly malignant astrocyto-
mas de novo [41]. Primary GBM apparently occurs de novo
and is associated with a short duration of symptoms.
Secondary GBM arises in patients who have had a previous
lower-grade astrocytic tumor or who have had symptoms for
several months. This difference in clinical presentation
suggests that primary and secondary GBMs have a different
pathogenesis. This hypothesis is supported by the observa-
tion that primary and secondary glioblastomas have a
different spectrum of genetic alterations. In fact, secondary
GBM has been found to have many of the alterations
observed in low-grade and anaplastic astrocytomas, along
with other anomalies, such as TP53 mutation [34], whereas
de novo GBM exhibits more frequent occurrences of EGFR
gene amplification [6]. Amplification of the EGFR locus is
found in approximately 40% of primary GBM but is rarely
found in secondary GBM [43]. Another important genetic
alteration seen more frequently in primary than in secondary
GBM involves the PTEN gene. Mutation of PTEN, ampli-
fication of EGFR, and loss of the q arm of chromosome 10
were statistically significant in GBM [37]. Besides its tumor
suppressor function, the protein encoded by the PTEN gene
regulates several processes that may play a critical role in the
cellular aggressiveness observed in GBM, including angio-
genesis, migration, and invasiveness [19]. Most gene alter-
ations induce cell-cycle dysfunction on a complex molecular
level. Further insight into tumor genesis bymeans of genomic
assays may aid in predicting the clinical behavior of
glioblastoma and in providing individualized potential targets
for therapeutic agents.
3. The Ras pathway as a target for GBM therapy
Standard treatment of GBM consisting of surgical
resection, radiation, and/or chemotherapy is rarely curative
[35]. Gene therapy of glioblastomas is limited by the short
survival of viral vectors and by their difficulty in reaching
glioblastoma cells infiltrating the brain parenchyma [5]. To
improve the prognosis of GBM, the development of new
therapeutic strategies is necessary. Glioblastoma multiformes
are suitably targeted by molecular therapy because they
display a set of defined molecular lesions and signaling
pathway alterations that may be used as targets for therapy.
Primary GBMs, those arising as de novo lesions, commonly
overexpress EGFR and its ligand-independent mutant
EGFRvIII [42]. This results in signaling through Ras-MAPK
and PI3K/Akt pathways. Target molecules extensively
studied include EGFR, PDGFR, PTEN, telomerase, and
signal pathway modulators for PI3K/Akt and Ras/MAPK
pathways. Therapies targeting these specific molecules may
result in killing tumor cells effectively while keeping normal
cells intact.
Phosphatidylinositol-3-kinase (PI3K) is a lipid kinase and
generates phosphatidylinositol-3,4,5-trisphosphate. Phos-
phatidylinositol-3,4,5-trisphosphate is a second messenger
essential for the translocation of Akt to the plasmamembrane,
where it is phosphorylated and activated by phosphoinosi-
tide-dependent kinases 1 and 2. Activation of Akt plays a
pivotal role in fundamental cellular functions such as cell
proliferation and survival by phosphorylating a variety of
substrates [21]. In recent years, it has been reported that
alterations to the PI3K-Akt signaling pathway are frequent in
human cancer. Constitutive activation of the PI3K-Akt
pathway occurs because of the amplification of the PIK3C
gene encoding PI3K or the Akt gene, or because of mutations
in the components of the pathway, for example, PTEN, which
inhibit the activation of Akt. Several small molecules,
designed to specifically target PI3K-Akt, have been devel-
oped and shown to induce cell-cycle arrest or apoptosis in
human cancer cells in vitro and in vivo [30]. Moreover, the
combination of an inhibitor with various cytotoxic agents
enhances the antitumor efficacy. Therefore, specific inhibi-
tion of the activation of Akt may be a valid approach to
treating human malignancies and overcoming the resistance
of cancer cells to radiation or chemotherapy.
Ras is an integral signaling element and has been
characterized as the primary switch that transmits external
signals through numerous intracellular signaling pathways.
Ras belongs to a superfamily of small molecular weight
guanine nucleotide–binding proteins with the intrinsic ability
to hydrolyze guanosine 5V-triphosphate known as GTPases.
