Prevention of head and neck cancer: Current status and future prospects

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Prevention of Head and NeckCancer: Current Status and Future

Prospects

S urgery, radiation therapy, and chemotherapy have been the cor-nerstone of management of most neoplastic processes for almostall of the previous century. Although the former two modalities

re critical in providing local control of a disease, chemotherapy, whileenerally effective in providing systemic control, has been fraught withoxicities. Additionally, treatments with chemotherapeutic agents have aimited appeal due to their nonspecific mode of action. It is only in the lasteveral years that emphasis has been placed in identifying and targetingpecific molecular abnormalities in hematological malignancies, as wells solid tumors. There is now a better understanding of the biology ofell cycle, as well as perturbations that contribute to carcinogenesis.lthough in its infancy, developments in molecular biology andenetics have already made a significant impact in the management ofalignancies. Efforts in these disciplines are now beginning to come

o fruition, with almost daily reports of biological agents demonstrat-ng activity in several different tumors. It is well recognized, however,hat cancer is a multi-step, sustained disease process which begins as alonal abnormality. A previously normal cell must undergo a series ofenetic alterations, which culminate in a malignant phenotype. Thedentification of tamoxifen as an agent that prevents breast carcinogene-is, and of certain COX-2 inhibitors as effective preventive strategies inolonic polyp formation, has sparked interest in preventive, interruptivend reversal strategies in head and neck cancer as well.Squamous cell cancer of the head and neck (SCCHN) is a significantublic health problem worldwide, especially in India and Southeast Asia,here this disease accounts for 25% of all malignancies.1 Although there

re approximately 500,000 cases annually worldwide,1 in the Unitedtates alone, there are approximately 40,000 new cases, with 12,000

2,3
eaths annually. Even though significant advances have been made in
urr Probl Cancer 2004;28:265-286.147-0272/2004/$30.00 � 0oi:10.1016/j.currproblcancer.2004.05.003

urr Probl Cancer, September/October 2004 265

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he surgical, radiation therapy, and chemotherapeutic approaches to theanagement of this disease, the anatomical complexity of the head and

eck region almost invariably portends some degree of functionalmpairment. Despite adequate, definitive therapy, a well-acknowledgedonsequence of primary aerodigestive tract malignancies is the sustained,ignificant risk for the development of second primary tumors (SPTs).econd primary tumors of the upper aerodigestive tract and lung occurith significantly higher frequency in this patient population, at a rate of

pproximately 3 to 4% per year.3

The importance of SPTs in determining outcome was reported in the980s, when Vikram and coworkers4 observed that adequately treatedead and neck cancer patients continued to succumb to second malignan-ies. Similar findings have been reported by Cooper and coworkers,5 whooted that despite improvements in the local management of head andeck cancer, overall survival remained dismal due to the development ofecond malignancies. The above observations have been ratified by thosef Licciardello and coworkers.6 A review of the SEER database hashown a lifetime risk of SPTs of up to 25-40% in patients with a prioristory of laryngeal cancer.7,8 From a clinical and an epidemiologicaltandpoint, however, it is important to note that smoking cessation doesesult in a significant reduction in the incidence of SPTs.9-14

olecular Biology of Head and Neckarcinogenesis

nitially identified as activated cellular genes in certain malignancies, onlyfew members of this class have been directly implicated in tumorigen-

sis.15 Cyclin D1 overexpression has been specifically identified inCCHN, and its amplification results from abnormalities (amplification)

n chromosome 11q13. Up to 30% of SCCHN are associated withmplification of cyclin D1.16-18 A p53 analog (p40/51/63) has beendentified as a protooncogene and found to be frequently amplified inquamous cell cancers.19 Amplification at this locus correlates with bothncreased mRNA transcription and increased protein levels.20

Other, less well established perturbations in SCCHN include those inhe ras pathway. The eukaryotic transcription factor (IF4E) binds toRNA, and its overexpression results in upregulation of proteins essen-

ial for cell growth and division.21-25 Elevated protein levels can contrib-te to oncogenic transformation, as well as the stimulation of angiogen-sis through the elaboration of vascular endothelial growth factor.26 Aeparate, distinct class of gene products overexpressed in SCCHN

