8
Gastroentérologie Clinique et Biologique (2009) 33, 1028—1035 CURRENT TREND Genetic alterations in precancerous pancreatic lesions and their clinical implications Les altérations génétiques dans les lésions précancéreuses pancréatiques et leurs implications en clinique O. Turrini a,b , C. Cano a , A. Legoffic a , J.R. Delpero b , J.C. Dagorn a , J. Iovanna a,a Inserm U624, stress cellulaire, parc scientifique et technologique de Luminy, 163, avenue de Luminy, CP 915, 13288 Marseille cedex 9, France b Département de chirurgie oncologique, institut Paoli-Calmettes, Marseille, France Available online 18 September 2009 Summary Pancreatic adenocarcinoma, with an incidence/death ratio of 0.99, has the worst prognosis of all cancers. Risk factors associated with the sporadic form of pancreatic ade- nocarcinoma are unknown and less than 10% of patients receive curative treatment (surgery associated with radiation therapy or chemotherapy) with a low 5-year survival rate (10 to 20%). In more than 90% of patients, the tumor discovered at diagnosis is not resectable or has already metastasized. Thus, a better understanding of the etiology of pancreatic cancer is essential to identify new prognostic markers and new therapeutic targets. There is a wealth of data on the identification of genetic alterations associated with pancreatic cancer and their role in its development. This review will focus on the current knowledge of genetic alterations associated with two pancreatic lesions that can potentially evolve into pancreatic adenocarcinoma, Pan- creatic Intraepithelial Neoplasia (PanIN) and Intraductal Papillary Mucinous Neoplasm (IPMN). These two lesions share a large panel of typical genetic alterations which are close to those found in pancreatic adenocarcinoma. A better understanding of these alterations may lead to therapeutic targets that could help prevent the progression of PanIN and IPMN to cancer. © 2009 Elsevier Masson SAS. All rights reserved. Résumé Le cancer pancréatique, pour lequel le rapport incidence/décès est de 0,99, est à ce jour le cancer dont le pronostic est le plus sombre. Les facteurs de risque associés à la forme sporadique de l’adénocarcinome pancréatique sont très mal connus et seuls 5 à 10 % des patients atteints sont en mesure de recevoir un traitement curatif (chirurgie associée DOI of original article:10.1016/j.gcb.2009.02.046. Retrouvez la version franc ¸aise de cette mise au point sur Internet à l’adresse www.emconsulte.com/revues/gcb (page d’accueil de Gastroentérologie clinique et biologique). Corresponding author. E-mail address: [email protected] (J. Iovanna). 0399-8320/$ – see front matter © 2009 Elsevier Masson SAS. All rights reserved. doi:10.1016/j.gcb.2009.08.007

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Page 1: Genetic alterations in precancerous pancreatic lesions and their clinical implications

Gastroentérologie Clinique et Biologique (2009) 33, 1028—1035

CURRENT TREND

Genetic alterations in precancerous pancreaticlesions and their clinical implications�

Les altérations génétiques dans les lésions précancéreuses pancréatiqueset leurs implications en clinique

O. Turrini a,b, C. Canoa, A. Legoffica, J.R. Delperob,J.C. Dagorna, J. Iovannaa,∗

a Inserm U624, stress cellulaire, parc scientifique et technologique de Luminy, 163, avenue de Luminy,CP 915, 13288 Marseille cedex 9, Franceb Département de chirurgie oncologique, institut Paoli-Calmettes, Marseille, France

Available online 18 September 2009

Summary Pancreatic adenocarcinoma, with an incidence/death ratio of 0.99, has the worstprognosis of all cancers. Risk factors associated with the sporadic form of pancreatic ade-nocarcinoma are unknown and less than 10% of patients receive curative treatment (surgeryassociated with radiation therapy or chemotherapy) with a low 5-year survival rate (10 to 20%).In more than 90% of patients, the tumor discovered at diagnosis is not resectable or has alreadymetastasized. Thus, a better understanding of the etiology of pancreatic cancer is essentialto identify new prognostic markers and new therapeutic targets. There is a wealth of data onthe identification of genetic alterations associated with pancreatic cancer and their role in itsdevelopment. This review will focus on the current knowledge of genetic alterations associatedwith two pancreatic lesions that can potentially evolve into pancreatic adenocarcinoma, Pan-creatic Intraepithelial Neoplasia (PanIN) and Intraductal Papillary Mucinous Neoplasm (IPMN).These two lesions share a large panel of typical genetic alterations which are close to thosefound in pancreatic adenocarcinoma. A better understanding of these alterations may lead totherapeutic targets that could help prevent the progression of PanIN and IPMN to cancer.© 2009 Elsevier Masson SAS. All rights reserved.