At least 20 members of this superfamily have been
identified. Three genes encode Ras proteins: N-ras, H-ras,
and K-ras. The 3 Ras proteins coded by these genes are
closely related to one another and are similar in their ability
to interact with regulators and effectors. Ras proteins are a
class of nucleotide-binding proteins that play pivotal roles in
the control of normal and transformed cell growth. Exper-
imental studies on the structure, function, and regulation of
Ras proteins indicate that they are key intermediates in signal
transduction pathways that mediate proliferative and other
types of signal largely from upstream of receptor kinases,
which control a wide variety of cellular processes, including
growth, differentiation, apoptosis, cytoskeletal organization,
and membrane trafficking [8,22]. For Ras to transduce
the extracellular signals provided by growth factors and
C.O. da Fonseca et al. / Surgical Neurology 65 (2006) S1:2–S1:9S1:4
cytokines, it must be associated with the inner surface of the
plasma membrane. Ras proteins are a family of membrane-
associated small GTPases that transmit signals from cell
surface receptors such as EGFR, EGFRvIII, and PDGFR,
promoting diverse cellular effects such as proliferation
survival and angiogenesis [26]. Membrane anchorage of
Ras, required for functional activity in signal transduction, is
facilitated by posttranslational modifications resulting in
covalent attachment of a farnesyl group to the cysteine in the
C-terminal CAAX motif. This attachment is mediated by
farnesyltransferase [22]. These lipophilic modifications
facilitate the association of isoprenylated proteins with an
intracellular membrane, which is a functional requirement.
Farnesylation of Ras enhances its ability to stimulate
downstream signaling enzymes, including mitogen-activated
protein kinase in mammalian cells. Unfarnesylated Ras
proteins do not associate the plasma membrane and are
incapable of cellular transformation. Farnesyltransferase
inhibitors represent a new class of agents that target signal
transduction pathways responsible for the proliferation and
survival of diverse malignant cell types. Although these
agents were developed to prevent a processing step
necessary for membrane attachment and maturation of Ras
proteins, recent studies suggest that farnesyltransferase
inhibitors block the farnesylation of additional cellular
polypeptides, thereby exerting antitumor effects independent
of the presence of activating ras gene mutations [22].
Although GBM does not display Ras mutations, it may have
enhanced expression of Ras. Also, GBM expresses high
levels of ligand-dependent and ligand-independent growth
factor (EGF and PDGF) receptors. Activation of these
receptors leads to tyrosine kinases activation and functional
upregulation of the Ras signaling pathway or expresses the
activated form of this protein [17]. Overexpression and
activation of receptor tyrosine kinases, such as PDGFR and
EGFR, lead to proliferation of human malignant astrocytoma
cells. Although oncogenic mutations affecting Ras are not
prevalent in human malignant astrocytomas, they might be
elevated in these tumors secondary to the mitogenic signals
originating from activated receptor tyrosine kinases. In
support of this hypothesis, high levels of Ras.GTP, similar
to those found in oncogenic Ras-transformed fibroblasts,
were present in 4 established human malignant astrocytoma
cell lines that express PDGFR and EGFR in 20 operative
malignant astrocytoma specimens [11]. Stimulation of
PDGFRs and EGFRs induced tyrosine phosphorylation of
the Shc adaptor protein and its association with Grb2,
suggesting a mechanism by which Ras may be activated in
human malignant astrocytoma cells. Furthermore, blocking
Ras activation by expression of the H-Ras–Asn17 dominant-
negative mutant, or by farnesyl transferase inhibitors,
decreased in vitro proliferation of the human astrocytoma
cell lines [13]. These results support the hypothesis that
proliferative signals from receptor tyrosine kinases
expressed by human malignant astrocytoma cells use the
Ras mitogenic pathway. Pharmacological inhibitors of the
Ras pathway may therefore be of therapeutic value in these
presently terminal tumors. Previous studies have demon-
strated that astrocytomas express elevated levels of activated
Ras.GTP despite the absence of activating Ras mutations.
The importance of increased Ras activity in GBM is
supported by studies demonstrating reduced growth of
preclinical GBM if treated with dominant-negative Ras
mutants or Ras pathway inhibitor drugs. Furthermore,
overexpresssion of H-Ras in a transgenic mouse model
resulted in early death from growth of multifocal malignant
astrocytomas [24]. Overall, the current data suggest that
activation of the Ras signaling pathway in malignant gliomas
is due to aberrant expression and overactivity of membrane
tyrosine kinase receptors, including EGFR, PDGFR, FGFR,
and IGF-IR.