27
ncludes COX-2. Two separate isoforms of COX (1 and 2) exist.
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lthough COX-1 is constitutive and facilitates the synthesis of prosta-landins for several physiological functions, COX-2 is inducible and notsually detected in normal tissues. Its expression, however, is enhanced inCCHN28 from a variety of stimuli, including cytokines, growth factors,nd oncogenes.29-35 Interestingly, wild-type p53 suppresses COX-2unction, whereas mutations in p53 (detected commonly in precancerousesions as well as SCCHN36,37) result in a loss of this suppression.ncreased expression of COX-2 is seen in both precancerous lesions andCCHN.38-50

Analogous to gain in function with the acquisition of oncogene activity,he loss of critical tumor suppressor function is a well studied mechanismf tumorigenesis. The most commonly characterized abnormality inCCHN involves chromosome 9p21 to 22.51,52 Abnormalities in thisegion are also present in a high frequency in dysplasia as well asarcinoma in situ.53 The specific, well-characterized abnormality at thisocus consists of loss of p16 function, which normally serves to abrogatehe cyclin D1 pathway. Therefore, loss of p16 function facilitatesncontrolled cell proliferation. Although only 10 to 15% of head and neckumors were subsequently found to be associated with p16 abnormalitiespecifically,54,55 alternative mechanisms of p16 function inhibition wereater identified, implicating p16 in a significant proportion of SCCHN.ne such perturbation was identified in p16�, which resides upstream of16 on chromosome 9p21.56 This transcript encodes for a protein throughn alternate reading frame (ARF) which normally functions to bind toDM2, which in turn inhibits p53 degradation.57,58 As would be

xpected, introduction of p16 or p16� into SCCHN cell lines results inotent growth inhibition.59 Izzo and coworkers reported on the increasedisk for the development of invasive disease in lesions associated withltered cyclin D1 expression, during and after treatment of patients withbiochemopreventive approach consisting of 13-cis-retinoic acid, �-in-

erferon, and �-tocopherol.60

The p53 gene resides on chromosome 17p, which is frequently deletedn up to 60% of SCCHN.61 Interestingly, p53 mutations occur much laterhan 17p abnormalities, suggesting that an alternate tumor suppressorene, not yet identified, probably resides on chromosome 17 and is likelynvolved in early tumorigenesis. Shin and coworkers proposed that p53lterations occur early in tumorigenesis and may serve as an importantntermediate biomarker of chemopreventive efforts. They performedmmunohistochemical analyses for p53 expression in 33 patients withCCHN and identified altered expression in 45% of the malignant lesions,
5% of the dysplastic lesions, 29% of the hyperplastic lesions, 21% of the

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ormal epithelium adjacent to the tumors, but in none of the controlsithout a diagnosis of SCCHN.62 Lippman and coworkers reported

imilarly on different levels of p53 expression in subjects with oralremalignant lesions.63 p53 expression was not altered in any of theontrols, but was notably altered in the deeper layers of the epithelium ofhe subjects with the premalignant lesions. Also, there was noted to be aignificant association between the level of p53 expression and theistologic grade. In a cohort of patients with SCCHN, p53 expression wasssociated with a worse outcome following definitive local therapy.64

hin and coworkers analyzed tumor specimens for p53 expression in 69atients with SCCHN treated with definitive surgery or radiation therapyt MD Anderson Cancer Center (MDACC) between 1980 and 1983. Theyoted that p53 expression was associated with an earlier tumor recurrenceate, as well as second primary tumor development in this patientopulation. Other loss of function abnormalities have also been associatedith chromosome 3p65,66 abnormalities, as well as loss of chromosome0q.67,68

The principal risk factors for the development of head and neck cancerre exposure to tobacco and alcohol. However, the majority of subjectsxposed to these carcinogens do not develop malignancies. There areresumably underlying genetic mechanisms that predispose certain indi-iduals to carcinogenesis. Genetic predisposition is further supported byhe observation that first-degree relatives of patients who have notecessarily been exposed to the carcinogens remain at an approximatelyhree- to eightfold higher risk of developing malignancies than thenrelated population.69-72 Advances in molecular biology now allow forsolation of genetic material (DNA) from the primary tumor for thessessment of specific molecular abnormalities (deletions/amplifications/haracterization of candidate oncogenes). Elucidation of the stepwiseequence of events in carcinogenesis now presents us with unique targetsor preventive, diagnostic, and therapeutic strategies.Several carcinogens have been identified in tobacco, and it is now well

