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.

lovisorlando@wnetrj.com.br (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.

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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