4. Perillyl alcohol—inhibition of the Ras/MAPK
pathway
Over the past 3 decades, we have made great strides in the
treatment of most, but not all, brain tumors. Dramatic
advances have occurred in diagnostic imaging, neurosurgery,
neuroanesthesia, radiotherapy, and chemotherapy for CNS
tumors. Unfortunately, our progress has not yet met our
expectations. Because of the infiltrative nature of most
primary brain tumors, neurosurgery can never be expected to
be curative for the majority of gliomas. Because infiltrative
tumors interdigitate with normal brain cells and are not
highly sensitive to irradiation, one cannot expect radiother-
apy to be curative without serious damage to normal brain
cells. The hope for a cure, then, rests with chemotherapy. The
ability to treat most advanced malignancies with classic
cytotoxic DNA-damaging agents is limited, with little
curative potential and rare durable remissions. This has led
to emphasis on the development of new therapeutic agents
with novel mechanism of action. Cancer researchers now
have a unified concept to guide their search for specific
genetic abnormalities. The genes and proteins that participate
in the conversion of normal into malignant cells are also
involved in the key processes that convert extracellular
events that culminate in division and growth.
The search for new chemotherapeutic drugs has increased,
especially for those that have a natural origin. Diverse
mevalonate-derived products of secondary metabolic path-
ways present in plants have both chemotherapeutic and
chemopreventive properties. The mevalonate pathway pro-
duces isoprenoids that are vital for diverse cellular functions,
ranging from cholesterol synthesis to growth control. Several
mechanisms for feedback regulation of low-density-lipopro-
tein receptors and of 2 enzymes involved in mevalonate
biosynthesis ensure the production of sufficient mevalonate
for several end products. Manipulation of this regulatory
system could be useful in treating certain forms of cancer
[16]. Early studies of the mechanism of the anticarcinogenic
actions of 2 pure isoprenoids, d-limonene and POH, revealed
that these cyclic monoterpenes suppressed the incorporation
C.O. da Fonseca et al. / Surgical Neurology 65 (2006) S1:2–S1:9 S1:5
of radiolabeled isoprenes into small G proteins in
carcinoma cell lines [unpublished data]. Perilyll alcohol,
also known as p-metha,1,7-diene-6-ol and sometimes as
4-isopropenyl-cyclohexenecarbinol, is composed of 2 iso-
prene units produced by the mevalonate pathway. It has
been found to be active in inducing apoptosis in tumor cells
with no impact on normal cells and can, in fact, turn back
tumor cells to a differentiated state [4].
Although the mechanism by which POH exerts its
anticancer activity is not clear, a number of potentially
important drug-related activities have been observed in
preclinical studies, including cellular effects such as an early
G1 arrest and the induction of apoptosis, biochemical effects
such as the inhibition of posttranslational modification of
proteins involved in signal transduction and differential gene
regulation with overexpression of M6P/IGF-II and TGF-btype II receptor genes. Indeed, it has been postulated that the
anticarcinoma activity of POH involves a decrease in the
levels of isoprenylated Ras and Ras-related proteins, thereby
reducing the physiological functioning of these proteins [1].
Protein isoprenylation involves the posttranslational modifi-
cation of a protein by the covalent attachment of a lipophilic
farnesyl isoprenoid group to a Cys residue at or near the
carboxyl terminus. Isoprenoid substrates for prenylprotein
transferase enzymes include farnesylpyrophosphate and
geranylgeranylpyrophosphate, 2 intermediates in the meval-
onate pathway [16]. This action was widely attributed to the
inhibition of farnesyl protein transferase activity. Farnesyla-
tion of Ras also greatly enhances its ability to stimulate
downstream signal-transduction enzymes, including mito-
gen-activated protein kinase in mammalian cells [19].