Résumé Le cancer pancréatique, pour lequel le rapport incidence/décès est de 0,99, est àce jour le cancer dont le pronostic est le plus sombre. Les facteurs de risque associés à laforme sporadique de l’adénocarcinome pancréatique sont très mal connus et seuls 5 à 10 %des patients atteints sont en mesure de recevoir un traitement curatif (chirurgie associée

DOI of original article:10.1016/j.gcb.2009.02.046.� Retrouvez la version francaise de cette mise au point sur Internet à l’adresse www.emconsulte.com/revues/gcb

(page d’accueil de Gastroentérologie clinique et biologique).∗ Corresponding author.

E-mail address: [email protected] (J. Iovanna).

0399-8320/$ – see front matter © 2009 Elsevier Masson SAS. All rights reserved.doi:10.1016/j.gcb.2009.08.007

Page 2: Genetic alterations in precancerous pancreatic lesions and their clinical implications

1029

les adénocarcinomes pancréatiques. Certaines d’entre elles pourraient être la base de ciblesthérapeutiques permettant d’éviter la transformation maligne.© 2009 Elsevier Masson SAS. Tous droits réservés.

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Introduction

Cancer of the pancreas is the fourth leading cause ofcancer death in western countries. In France, 7,200 personsdie from this disease every year; in the USA, the annualdeath rate is about 30,000 [1]. With a incidence/deathratio of 0.99, pancreatic cancer is a crucial public healthissue [2]. Although perioperative radiation therapy andchemotherapy be slightly advantageous in terms of survival,there is no proven effect on prognosis [3]. Surgery offersthe only possible cure if resection margins remain healthy.But only 10 to 15% of patients have localized resectabletumors at diagnosis [4] and most operated patients rapidlydevelop locoregional or metastatic disease progression[5]. Screening for pancreatic cancer, which could result inearly diagnosis and thus increase the chances of curativetreatment, is not a currently available practice. There isno known biological or clinical screening test with a provenefficacy and the risk factors associated with the sporadicforms of the disease are unknown. Thus, this tumor has avery poor prognosis. A better understanding of the mecha-nisms of development of pancreatic cancer could probablycontribute identifying new molecular targets and improvingdiagnosis and treatment. A large body of work has beendevoted to the identification of genetic alterations and theirrole in the development of pancreatic cancer. In this review,we will focus on genetic anomalies associated with two wellrecognized precancerous pancreatic lesions, PancreaticIntra-epithelial Neoplasia (PanIN) and Intraductal PapillaryMucinous Neoplasm (IPMN). Both of these anomalies mainlyinvolve either oncogenes or tumor suppressor genes. Onco-genes are genes which favor the expression of malignanttransformation. These genes are either mutated genes ormutation-free genes which are abnormally over-expressed.Tumor suppressor genes are genes which normally protectcells from degeneration. Their mutation or their inhibitionfavors the development of cancer cells.

Pancreatic intra-epithelial neoplasia

Although precursor lesions of pancreatic cancer were welldocumented more than a century ago, a rigorous classifica-

ogio[

ion of these lesions was not obtained until the last decade.orphological analyses of resected pancreatic cancersould suggest that pancreatic adenocarcinomas do notevelop de novo but are a result of a step-wise progressioneading to the generation of invasive lesions [6—8]. Aroundhe late 1990s, a plethora of often biologically impreciseerms were used to describe these lesions. In 2001, an inter-ational agreement [9] was reached on the nomenclaturesed for a vast spectrum of lesions ranging from low-gradeesions (PanIN-1) to in situ carcinomas (PanIN-3), considereds the stage preceding invasion of the neighboring stroma.detailed description of the histological features of PanIN

s beyond the scope of this review. However, it should beoted that the histological progression of PanIN is (with aew exceptions) associated with progressive accumulationf the same molecular anomalies that are observed innvasive cancer [10]. These molecular alterations have beenelpful in demonstrating that PanINs are clonal precursorsf pancreatic adenocarcinoma. Although the exact naturalistory of PanIN has been difficult to establish, theseesions are thought to exist well before the appearancef the adenocarcinoma. Because of the poor prognosisf pancreatic cancer, it is important to obtain a geneticharacterization of the pancreatic tissue harboring PanINesions, and in particular PanIN-3 lesions, before they trans-orm into malignant lesions. The identification of molecularnomalies specific for PanIN would enable early screeningor the pancreatic transformation process and might fur-ish important elements for new targets for therapeutictrategies.