stablished that there are differences in the individual ability to metabo-ize these products.73 It is conceivable that individuals exposed toersistently elevated levels of carcinogens, not just tobacco products, forprolonged period of time are at greater risk for tumorigenesis. Phase I

n the metabolism of tobacco components involves the activation ofrecursors through the CYP, cytochrome P450, enzyme systems, whereashase II involves detoxification of these products through conjugationechanisms. Although CYP-related polymorphisms exist, they have not

een found to significantly contribute to differences in risks for


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CCHN.74-84 It is the phase II mechanisms which likely play a moremportant role in accounting for differential risks, as described below.The glutathione S-transferase (GST) genes are responsible for the

onjugation steps that abrogate tobacco toxin injury to DNA. Althoughhere are several components of the GST complex, GSTM1 and GSTT1re two well studied isoenzymes. It is now well recognized that there existndividual, racial, and ethnic differences in expressivity of this complex.ndividuals possessing the null genotype, in which the enzyme isnderexpressed or not expressed at all, are at a higher risk for theevelopment of squamous cell malignancies. This risk is higher than thatn controls when either of the isoenzymes is underexpressed and almostwice as high when both are underexpressed.85-95

hemopreventionA term first coined by Sporn and coworkers in 1976, chemoprevention

mplies the use of a natural or synthetic chemical for the reversal,uppression, or prevention of conversion of a premalignant lesion to annvasive form.96 This concept is of particular relevance to the uppererodigestive tract, where synchronous as well as metachronous lesionsre a significant cause of morbidity and mortality. The term “fieldancerization” was introduced by Slaughter and coworkers, who attrib-ted this to a field defect in the aerodigestive tract, which allowed forndependent generation of neoplastic clones at a number of sites.97

ladder cancer is the prototype of such skip lesions, but in SCCHN,imilar skip lesions have been identified. Interestingly, lesions surroundedy normal intervening mucosa share identical genetic abnormalities withhe primary or dominant invasive lesion. It is therefore hypothesized thatsingle cell first develops a clonal abnormality after sustaining a criticalenetic insult. It then migrates through the mucosa and deposits nests ofells in the adjacent regions, which tend to grow independently atifferent rates. Over a period of time, the dominant lesion becomesanifest as a malignancy, while others, although they share the same

enetic abnormalities, are not yet fully transformed.In accordance with this hypothesis, observations have been made byifferent investigators which add credence to the concept of fieldancerization. Califano and coworkers evaluated 18 patients with cervicalodal metastases (of squamous histology) from unknown head and neckrimary sites.98 In 10 of these patients, the investigators were able toemonstrate genetic abnormalities identical to the cervical lymph nodaletastases in benign appearing mucosa, and 3 of these patients eventually

eveloped invasive cancers.


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linical Trials of Chemoprevention in Oral PremalignantesionsOral premalignant lesions have provided for an excellent model for

ssessing the efficacy of preventive agents. Several trials have beenompleted in patients with leukoplakia and erythroplakia. Leukoplakia isssociated with a less than 5% risk for malignant transformation, aspposed to erythroplakia, which carries a higher incidence, 30-50%, ofalignant transformation. The former has an approximately 30-50% rate

f spontaneous regression, in contrast to erythroplakia and dysplasticesions, where this rate is only 5%.104

The premise of chemopreventive efforts in the upper aerodigestive tractas been targeting of critical molecular lesions based on the aforemen-ioned observations. Although a number of genetic lesions, as outlinedarlier, have been identified in populations at risk for the development ofCCHN, and several others are yet to be discovered, most of the effortsave been targeted on the use of retinoids. Incidental observations wererst made on cattle deprived of vitamin A by Wolbach and Howe99; thesenimals had a much higher incidence of lung cancer and upper aerodi-estive tract malignancies.Retinoids have diverse biological effects, including the modulation ofifferentation, proliferation, and apoptosis in normal as well as malignantissues (Fig 1).100,101 Both the retinoid ligand and the receptor displayignificant diversity.102,103 There are at least two broad categories ofuclear retinoid receptors, RARs and RXRs, and each of them haveeveral subclasses. RXR ligands are more versatile since they can bind to