Farnesylation is the most critical part of the process that
leads to the activation of Ras [12], and farnesyl transferase
inhibitors exert their antitumor effect, in part, by inhibiting
Ras-mediated signaling. Nevertheless, farnesyl transferase
inhibition may also block signaling from other pathways that
also require farnesylation, including the Rho B and PI3K/Akt
pathways [21]. Also, follow-up studies revealed that POH
suppresses the synthesis of small G proteins and HMG CoA
reductase [28]. In addition, POH has been shown to induce
apoptosis and cause a G0/G1 arrest in liver tumors [29], colon
cancer cell [32], mammary carcinomas cells [36], leukemic
cell lines [33], and a transitory G2 arrest and Fas-mediated
apoptosis in prostate cells [20] and GBM cells [31]. In
pancreatic ductal adenocarcinoma cells, the apoptotic effect
of POH appears to involve the increase of the proapoptotic
protein Bak in a wild-type p53-dependent way [38]. But in
human Philadelphia chromosome–positive leukemia cells,
POH reduces the apoptotic threshold by lowering the
expression of Bcl-xL, which allows a c-Myc-dependent
apoptosis to proceed [9].
However, in contrast to what has been reported for
carcinomas, POH-mediated anticancer activity is not associ-
ated with inhibition of Ras protein prenylation in other
malignant cells. For instance, in Philadelphia chromosome–
positive leukemia cells that express the Bcr/Abl tyrosine
kinase, POH causes a rapid and profound G1 arrest that is
followed by apoptosis, without affecting Ras prenylation or
activity [9]. Rather, the POH antileukemia effect was
associated with a rapid (within 2-4 hours) reversal of the
activating phosphorylation of the MAPK and of the Stat5
transcription factor [personal communication, Yang X,
Clark SS]. Reversal of Mek activation blocks signaling
through the mitogen-activated protein kinase pathway,
whereas reduced expression of active Stat5 leads to reduced
expression of Bcl-xL and increased sensitivity to apoptosis
[personal communication, Yang X, Clark SS]. It remains to
be seen whether the Ras-independent antileukemia effect
will be found in other malignant cell types. Although it is
not clear how POH exerts these antileukemia effects in the
absence of an effect on Ras, it is possible that the
monoterpene uses different mechanisms to exert its anti-
cancer activity on different types of malignant cells.
Alternatively, as the half-life of farnesylated Ras is more
than 24 hours metabolic turnover of human c-rasH p21
protein of EJ bladder carcinoma and its normal cellular and
viral homologs [40], the anticarcinoma activity that has been
associated with loss of Ras prenylation might arise only
after longer (ca 48 hours) exposure to POH. Shorter
treatment with POH (4-16 hours), as reported in the
leukemia model, would not be expected to significantly
affect Ras prenylation or activity. Whether via change in
prenylation or via other signaling-related activities, POH
treatment alters the expression of a limited number of
different gene products in treated tumors and leukemias
[7,8]. Together, these observations raise the possibility that
POH can exert anticancer activity in both a Ras-dependent
and a Ras-independent fashion.
Our experiments [15] showed that in vitro treatment of
POH consistently inhibited proliferation, produced marked
changes in cell morphology, inhibited protein synthesis,
caused marked alteration in membrane permeability and
drastic changes in the cytoarchitecture of C6, U87MG, and
A172 cells. We have previously shown that in in vivo
treatment of glioblastoma cells, POH showed inhibition of
cell migration and antimetastatic activity in the model of the
chick embryo with C6 cell line [39]. Such results indicate the
chemotherapeutic action of POH by promoting cytotoxicity
and arresting migration of murine and human glioblastoma
cell lines. Also, another study of our group [14] showed that
treatment of both human primary cultures and established
cell lines of GBM (U87 and A172) with 1 mmol/L of POH
led to marked alterations in cell morphology, decrease in cell
viability, and death by apoptosis.
5. Perillyl alcohol—clinical trials
Because Ras activation plays an important role in the
pathogenesis of GBM [13], it makes sense that inhibition of
elevated Ras activity represents a pharmacological-based
approach for a potential treatment of these tumors. Perillyl
alcohol inhibits Ras/MAPK signaling pathway [10], making
Fig. 1. A: Preoperative MRI revealing the extent of the tumor in the right hemisphere. B and C: Glioblastoma. Dense cellularity and palisading nuclei, ischemic
area, and necrosis.
C.O. da Fonseca et al. / Surgical Neurology 65 (2006) S1:2–S1:9S1:6
it feasible to begin considering its use for patients with
relapsed GBM. Phase I and phase II trial protocols in humans
indicate that oral administration of POH affects mainly the
gastrointestinal tract, causing nausea, vomiting, and diarrhea.