While animal models have not yet been developedor IPMN, several genetically modified mouse models areurrently available for PanIN. These models have shownn the mouse that PanIN lesions develop into pancreaticdenocarcinomas. The process has always involved pan-reatic expression of the mutated KRAS gene [11—13].xpression of the mutated KRAS gene induces formationf PanIN but not its malignant transformation, which isnly obtained after loss of activity of a tumor suppressorene (CDKN2A/p16, SMAD4 or other molecules participat-

Genetic alterations in precancerous pancreatic lesions and their clinical implications

à la radiothérapie ou chimiothérapie périopératoire) avec un taux de survie faible à cinqans (10 à 20 % selon les séries). En effet, plus de 90 % des patients présentent au momentdu diagnostic une tumeur inextirpable ou des métastases. Il est donc impératif de mieuxcomprendre l’étiologie du cancer pancréatique afin d’identifier de nouveaux marqueurs depronostic et de nouvelles cibles thérapeutiques. De nombreuses équipes se sont intéressées àl’identification des altérations génétiques associées au cancer pancréatique et au rôle de cesaltérations dans son développement. Cette revue fait le point des connaissances actuelles surles anomalies génétiques associées aux deux lésions pancréatiques à potentiel malin que sontles néoplasies intra-épithéliales pancréatiques ou PanIN et les tumeurs intracanalaires papil-laires et mucineuses du pancréas ou TIPMP. La plupart des anomalies génétiques détectéessont communes à ces deux situations précancéreuses et diffèrent peu de celles observées dans

ng in the TGF beta and TP53 signaling pathways) [14—20]r in the presence of chronic pancreatic inflammation21].

Page 3: Genetic alterations in precancerous pancreatic lesions and their clinical implications

1

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enetic alterations in pancreatic intra-epithelialeoplasia

he characterization of genetic alterations observed inanIN has progressed further than that for IPMN.

ncogenesrotein ERBB2 encodes tyrosine kinase growth factor recep-or, whose activation induces cell proliferation. ProteinRBB2 is thus considered to be a powerful oncogene. Itsverexpression is one of the earliest events in the oncogenicrocess in the pancreas. It is detected in 82% of PanIN-1Aesions and in 100% of PanIN-3 lesions [22].

The oncogene KRAS is activated in approximately 90%f all pancreatic adenocarcinomas. These mutations affectodons 12 (the most common mutation), 13 and 61 [23].utated protein KRAS facilitates progression along the cellycle via activation of MAP kinases and AKT kinase [24].hese KRAS gene mutations also appear very early in pan-reatic cancer. They are found in 36% of PanIN-1A lesions,4% of PanIN-1B and PanIN-2 lesions, and 87% of PanIN-3esions [25]. It is important to note that PanIN lesions car-ying different mutations can coexist in the same organ;hese mutations thus come from different clones [26]. Nev-rtheless, only one of the clones progresses to an invasiveorm, the clone with the mutations that are always found inancreatic adenocarcinomas.

umor suppressor genescertain number of tumor suppressor genes are inacti-

ated in PanIN. They are also inactivated in adenocarinomas.he CDKN2A/p16 gene encodes a protein which binds toyclin-dependent kinases Cdk4 and Cdk6, thus inhibitingheir linkage with cyclin D1 and halting the cell cycle at G1/S27]. This loss of activity, which is observed in more than0% of pancreatic adenocarcinomas, can occur via at leasthree mechanisms: (a) homozygous deletion, (b) mutation ofne allele plus deletion on the second, or (c) hypermethy-ation of its promoter resulting in nearly total suppressionf its expression [28—30]. Loss of CDKN2A/p16 expression islso observed in PanIN lesions with an incidence of 30% inanIN-1A and PanIN-1B, 55% in PanIN-2 and more than 70%n PanIN-3 [31].

TP53 tumor gene suppressor is inactivated in more than0% of pancreatic adenocarcinomas. The mechanism of inac-ivation is often associated with loss of one allele and annactivating mutation of the other [32]. Inactivation of TP53s rarely observed in PanIN-1A or PanIN-1B lesions, indicatinghat the loss of TP53 function occurs late in the tumorigen-sis process [33].

SMAD4 tumor suppressor gene is frequently inactivatedn pancreatic adenocarcinomas [34]. Protein SMAD4 is impli-ated in the signaling cascade of TGF beta; its inactivationlocks the inhibitor effect of TGF beta and other members ofts family on cell growth, thus allowing uncontrolled growthf tumor cells [35]. SMAD4 expression is preserved in PanIN-1nd PanIN-2 lesions, but lost in 40% of PanIN-3 lesions [36].

ther anomalies observedoss of telomere integrity in the ductal epithelium is prob-bly the cause of the genomic instability observed in PanIN

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O. Turrini et al.

37]. Telomeres are TTAGGG repeats that cap the ends ofhromosomes, inducing a certain degree of stability dur-ng cell division [38]. Telomere shortening is one of the firstenetic aberrations detected in PanIN lesions, occurring inore than 90% [37]. Intact telomeres act like ‘‘guardians’’

f the pancreatic ductal genome; their shortening in PanINeads to progressive accumulation of chromosomal anoma-ies and finally malignant transformation.