IG 1. Molecular model of retinoid chemoprevention. ATRA, all-trans-retinoic acid; 13cRA, 13-cisetinoic acid; RAR-beta, retinoic acid receptor beta. Adapted with permission from Cancer, Principlesnd Practice of Oncology, DeVita, Hellman, Rosenberg, Lippincott, Wilkins and Williams, 6th edition,587, 2001.

host of receptors, whereas RARs can only form heterodimers with


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XRs. Some of the retinoids tested in clinical trials include 13-cis-etinoic acid (13cRA), 9-cis-retinoic acid, 4-N-4hydroxyphenyl reti-amide (fenretinide), bexarotene, and etretinate (Fig 2).Vitamin A Analogs. One of the pivotal studies testing the efficacy of

etinoids was completed in 1986 by Hong and coworkers.105 Patients withral premalignant lesions were randomized to high doses of 13-cis-etinoic acid at 1 to 2 mg/kg/day versus placebo. There was a 67%esponse rate with the use of the retinoid. However, toxicities (cheilitis,ypertriglyceridemia, erythema) were prohibitive during therapy, andelapses occurred frequently following cessation of therapy. This led to aeparate maintenance trial, in which patients were randomized, after 3onths of high-dose 13-cis-retinoic acid at 1.5 mg/kg/day, to either low

oses of maintenance 13-cis-retinoic acid at 0.5 mg/kg/day or �-carotenet 30 mg/day.106 This trial showed a significant advantage for mainte-ance with 13-cis-retinoic acid in comparison to �-carotene. However,ong-term follow up has failed to show an advantage for either

IG 2. Structures of selected retinoids and the carotenoid, beta-carotene. Retinol, 13-cis retinoic acid,nd beta-carotene are naturally occurring; 4HPR and etretinate are synthetic retinoids. Adapted withermission from Cancer, Principles and Practice of Oncology, DeVita, Hellman, Rosenberg, Lippin-ott, Wilkins and Williams, 6th edition, p576, 2001.

ntervention.


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Papadimitrakopolou and coworkers107 conducted a trial in which 36atients with advanced premalignant lesions (defined as moderate toevere dysplasia or carcinoma in situ) of the larynx and oral cavity werereated with a combination of �-interferon (3 � 106 IU/m2 three times aeek sq) with 13-cis-retinoic acid at 50 mg/m2/day and �-tocopherol at200 IU daily. Interestingly, almost 50% of the patients with laryngealesions demonstrated a complete reversal of their premalignant lesion, andeveral patients had long-lasting durable remissions for several months.atients with oral cavity lesions, however, did not have durable responses.n fact, disease progression occurred commonly.Stich and coworkers108 evaluated the effects of vitamin A in 65 patientsith oral leukoplakia in betel nut chewers and tobacco users in India.hirty-one of these patients were randomized to receive 100,000 IU ofitamin A and the other 34 placebo. Although 12 patients in the treatmentrm had responses, with no progressions reported, only 1 patient in thelacebo arm had a spontaneous regression, and the majority progressed.n a separate trial, the same investigators conducted a randomized studyf vitamin A administration at doses of 200,000 IU a week compared withlacebo in patients with leukoplakia. They reported a near 100%uppression in the incidence of leukoplakia, with almost a 50% completeesponse rate. Results of this study, as well as others, led to the conductf EUROSCAN, a large randomized study in patients with a prioriagnosis of head and neck cancer or lung cancer (see below). The first ofhree arms was randomized to the administration of vitamin A in doses of00,000 IU daily for the first year, followed by half the doses in theecond year. A second arm consisted of the above intervention inombination with N-acetylcysteine (600 mg) daily for 2 years, and a thirdrm of no intervention. The primary endpoint was assessment of the ratef SPT development. However, this trial failed to a show differences inecurrence rates, SPT development, or overall death rate.109