No evidence of hepatic, renal, or neurobiological toxicity has
been observed [2,3,18,25,27], and the maximum tolerated
dose determined was 8.4 g/M2 per day delivered orally in 4
doses. Modest clinical responses were attained in trials.
Efforts are now directed toward building high-dose formu-
lations of POH free of gastrointestinal toxicity.
In accordance with these directions, we are developing a
new method for POH delivery, that is, POH inhalation (US
Patent Application 20040087651, May 6, 2004). This
intranasal delivery allows POH to cross the blood-brain
barrier and reach the CNS, eliminating the need for systemic
Fig. 2. Computed tomography scans showing a response to intranasal delivery of
tumor size between the initial computed tomography scan (A) and the follow-up
delivery and reducing the side effects. Based on the favorable
therapeutic ratio observed in vitro and in vivo treatment,
commercial availability, low cost, and low toxicity, we are
developing clinical trial phase II that delivers POH by
inhalation in patients with relapsed GBM. This trial was
approved by the Brazilian Committee of Ethics and Research
(CONEP 9681 no. 25000.009267/2004-25, July 12, 2004). In
4 patients with relapsed GBM, POH was administered by
inhalation 4 times daily. Dose escalation was carried out
according to a standard phase II design. The starting dose was
based on experience obtained with POH given on a
continuous 3-times-a-day schedule. The patient was treated
and evaluated for 4 weeks at each dose level before dose
escalation. Dose-limiting toxicity was defined as any toxicity
of grade 3 or higher according to the National Cancer Institute
POH treatment in a patient with relapsed glioblastoma. Note the decrease in
performed after 3 months (B) and after 5 months (C) of treatment.
C.O. da Fonseca et al. / Surgical Neurology 65 (2006) S1:2–S1:9 S1:7
common toxicity criteria that occurred within the first
4 weeks on study with duration, grade 2 or higher diarrhea
of days’ or more duration, grade 2 or higher creatinine, and
patient refusal to continue on therapy because of drug
intolerance regardless of the grade of toxicity. Hematological
and nonhematological parameters were monitored each
week. The patient remained at home, receiving 3 meals a
day. After 3 months of POH treatment, 3 patients showed
stable tumor size, suggesting an inhibition of tumor growth.
Brain computed tomography scans of a patient who was
under treatment for 5 months showed a decrease in tumor size
(Figs. 1 and 2); as these patients underwent radiotherapy
before beginning POH treatment, we are still unable to
conclude whether the effects observed in the patients are due
to a direct or an adjuvant effect of POH.
References
[1] Ariazi EA, Satomi Y, Elis MJ, Haag JD, Shi W, Satler CA, Gould MN.
Activation of the transforming growth factor beta signaling pathway
and induction of cytostasis and apoptosis in mammary carcinomas
treated with the anticancer agent perillyl alcohol. Cancer Res 1999;
59:1917-28.
[2] Azzoli CG, Miller VA, Ng KK, Krug LM, Spriggs DR, Tong WP,
Riedel ER, Kris MGA. Phase I trial of perillyl alcohol in patients with
advanced solid tumors. Cancer Chemother Pharmacol 2003;51:493 -8.
[3] Bailey HH, Levy D, Harris LS, Schink JC, Foss F, Wadler AS. Phase
II trial of daily perillyl alcohol in patients with advanced ovarian
cancer. East Coop Oncol 2002;3:464 -8.
[4] Belanger JT. Perillyl alcohol: applications in oncology. Altern Med
Rev 1998;3:445-8.
[5] Benedetti S, Pirola B, Pollo B, Magrassi L, Bruzzone MG, Rigamonti
D, Galli R, Selleri S, Di Meco F, De Fraja C, Vescovi A, Cattaneo E,
Finocchiaro G. Gene therapy of experimental brain tumors using
neural progenitor cells. Nat Med 2000;6:447 -50.
[6] Benjamin R, Capparella J, Brown A. Classification of glioblastoma
multiforme in adults by molecular genetics. Cancer 2003;9:82 -90.
[7] Clark SS, Perman SM, Sahin MB, Jenkins GJ, Elegbede JA.
Antileukemia activity of perillyl alcohol (POH): uncoupling apoptosis
from G0/G1 arrest suggests that the primary effect of POH on Bcr/Abl
cells is to induce cell arrest. Leukemia 2002;16:213 -22.