In addition, other proteins are overexpressed during thetepwise progression towards PanIN; these proteins couldhus be interesting markers of pancreatic carcinogenesis.xpression of protein Ki-67 is associated with cell prolifer-tion. Ki-67 is expressed more frequently in the nuclei ofigh-grade PanIN cells (22% in PanIN-3) than in early forms0.7% in PanIN-1A) [33]. The index is 35 to 40% in adeno-arcinomas [39]. Expression of topoisomerase II, necessaryor DNA relaxation before replication, follows that of pro-ein Ki-67 [33]. Cyclin D1, which is a key element of the cellycle, is overexpressed in 80% of pancreatic adenocarcino-as, 50% of PanIN-3 and 30% of PanIN-2 [40]. The expression

f these markers is not due to a genomic or epigenetic event.t is simply the consequence of accelerated cell proliferationue to the malignant transformation at an intensity directlyelated to the transformation.

ancreatic intra-epithelial neoplasia and accumulationf genetic anomaliesrogression from PanIN-1A to PanIN-3 then to pancreaticdenocarcinoma is associated with progressive accumu-ation of genomic alterations. This model of progressionas been described by several laboratories [33,41,42] andppears to be well established, with new characterizationsf new anomalies in precancerous lesions regularly pub-ished. Activation of the KRAS oncogene and amplification ofRBB2 are the earliest genetic alterations observed in PanIN-A. Telomere shortening is also observed at an early stage.oss of CDKN2A/p16 activity comes slightly later in stagesanIN-1B and PanIN-2. Cyclin D1 suppression is seen in PanIN-. The latest events, observed in PanIN-3, are inactivationf TP53 and SMAD4 and expression of Ki-67 (Fig. 1).

ntraductal papillary mucinous neoplasm

PMN is a relatively uncommon lesion. The macroscopicppearance is a dilated pancreatic duct filled with mucus;here is a strong potential for degeneration [43]. IPMN canlso be seen as a small cyst in the lumen of a secondaryancreatic duct or as a set of large multicystic lesions involv-ng the main pancreatic duct and several secondary ducts.alignant transformation is mainly seen in this second type

44]. IPMNs occur more readily in older men and less fre-uently in women [45]. Discovery is generally fortuitous;ometimes made because of acute obstructive pancreatitisesulting from mucus filled ducts [43]. Microscopically, IPMNsre composed of a mucin-producing ductal type epithe-ium. IPMNs are classed by architecture into four types:

astric, intestinal, pancreaticobiliary and oncocytic [46].uctal cells show diverse degrees of atypia ranging from

ow-grade to high-grade lesions, the latter correspondingo in situ carcinoma. In 1996, the World Health Organiza-ion defined four groups of IPMN as a function of cell atypia:

Page 4: Genetic alterations in precancerous pancreatic lesions and their clinical implications

Genetic alterations in precancerous pancreatic lesions and their clinical implications 1031

ncer

iaf

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Figure 1 Genetic anomalies associated with preca

adenomas, borderline tumors, in situ carcinomas, and inva-sive tumors. Disease prognosis depends upon the presenceof invasive carcinoma at diagnosis. Unfortunately, invasivecarcinoma progresses much like classic adenocarcinoma.

Genetic alterations in intraductal papillarymucinous neoplasm

Three oncogenes (KRAS, ERBB2 and AKT) and five tumor sup-pressor genes (CDKN2A/p16, TP53, SMAD4, LKB1 and DUSP6)are involved.

OncogenesThe oncogene KRAS is activated by mutation in approxi-mately 72% of IPMN lesions [47]. This mutation has beenidentified in low-grade and high-grade lesions. Although itis not particularly specific, this mutation appears to be nec-essary for the development of IPMN. It is interesting to notethat different mutations which activate KRAS are sometimesobserved in the same patient at different IPMN foci. Therecould be several explanations for this, including polyclonaldevelopment of IPMN and KRAS mutation secondary to thedevelopment of the lesions. Yoshizawa et al. have suggestedthat low-grade forms arise via a polyclonal mechanism whilehigh-grade forms develop from low-grade lesions in a clonalmanner [48]. Overexpression of ERBB2, which is a common

and early event in pancreatic cancer as mentioned above,has been observed in 63% of IPMN lesions [49]. AKT is a pro-tein kinase involved in KRAS signaling. It plays a key rolein cell growth and survival. A recent study showed that theactivated phosphorylated form of this kinase was induced

ensdc

ous lesions, PanIN, IPMN, and chronic pancreatitis.

n 63% of lesions (versus 70% in adenocarcinomas). It waslso interesting to note that AKT activation is slightly morerequent in high-grade than in low-grade forms [50].