Chiesa and coworkers110,111 randomized 61 patients with oral prema-ignant lesions to either 4 months of etretinate (4-N-4hydroxyphenyletinamide/fenretinide) at 40 mg/day versus placebo. Major responsesere seen in 27 of the 31 patients treated with etretinate versus 5 of the0 patients in the placebo arm. Toxicity was minimal.The observations from some of the studies mentioned above are

ppealing from the standpoint of being able to identify an aberrant targetretinoic acid receptor) in SCCHN and being able to then disrupt theownstream signaling mediated by ligands (retinoids). Although theechanisms of vitamin A deficiency-induced carcinogenesis are not

learly understood, it is possible that this family of agents promote growth


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nd differentiation through restoration of retinoic acid receptor-� expres-ion.112 Although a significant advance in our understanding of theechanisms of tumorigenesis in SCCHN, there are several limitations to

ur current approaches, which now warrant more complete, well-de-igned strategies in preventive approaches.Vitamin E. Vitamin E has been known to have antioxidant properties.hile most of the information pertaining to its effects in oral malignan-

ies is available from animal experiments, the mechanisms by whichitamin E may exert a protective function are speculated to be related tohis agent’s interaction with the immune system.113 Animal experimentsuggest a possible immunomodulatory role for �-tocopherol against bothacterial and viral infections.114 There are also epidemiological studies,imilar to those with selenium, which suggest a protective role for vitamin

as well, in geographic regions where the consumption of this agent isigher compared with other populations. These observations haverompted the investigation of vitamin E’s protective role against malig-ancies. There are limited data from human trials,115 in one of which 43atients with oral leukoplakia were treated with 400 IU of vitamin E twiceaily for 24 weeks. There were histological responses in 21% and clinicalegressions in the lesions in an additional 46% of the patients.116

Selenium. There are epidemiological observations that death rates fromalignancies, including SCCHN, are low in geographic regions of theorld where the soil is rich in selenium.117 The use of selenium inCCHN has been associated with an enhancement of cell-mediated

mmune responses. There are currently ongoing clinical trials evaluatinghe role of selenium in the adjuvant setting in lung cancer.118

linical Trials of Second Primary TumorhemopreventionFirst-Generation Trials. As mentioned previously, even after a curative

reatment, upper aerodigestive tract malignancies are associated with aignificant risk of recurrence; this is not limited to the local mucosa, buto the entire squamous epithelium which has been exposed to the sameisk factors.With this understanding, and based on their own observations of

eversibility of oral premalignant lesions, Hong and coworkers119 hypoth-sized that an intervention in upper aerodigestive tract malignanciesollowing adequate treatment (surgery/XRT) of SCCHN would be asso-iated with a reduction in the risk of second primary tumors. A doublelind, placebo-controlled trial was conducted of high-dose 13-cis-retinoic
cid (50-100 mg/m2/day) or placebo for 12 months. After a median

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ollow up of at least 32 months, there were significantly fewer SPTs in thereatment arm (4%) than in the placebo arm (24%; P � .005). In the samenalysis, the majority of the SPTs occurred in the epithelium at risk fromrior exposure to tobacco (93%), including the upper aerodigestive tract,ungs, and esophagus. However, similar to the observations made inarlier trials, toxicities (dermatological, gastrointestinal) were limiting.pproximately one third of the patients required discontinuation of

herapy due to these toxicities. Additionally, overall survival was notmpacted.After a median follow up of 4.5 years in this trial, Benner and

oworkers120 determined from analyzing these data that treated patientsontinued to have significantly fewer SPTs (14%) versus the placebo arm31%; P � .042). Most of this advantage actually occurred in thepithelium at risk, rather than at distant sites, and although this differenceersisted for as long as 2 years, it was no longer significant at 3 years.Bolla and coworkers,121 with a similar hypothesis, assessed the efficacyf etretinate in patients with a prior history of SCCHN. Patients wereandomized to either the retinoid at 50 mg/day for 1 month followed byalf the dose for 24 months or placebo. After a median follow up of 41onths, there was no difference in outcome, including the incidence of

econd primary tumors.Second-Generation Trials. Based on observations by Hong et al, asell as those by Pastorino and coworkers109 in lung cancer, the Europeanrganization for Research and Treatment of Cancer initiated the EURO-CAN trial. This was a large, multiinstitutional randomized study of 2592atients with completely resected/adequately treated stage I and II nonmall cell lung cancer as well as stages I, II, and III SCCHN. In a 2 � 2actorial design, patients were randomized on retinyl palmitate at 300,000U daily for 1 year followed by 150,000 IU in the second year, or to-acetylcysteine at 600 mg/d ayfor 2 years, or to both compounds, or too intervention at all. After a median follow up of 49 months, there wereo differences in outcome in any of the arms. The authors concluded thatnterventions consisting of 2 years of either retnyl palmitate or N-acetyl-ysteine did not provide protection from SPTs.In 1991, the University of Texas MD Anderson Cancer Center launchedrandomized, double blind trial of low-dose 13-cis-retinoic acid (30g/day) versus placebo to prevent SPTs in patients adequately treated for

tages I and II SCCHN. Of the 1384 patients randomized, 1189 wereonsidered eligible and evaluable. This study indicated that although thereas a transient protective effect of low-dose 13cRA on prevention of

ecurrence while the patients were on the study, there was no impact on


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he primary endpoint of second primary tumor prevention between thereatment and the placebo arm. Interestingly, however, continued smok-ng correlated with a poor outcome in these patients in terms of both ratef SPT development and survival.122,123