[8] Clark SS, Zhong L, Filiaut D, Perman SM, Ren Z, Gould M, Yang X.
Anti-leukemia effect of perillyl alcohol in Bcr/Abl-transformed cells
indirectly inhibits signaling through Mek in Ras- and Raf-independent
fashion. Clin Cancer Res 2003;9:4494-504.
[9] Clark SS, Zhong L, Perman SM, Ren Z, Gould MN, Yang X. Anti-
leukemia effect of perillyl alcohol (POH): POH indirectly inhibits
signaling through the Erk pathway in a Ras- and Raf-independent
fashion. Clin Cancer Res 2003;9:4494-504.
[10] Crowell PL, Ren Z, Vedejs E, Gould MN. Structure-activity relation-
ships among monoterpene inhibitors of protein isoprenylation and cell
proliferation. Biochem Pharmacol 1994;47:1405-15.
[11] Ding H, Roncari L, Shannon P, Wu X, Lau N, Karaskova J, Gutmann
DH, Squire JA, Nagy A, Guha A. Astrocyte-specific expression of
activated p21-ras results in malignant astrocytoma formation in a
transgenic mouse model of human gliomas. Cancer Res 2001;61:
3826-36.
[12] Feldkamp MM, Lau N, Guha A. Growth inhibition of astrocytoma
cells by farnesyl transferase inhibitors is mediated by a combination of
anti-proliferative, proapoptoic and anti-angiogenic effects. Oncogene
1999;18:7514-26.
[13] Feldkamp MM, Lau N, Roncari L, Guha A. Isotype-specific
Ras.GTP-levels predict the efficacy of farnesyl transferase inhibitors
against human astrocytomas regardless of Ras mutational status.
Cancer Res 2001;61:4425-31.
[14] Fernandes J, da Fonseca CO, Teixeira A, Gatass CR. Perillyl alcohol
induces apoptosis in human glioblastoma multiforme cells. Oncol Rep
2005;13:943 -7.
[15] Fonseca CO. Effects of in vivo and in vitro treatment of monoterpene
perillyl alcohol on proliferation and gene expression control of high
grade gliomas. Arq Neuro-Psiquiatr 2004;62(no. 4):1117 -8.
[16] Goldstein JL, Brown MS. Regulation of the mevalonate pathway.
Nature 1990;343:425-30.
[17] Guha A, Feldkamp MM, Lau N, Boss G, Pawson A. Proliferation of
human malignant astrocytomas are dependent on Ras activation.
Oncogen 1997;15:2755-65.
[18] Hudes GR, Szarka CE, Adams A, Ranganathan S, McCauley RA,
Weiner LM, Gallo JM. Phase I pharmacokinetic trial of perillyl
alcohol in patients with refractory solid malignancies. Clin Cancer Res
2000;6:3071 -80.
[19] Itoh T, Kaibuchi K, Masuda T, Yamamoto T, Matsuura Y, Maeda A,
Shimizu K, Takai Y. The post-translational processing of ras p21 is
critical for its stimulation of mitogen-activated protein kinase. J Biol
Chem 1993;15:3025-8.
[20] Jeffers L. The effect of perillyl alcohol on the proliferation of human
prostatic cell lines. Proc Am Assoc Cancer Res 1995;36:303.
[21] Jiang K, Coppola D, Crespo NC, Nicosia SV, Hamilton AD, Sebti
SM, Cheng JQ. The phosphoinositide 3-OH kinase/AKT2 pathway as
a critical target for farnesyltransferase inhibitor-induced apoptosis.
Mol Cell Biol 2000;20:139 -48.
[22] Karp JE, Kaufmann SH, Adjei AA, Lancet JE, Wright JJ, End DW.
Current status of clinical trials of farnesyltransferase inhibitors. Curr
Opin Oncol 2001;3:470 -6.
[23] Kubiatowski T, Jang T, Lachyankar MB, et al. Association of
increased phosphatidylinositol-3-kinase signaling with increased
invasiveness and gelatinase activity in malignant gliomas. J Neurosurg
2001;95:480 -8.
[24] Levin VA. Chemotherapy for brain tumors of astrocytic and
oligodendroglial lineage: the past decade and where we are heading.