umor suppressor geness mentioned above, CDKN2A/p16 plays a major role in cyclerrest. CDKN2A/p16 expression is lost in about half of allPMN lesions [51]. This diminished expression appears toesult in most cases from hypermethylation of its promoter52]. The tumor suppressor gene TP53 is a transcription fac-or which induces the expression of several genes involved inell cycle arrest, DNA repair, and apoptosis induced by DNAamage. TP53 activation is lost in half of human tumors,ither by deletion or mutation of its gene, or by proteoso-al hyperdegradation. Loss of TP53 activity is observed in

alf of IPMNs with a higher frequency in high-grade lesions46,47,49]. The current hypothesis is that loss of TP53 activ-ty in this type of lesion induces loss of genome integrityhich in turn leads to malignant transformation. The chro-osome region 18q21.1 frequently exhibits deletion of its

wo alleles in pancreatic adenocarcinomas [53—55]. Theumor suppressor SMAD4 is located in this region. Never-heless, homozygous mutations of SMAD4 are rare in IPMN.oreover, expression of the SMAD4 protein is generally pre-

erved in these lesions, independent of the degree of atypia56]. Loss of the SMAD4 function appears to be a very late

vent in IPMN transformation, unless this transformationever involves the SMAD4 pathway. The LKB1 gene is respon-ible for Peutz-Jeghers syndrome (transmitted by autosomalominant inheritance, this syndrome associates periorifi-ial lentiginosis and intestinal polyposis; the development
Page 5: Genetic alterations in precancerous pancreatic lesions and their clinical implications

1 O. Turrini et al.

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Table 1 Genetic alterations observed in precancerouslesions of the pancreas.

PanIN IPMN

Size of lesion Microscopic Macroscopic

Oncogenes ERBB2KRAS

KRASERBB2AKT

Tumor suppressor genes CDKN2A/p16TP53SMAD4

CDKN2A/p16TP53SMAD4(late)LKB1DUSP6

Others Telomere MUC2

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f several types of cancer including pancreatic cancer cane found during the course of the disease). The role of LKB1nvolves preservation of cell polarity [57]. Mutation of LKB1as been identified in 25% of IPMN lesions in patients withouteutz-Jeghers syndrome and also in a few cases of pancre-tic adenocarcinoma [58]. LKB1 might thus play a role in theevelopment of IPMN in some patients. Finally, the expres-ion of DUSP6, a phosphatase which interacts with the kinaseAPK1 regulating its activity, appears to be lost or greatlyiminished in a few IPMNs [52].

anagement and treatment of intraductalapillary mucinous neoplasm patients

linically, the major progress in recent years for patientsith pancreatic cancer has been the use of endoscopic ultra-

ound, CT scan or MRI to explore the pancreatic ducts anddentify lesions so that prophylactic pancreatic resectionan be performed before they degenerate into invasive car-inoma. Criteria have been defined to establish the formalndication for pancreatic resection when the short-term riskf degeneration is high: symptomatic IPMN, involvement ofhe main duct, cysts measuring more than 20 mm, the pres-nce of an intramural nodule at the level of the cyst(s) orn the main duct. However, there is no agreement about theppropriate examinations or their schedule in patients withPMN with a low risk of degeneration. Because benign IPMNan degenerate rapidly, certain teams have decided to oper-te on all patients with a diagnosis of IPMN. Surgery for IPMNs highly variable depending on the extent of the lesion(s),anging from simple enucleation to total pancreatectomyith resulting and difficult-to-control diabetes. Prophylactic

urgery results in substantial morbidity and mortality pre-enting a major challenge for decision making and patientanagement (convincing an asymptomatic patient of the

eed for a major operation with significant sequella). Obvi-usly, the risk of surgery must be balanced against the riskf degeneration and the development of pancreatic cancerith a known fatal outcome. Several teams have tried toefine factors predictive of malignity in patients with IPMN.roposed methods for distinguishing between benign andegenerated IPMN include endoscopic ultrasound-guidedne needle aspiration for cytological and immunohistochem-

cal analysis [59—61] and biochemical analysis or tumorarker assay of endoscopic ultrasound-guided fine needle

spiration products [62] or pancreatic juice [63]. To date,he precision of these methods has not been shown.