In a phase II clinical trial of biochemoprevention, Shin and coworkersdministered isoretinoin (50 mg/m2 orally daily), �-tocopherol (1200 IUrally daily), and IFN-� (3 � 106 units subcutaneously three times aeek) for 12 months in patients with treated locally advanced SCCHN.fter a median follow up of 49.4 months, the DFS rates at 1, 3, and 5ears were reported at 89, 78, and 74%, respectively. The overall survivalates at 1, 3, and 5 years were 98, 89, and 81%, respectively. Eighty-sixercent of the patients completed their therapy, suggesting acceptableolerability and compliance. Toxicities included weakness, peripheraleuropathy, as well as one report of optic neuritis. Based on these results,phase III randomized trial has been designed with reduced doses of

soretinoin (40 mg/m2 orally daily) and IFN-� (2 � 106 units subcuta-eously three times a week).124

uture DirectionsThere is now irrefutable evidence that one of the most effectivereventive interventions, particularly for SCCHN, is cigarette smokingessation. It is imperative that this important aspect be continuallymphasized by health care professionals to their patients. There isounting evidence that even in patients who have developed tobacco-

elated malignancies, smoking cessation results in an improved outcomeompared with those subjects who continue to smoke.For patients at risk for the development of SCCHN, as well as otherpper aerodigestive tract malignancies, a large-scale, concerted effort isecessary to better elucidate the mechanisms underlying carcinogenesis,s well as the development of targeted/molecular interventions whichould permit reversal or abrogation of the carcinogenic process. Al-

hough extensive work has been done to address some of these issues, asentioned previously, this effort should be used as a foundation for the

esign and execution of well-designed international trials.The following is an outline of potential candidate agents, as well asrospective studies.H-ras, a member of the ras gene family, is commonly altered in a varietyf solid tumors, including SCCHN. Approximately 30% of leukoplakiaesions and almost 50% of invasive SCCHN are associated with H-ras
utations. The role of farnesyl transferase inhibitors, which inhibit the

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nzymatic activation of H-ras, is currently being evaluated in phase I andI clinical trials, in both SCCHN and lung cancer.125-127

Cyclo-oxygenase, an enzyme that catalyzes the the synthesis of pros-aglandins, has been found to be overexpressed in a variety of premalig-ant and malignant states,128 including leukoplakia and SCCHN. In-reased levels of COX-2 may contribute to carcinogenesis through aariety of mechanisms, including the modulation of apoptosis andngiogenesis (Figs 3 and 4). In experimental models, COX-2 inhibitorsnhibit the carcinogenic process and suppress metastases, as well asnhance the effects of radiation therapy and chemotherapy. Lee andoworkers determined that at least seven SCCHN cell lines constitutivelyxpressed COX-2. They then cultured these cell lines in the presence ofS-398 and indomethacin, both of which are known to be inhibitors of

he COX-2 enzyme. After 72 h, there was a significant dose-dependentnhibition of cell growth, as well as a significant increase in the numberf cells that transitioned into the G0/G1 phase. Likewise, there was a

IG 3. COX 1 and 2 mediated prostaglandin synthesis. Adapted with permission from Dannenbergt al., 2002.