Neuro-oncology 1999;1:69-80.
[25] Liu G, Oettel K, Bailey H, Ummersen LV, Tutsch K, Staab MJ,
Horvath D, Alberti D, Arzoomanian R, Rezazadeh H, McGovern J,
Robinson E, DeMets D, Wilding G. Phase II trial of perillyl alcohol
(NSC 641066) administered daily in patients with metastatic androgen
independent prostate cancer. Invest New Drugs 2003;21:367 -72.
[26] Lokker NA, Sullivan CM, Hollenbach SJ, Israel MA, Giese NA.
Platelet-derived growth factor (PDGF) autocrine signaling regulates
survival and mitogenic pathways in glioblastoma cells: evidence that
the novel PDGF-C and PDGF-D ligands may play a role in the
development of brain tumors. Cancer Res 2002;62:3729-35.
[27] Meadows S, Meelkerin D, Berlin J, Bailey H, Thomas J. Phase II trial
of perillyl alcohol with metastatic colorectal cancer. Int J Gastrointest
Cancer 2003;32:125-8.
[28] Mo H, Elson CE. Studies of the isoprenoid-mediated inhibition of
mevalonate synthesis applied to cancer chemotherapy and chemo-
prevention. Exp Biol Med (Maywood) 2004;229:567-85.
[29] Mills JJ, Chari SS, Boyer IJ, Gould MN, Jirtle RL. Induction
apoptosis in liver tumors by the monoterpene perillyl alcohol. Cancer
Res 1995;55:979-83.
[30] Mizoguchi M, Nutt CL, Mohapatra G, Louis DN. Genetic alterations
of phosphoinositide 3-kinase subunit genes in human glioblastomas.
Brain Pathol 2004;14:372-7.
[31] Rajesh D, Stenzel RA, Howard SP. Perillyl alcohol as a radio-/
chemosensitizer in malignant glioma. J Biol Chem 2003;19:
35968-78.
[32] Reddy BS, Wang CX, Samaha H, et al. Chemoprevention of colon
carcinogenesis by dietary perillyl alcohol. Cancer Res 1997;57:420-5.
[33] Sahin MB, Perman SM, Jenkins G, Clark SS. Perillyl alcohol
selectively induces G0/G1 arrest and apoptosis in Bcr/Abl-trans-
formed myeloid cell lines. Leukemia 1999;13:1581-9.
C.O. da Fonseca et al. / Surgical Neurology 65 (2006) S1:2–S1:9S1:8
[34] Shapiro JR. Genetics of nervous system tumors. Hematol Oncol Clin
North Am 2001;15:961 -77.
[35] Shapiro WR, Green SB, Burger PC, Mahaley Jr. MS, Selker RG,
VanGilder JC, Robertson JT, Ransohoff J, Mealey Jr. J, Strike TA,
et al. Randomized trial of three chemotherapy regimens and two
radiotherapy regimens in postoperative treatment of malignant glioma.
J Neurosurg 1989;71:1 -9.
[36] Shi W, Gould MN. Induction of cytostasis in mammary carcinoma
cells treated with the anticancer agent perillyl alcohol. Carcinogenesis
2002;23:131-42.
[37] Smith JS, Tachibana I, Passe SM, Huntley BK, Borell TJ, Iturria N,
Fallon O’JR, Schaefer PL, Scheithauer BW, James CD, Buckner JC,
Jenkins RB. PTEN mutation, EGFR amplification, and outcome in
patients with anaplastic astrocytoma and glioblastoma multiforme. J
Natl Cancer Inst 2001;93:124-1256.
[38] Stayrook KR, McKinzie JH, Burke YD, Burke YA, Crowell PL.
Induction of the apoptosis-promoting protein Bak by perillyl alcohol
in pancreatic ductal adenocarcinoma relative to untransformed ductal
epithelial cells. Carcinogenesis 1997;18:1655-8.
[39] Teruszkin I, Alves S, Silva HN, Curie CM, Bozza M, Fonseca CO, Da
Costa CMG. Effects of perillyl alcohol in glial cell line in vitro and
anti-metastatic activity in chorioallantoic membrane model. Int J Mol
Med 2002;10:785 -8.
[40] Ulsh L, Shih TY. Metabolic turnover of human c-rasH p21 protein of
EJ bladder carcinoma and its normal cellular and viral homologs. Mol
Cell Biol 1984;4:1647-52.