Progress in IPMN research could help determine appro-riate management practices. New predictive criteria basedn genomic or proteomic analyses of endoscopic ultrasound-uided fine needle aspirations rather than solely on imagingesults could help limit the indication for mutilating surgeryo patients with IPMNs with a clear prognosis of malignantegeneration.

ifferences between pancreaticntra-epithelial neoplasia and intraductal

apillary mucinous neoplasm

anIN and IPMN are both intraductal lesions with a potentialor malignant degeneration. They are nevertheless very dif-

nwaca

shorteningKi67Cyclin D1

erent lesions. The most important difference is their size.anINs are microscopic lesions while IPMNs are macroscopic.t the present time, PanINs cannot be detected withrdinary endoscopic or radiological methods. They are dis-overed fortuitously during histological analysis of surgicalpecimens. Immunoreactivity can also distinguish betweenhe two types of lesions. MUC2 is found in most IPMNs butever in PanINs. SMAD4 expression, often lost in PanIN-3,s almost systematically preserved in both degenerated andon-degenerated IPMNs [56]. DUSP6 expression is preservedn PanINs but strongly reduced in most IPMNs [46]. Theseifferences suggest distinct molecular mechanisms under-ying the development of these two precancerous lesionsTable 1).

hronic pancreatitis: a precancerous state

t is now generally accepted that patients with chronicancreatitis have a higher risk of developing adenocarci-oma of the pancreas [64], and more particularly if there isereditary chronic pancreatitis [65]. Chronic inflammationight cause DNA damage which accumulates over time, as

n PanIN. Several teams have described PanIN-like lesionsn chronic pancreatitis tissue [66,67]. This could be highlyignificant for the well-known problem of the differentialiagnosis between ‘‘tumor-like’’ chronic pancreatitis andancer: despite the contribution of endoscopic ultrasound,definite diagnosis of adenocarcinoma is difficult to obtain.urthermore, a negative specimen does not necessarilyxclude a diagnosis of cancer in these patients [68—70].he usefulness of searching for genetic anomalies in samplesbtained from patients with chronic pancreatitis has alreadyeen studied: the search for the KRAS oncogene mutation inissue [71], serum [72], and endoscopic ultrasound-guidedeedle aspirations [73] appears to be more useful than aearch in pancreatic juices [74,75]. Nevertheless, there is

o validated screening strategy to monitor these patientsith chronic pancreatitis. A better knowledge of the geneticlterations and of the carcinogenesis of pancreatic tissueould be helpful in establishing earlier diagnosis of pancre-tic cancer in these patients.
Page 6: Genetic alterations in precancerous pancreatic lesions and their clinical implications

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Genetic alterations in precancerous pancreatic lesions and t

Conclusions and perspectives

PanIN and IPMN are distinctly different precancerous lesions.For the time being, the diagnosis of PanIN is based on patho-logical analysis of a surgical specimen. A set of geneticmarkers of PanIN is needed to obtain a simple, specificand early diagnosis so that prophylactic measures can betaken, particularly in the context of familial pancreatitis.Although the diagnosis of IPMN is made earlier and earlier,the decision for surgery is still a major challenge. If genomicor proteomic markers of IPMN could be detected in endo-scopic ultrasound fine needle aspirations, patients at highrisk of malignant degeneration could be identified earlierso that the most appropriate preventive intervention couldbe proposed. These observations illustrate the importanceof obtaining better knowledge of the genetic alterationsin PanIN, which could be useful for familial screening. Ifthe genetic alterations of IPMN were better understood,patients could be selected for close surveillance or surgery.

It is also important to note that most genetic anoma-lies observed in PanIN and IPMN do not differ fromthose observed in established pancreatic adenocarcinoma,except for the major chromosomal alterations (duplications,deletions, rearrangements) which are only present in ade-nocarcinomas. These anomalies may result from anomalousmitosis, with poor redistribution of the genetic materialbetween the two daughter cells, which most probably occursin advanced adenocarcinoma. This suggests that geneticalterations are necessary for the malignant transformationof precancerous lesions but are not sufficient for progressionto cancer. Other factors are necessary. Characterizing thesefactors, which one would expect to be similar to the predis-posing factors for cancer (chronic inflammation, cigarettesmoking, certain diets, etc.), could result in better prophy-laxis for pancreatic cancer.

Conflits of interest

None.

References

[1] Greenlee RT, Hill-Harmon MB, Murray T, Thun M. Cancer statis-tics, 2001. CA Cancer J Clin 2001;51:15—36.

[2] Devesa SS, Blot WJ, Stone BJ, Miller BA, Tarone RE, FraumeniJr JF. Recent cancer trends in the United States. J Natl CancerInst 1995;87:175—82.

[3] Yeo CJ, Abrams RA, Grochow LB, Sohn TA, Ord SE, HrubanRH, et al. Pancreaticoduodenectomy for pancreatic adeno-carcinoma: postoperative adjuvant chemoradiation improvessurvival. A prospective, single-institution experience. Ann Surg1997;225:621—33.

[4] Warshaw AL, Gu ZY, Wittenberg J, Waltman AC. Preoperativestaging and assessment of resectability of pancreatic cancer.Arch Surg 1990;125:230—3.

[5] Lillemoe KD, Yeo CJ, Cameron JL. Pancreatic cancer: state-of-the-art care. CA Cancer J Clin 2000;50:241—68.