ignificant decrease in the number of cells in the S phase compared with


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hat in controls. Additionally, NS-398 was capable of inducing apoptosisn two of the three cell lines.Wang and coworkers demonstrated a significant reduction of newasculature development at tumor sites in mice treated with celecoxib.129

dditionally, there are reports of possible synergism between COX-2nhibitors and agents that target the epidermal growth factor receptorEGFR)130 (see below).The EGFR is frequently overexpressed in SCCHN. 131 Additionally,verexpression of EGFR is associated with a poor outcome, including thebservation of more aggressive growth in such tumors.132 The EGFRamily consists of at least four closely related growth factors, includingGFR/Her-1/erb-B1, Her-2/erb-B2/neu, Her-3/erb-B3, and Her-4/erb-B4.igand binding to EGFR results in the initiation of cytoplasmic signaling,hich is mediated through several different effectors, including Ras andI3K (phosphatidylinositiol-3-kinase). Downstream of this activation,ne of the signaling events involves phosphorylation of Akt, which in turnediates its antiapoptotic effect through inactivation (through phosphor-

lation) of BAD. Experiments with agents targeted to the EGFR havehown an abrogation of Akt phosphorylation and therefore increasedytotoxicity. Gupta and coworkers133 observed a significantly enhancedadiation-induced cytotoxic effect in patients with SCCHN when theyere also treated with gefitinib, an agent directed to the EGFR.A combined approach with the use of EGFR inhibitors and a COX-2

nhibitor has been explored by Shin and coworkers.134 The authorsypothesized that this combination may provide synergistic activitygainst SCCHN. In separate experiments, they treated two head and neckancer cell lines with gefitinib, an EGFR tyrosine kinase inhibitor alone,elecoxib alone, or the combination. Synergism with the combination was

134

IG 4. Levels of COX-2 protein are increased in HNSCC, as shown in this experiment, wheremmunoblotting was performed on HNSCC from six patients (odd lanes) and normal oral mucosa fromix healthy volunteers (even lanes). The immunoblot was probed with antibody specific for COX-2.urified ovine COX-2 was used as a standard. Reprinted with permission from Cancer Research.

eported. Based on these encouraging in vitro observations, the


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nvestigators conducted similar experiments in nude mice (xenograftodels) in whom the Tu212 SCCHN cell line was introduced. Four

roups of animals were subjected to gefitinib alone, celecoxib alone, theombination, or a placebo vehicle. Significant synergism was reportedith the combination, leading the investigators to propose phase I/II trialsith this combination.135

Magne and coworkers136 tested the antiproliferative activity of gefitinibgainst SCCHN cell lines and were able to establish increasing activityith higher growth factor receptor levels. They furthered this research to

ssess the specific molecular mechanisms underlying this activity, as wells explored the possibility of synergism between gefitinib and chemo-herapeautic agents. They cultured CAL 33 cells, a SCCHN cell line, withefitinib, followed 48 h later with the addition of 5-FU and cisplatin.lthough gefitinib alone was able to induce accumulation of these cells in

he G0/G1 phase of the cell cycle, with a concomitant increase in p21,27, and BAX as well as a decrease in Akt phosphorylation, the additionf the chemotherapeutic agents resulted in a dramatic increase in caspaseactivity as well.137

Cohen and coworkers recently reported on 47 patients treated forecurrent SCCHN. About 11% of the patients responded to 500 mg ofeftinib administered daily.138 Although dermatological and gastrointes-inal toxicities are prominent, the reports of rare cases of interstitialneumonitis as well may limit the application of this strategy in areventive setting. There are ongoing trials of lower doses of geftinib,hich are not yet mature enough to report on toxicities or efficacy.Alterations in p53 are commonly reported in SCCHN. Due to the poorrognosis associated with p53 mutations, an ongoing trial, ONYX-015,as preliminarily reported on the use of p53 administered with andenoviral vector in patients with oral premalignant lesions. Two of therst 10 patients treated with weekly administration as a mouthwash haveemonstrated resolution of dysplasia.139 An ongoing study is evaluatinghe role of tumoral injections of the adenoviral vector in patients withecurrent SCCHN.With advances in our current understanding of the molecular biology ofCCHN, there are now several more experimental approaches availableith novel agents that may provide for more effective chemopreventive

pproaches. Future trials in chemoprevention will require a betternderstanding of the intermediate biomarkers of response, so thatnterventions can be continued, or interrupted, based on the prognosticnd predictive value of these intermediary biomarkers of response.
owever, it is imperative that the scientific community not repeat some

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f the previous efforts that have resulted in suboptimal results. Singlegent interventions have not necessarily resulted in meaningful outcomes,nd the challenge will be to identify effective combinations of biologicalgents that would be safe, as well as effective in chemoprevention ofalignancies

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Prevention of head and neck cancer: Current status and future prospects