[41] von Deimling M, von Ammon K, Schoenfeld D, Wiestler OD,
Seizinger BR, Louis DN. Subsets of glioblastoma multiforme defined
by molecular genetic analysis. Brain Pathol 1993;3:19 -26.
[42] Yarden Y. The EGFR family and its ligands in human cancer signaling
mechanisms and therapeutic opportunities. Eur J Cancer
2001;37(Suppl. 4):S3-8.
[43] Zhu Y, Parada LF. The molecular and genetic basis of neurological
tumors. Nat Rev Cancer 2002;2:616 -26.
Commentary
This review on malignant gliomas is done extensively
with a remarkable input on the pathways, classifications, and
experimental update on antitumor agent POH. It is already
evident from literature that many genes are involved in
glioma tumorigenesis (chromosomal amplifications and
deletions). Study of gliomagenesis as stated by the authors
may assign a few more candidate genes from genomic
assays. This candidate gene approach may help in the initial
screening but may not be an ideal way. The authors also
claim POH to be a potential chemotherapeutic agent for
GBMs because POH inhibits Ras/MAPK signalling path-
way. Perillyl alcohol, being a natural monoterpene, may be a
better option to use as compared to other agents. The critical
part is its administration on individuals without any adverse
effect even at high dosages.
Several receptor tyrosine kinase inhibitors, including
imatinib mesylate (Gleevec, Novartis, Basel, Switzerland),
gefitinib (Iressa, Astra Zeneca, Wilmington, DE), and
erlotinib (Tarceva, Genentech, South San Francisco, CA),
have entered clinical trials for high-grade glioma patients.
Farnesyl transferase inhibitors, such as tipifarnib (Zarnestra,
Johnson & Johnson, New Brunswick, NJ), which impair
processing of proRas and inhibit the Ras signaling pathway,
have also entered clinical trials for patients with malignant
gliomas. It is clear that glioma cell migration is a complex
combination of multiple molecular processes, including the
alteration of tumor cell adhesion to a modified extracellular
matrix, the secretion of proteases by the cells, and
modifications to the actin cytoskeleton. Potential targets
include growth factor receptors and other protein tyrosine
kinases, internal signal transduction pathways, ras activa-
tion, and matrix metalloprotease activity. Particular inhib-
itors therefore should only be chosen if the target is present
in the tumor tissue, but this is only possible if individual
patients are submitted to the molecular profiling of their
tumors before undergoing any treatment to combat their
migratory glioma cells. That is where the whole genome
scans of individuals become necessary. New molecular
approaches using whole-genome scans of these individual
patients with complete genomic information may help
classify these individuals, as these patients were first
grouped based on World Health Organization classification
and then were subset based on observed genome variation
in that population. These data can help in classifying the
metastatic phenotype and also in evaluating individuals
with certain genotypes for any adverse effect of the drug,
thus avoiding the risk.
Radhika Gade Andavolu, PhD, MBA
Genetics Research Laboratories
Eisenhower Medical Center
Rancho Mirage, CA 92270, USA
This article by da Fonseca et al outlines some of the
recent advances in the molecular genetics of malignant
gliomas. In particular, the authors present a review of the
advances in our understanding of the Ras signaling
pathway and suggest that perillyl alcohol (POH) may be
an effective antitumor agent by inhibiting the Ras
pathway. The authors describe in detail various aspects
pertinent to Ras signaling and then proceed from there to
further describe the rationale of using POH against solid
tumors. They summarize some previously published work
from their own laboratory to support the use of POH for
the treatment of glioblastoma and review other phases 1
and 2 clinical trials of POH for other cancers. In this
article, the authors present some interim results (n = 4) of
a phase 2 trial using intranasal delivery of POH in patients
with recurrent glioblastoma. They show computed tomo-
graphic scans of 1 patient with an apparent radiographic
response after POH treatment, although they concede that
this could simply be a delayed effect of the patient’s prior
radiation therapy.
In general, I think the topic of glioma molecular genetics
is an important subject for clinical neurosurgeons and neuro-
oncologists. Furthermore, the introduction of new therapeu-
tic agents would be of interest to the field. Although the
numbers reported in this clinical study are small (n = 4) and
Top Related