[6] Cubilla AL, Fitzgerald PJ. Morphological lesions associated withhuman primary invasive nonendocrine pancreas cancer. CancerRes 1976;36:2690—8.

[7] Klimstra DS, Longnecker DS. K-ras mutations in pancreatic duc-tal proliferative lesions. Am J Pathol 1994;145:1547—50.

[

clinical implications 1033

[8] Kozuka S, Sassa R, Taki T, Masamoto K, Nagasawa S, Saga S,et al. Relation of pancreatic duct hyperplasia to carcinoma.Cancer 1979;43:1418—28.

[9] Hruban RH, Adsay NV, Albores-Saavedra J, Compton C, GarrettES, Goodman SN, et al. Pancreatic intraepithelial neoplasia:a new nomenclature and classification system for pancreaticduct lesions. Am J Surg Pathol 2001;25:579—86.

10] Takaori K, Hruban RH, Maitra A, Tanigawa N. Current topics onprecursors to pancreatic cancer. Adv Med Sci 2006;51:23—30.

11] Grippo PJ, Nowlin PS, Demeure MJ, Longnecker DS, Sand-gren EP. Preinvasive pancreatic neoplasia of ductal phenotypeinduced by acinar cell targeting of mutant Kras in transgenicmice. Cancer Res 2003;63:2016—9.

12] Guerra C, Mijimolle N, Dhawahir A, Dubus P, Barradas M,Serrano M, et al. Tumor induction by an endogenous K-rasoncogene is highly dependent on cellular context. Cancer Cell2003;4:111—20.

13] Hingorani SR, Petricoin EF, Maitra A, Rajapakse V, King C,Jacobetz MA, et al. Preinvasive and invasive ductal pancre-atic cancer and its early detection in the mouse. Cancer Cell2003;4:437—50.

14] Aguirre AJ, Bardeesy N, Sinha M, Lopez L, Tuveson DA, HornerJ, et al. Activated Kras and Ink4a/Arf deficiency cooperate toproduce metastatic pancreatic ductal adenocarcinoma. GenesDev 2003;17:3112—26.

15] Bardeesy N, Aguirre AJ, Chu GC, Cheng KH, Lopez LV, Hezel AF,et al. Both p16(Ink4a) and the p19(Arf)-p53 pathway constrainprogression of pancreatic adenocarcinoma in the mouse. ProcNatl Acad Sci U S A 2006;103:5947—52.

16] Hingorani SR, Wang L, Multani AS, Combs C, DeramaudtTB, Hruban RH, et al. Trp53R172H and KrasG12D cooperateto promote chromosomal instability and widely metastaticpancreatic ductal adenocarcinoma in mice. Cancer Cell2005;7:469—83.

17] Ijichi H, Chytil A, Gorska AE, Aakre ME, Fujitani Y, Fujitani S,et al. Aggressive pancreatic ductal adenocarcinoma in micecaused by pancreas-specific blockade of transforming growthfactor-beta signaling in cooperation with active Kras expres-sion. Genes Dev 2006;20:3147—60.

18] Izeradjene K, Combs C, Best M, Gopinathan A, Wagner A, GradyWM, et al. Kras(G12D) and Smad4/Dpc4 haploinsufficiencycooperate to induce mucinous cystic neoplasms and invasiveadenocarcinoma of the pancreas. Cancer Cell 2007;11:229—43.

19] Kojima K, Vickers SM, Adsay NV, Jhala NC, Kim HG, Schoeb TR,et al. Inactivation of Smad4 accelerates Kras(G12D)-mediatedpancreatic neoplasia. Cancer Res 2007;67:8121—30.

20] Sharpless NE, Ramsey MR, Balasubramanian P, CastrillonDH, DePinho RA. The differential impact of p16(INK4a) orp19(ARF) deficiency on cell growth and tumorigenesis. Onco-gene 2004;23:379—85.

21] Guerra C, Schuhmacher AJ, Canamero M, Grippo PJ, Verda-guer L, Perez-Gallego L, et al. Chronic pancreatitis is essentialfor induction of pancreatic ductal adenocarcinoma by K-Rasoncogenes in adult mice. Cancer Cell 2007;11:291—302.

22] Day JD, Digiuseppe JA, Yeo C, Lai-Goldman M, Anderson SM,Goodman SN, et al. Immunohistochemical evaluation of HER-2/neu expression in pancreatic adenocarcinoma and pancre-atic intraepithelial neoplasms. Hum Pathol 1996;27:119—24.

23] Caldas C, Kern SE. K-ras mutation and pancreatic adenocarci-noma. Int J Pancreatol 1995;18:1—6.

24] Hingorani SR, Tuveson DA. Ras redux: rethinking how and whereRas acts. Curr Opin Genet Dev 2003;13:6—13.

25] Lohr M, Kloppel G, Maisonneuve P, Lowenfels AB, Luttges J. Fre-

quency of K-ras mutations in pancreatic intraductal neoplasiasassociated with pancreatic ductal adenocarcinoma and chronicpancreatitis: a meta-analysis. Neoplasia 2005;7:17—23.

26] Laghi L, Orbetegli O, Bianchi P, Zerbi A, Di CV, Boland CR,et al. Common occurrence of multiple K-RAS mutations in

Page 7: Genetic alterations in precancerous pancreatic lesions and their clinical implications

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[

[

[

[

[

[

[

[

[

[

[

[

[

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[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

[

034

pancreatic cancers with associated precursor lesions and inbiliary cancers. Oncogene 2002;21:4301—6.

27] Sherr CJ. Cell cycle control and cancer. Harvey Lect2000;96:73—92.

28] Caldas C, Hahn SA, da Costa LT, Redston MS, Schutte M, SeymourAB, et al. Frequent somatic mutations and homozygous dele-tions of the p16 (MTS1) gene in pancreatic adenocarcinoma.Nat Genet 1994;8:27—32.

29] Schutte M, Hruban RH, Geradts J, Maynard R, Hilgers W,Rabindran SK, et al. Abrogation of the Rb/p16 tumor-suppressive pathway in virtually all pancreatic carcinomas.Cancer Res 1997;57:3126—30.

30] Ueki T, Toyota M, Sohn T, Yeo CJ, Issa JP, Hruban RH, et al.Hypermethylation of multiple genes in pancreatic adenocarci-noma. Cancer Res 2000;60:1835—9.

31] Wilentz RE, Geradts J, Maynard R, Offerhaus GJ, Kang M, Gog-gins M, et al. Inactivation of the p16 (INK4A) tumor-suppressorgene in pancreatic duct lesions: loss of intranuclear expression.Cancer Res 1998;58:4740—4.

32] Redston MS, Caldas C, Seymour AB, Hruban RH, da Costa L,Yeo CJ, et al. p53 mutations in pancreatic carcinoma and evi-dence of common involvement of homocopolymer tracts in DNAmicrodeletions. Cancer Res 1994;54:3025—33.

33] Maitra A, Adsay NV, Argani P, Iacobuzio-Donahue C, DeMarzo A, Cameron JL, et al. Multicomponent analysis of thepancreatic adenocarcinoma progression model using a pan-creatic intraepithelial neoplasia tissue microarray. Mod Pathol2003;16:902—12.

34] Hahn SA, Schutte M, Hoque AT, Moskaluk CA, da Costa LT, Rozen-blum E, et al. DPC4, a candidate tumor suppressor gene athuman chromosome 18q21.1. Science 1996;271:350—3.

35] Truty MJ, Urrutia R. Basics of TGF-beta and pancreatic cancer.Pancreatology 2007;7:423—35.

36] Wilentz RE, Iacobuzio-Donahue CA, Argani P, McCarthy DM,Parsons JL, Yeo CJ, et al. Loss of expression of Dpc4 inpancreatic intraepithelial neoplasia: evidence that DPC4 inac-tivation occurs late in neoplastic progression. Cancer Res2000;60:2002—6.

37] van Heek NT, Meeker AK, Kern SE, Yeo CJ, Lillemoe KD,Cameron JL, et al. Telomere shortening is nearly univer-sal in pancreatic intraepithelial neoplasia. Am J Pathol2002;161:1541—7.

38] Gisselsson D. Chromosome instability in cancer: how, when,and why? Adv Cancer Res 2003;87:1—29.

39] Klein WM, Hruban RH, Klein-Szanto AJ, Wilentz RE. Directcorrelation between proliferative activity and dysplasia in pan-creatic intraepithelial neoplasia (PanIN): additional evidencefor a recently proposed model of progression. Mod Pathol2002;15:441—7.

40] Gansauge S, Gansauge F, Ramadani M, Stobbe H, Rau B,Harada N, et al. Overexpression of cyclin D1 in human pan-creatic carcinoma is associated with poor prognosis. CancerRes 1997;57:1634—7.

41] Hruban RH, Goggins M, Parsons J, Kern SE. Progression modelfor pancreatic cancer. Clin Cancer Res 2000;6:2969—72.

42] Bardeesy N, De Pinho RA. Pancreatic cancer biology and genet-ics. Nat Rev Cancer 2002;2:897—909.

43] Tanaka M. Intraductal papillary mucinous neoplasm of the pan-creas: diagnosis and treatment. Pancreas 2004;28:282—8.

44] Kobari M, Egawa S, Shibuya K, Shimamura H, Sunamura M,Takeda K, et al. Intraductal papillary mucinous tumors ofthe pancreas comprise 2 clinical subtypes: differences inclinical characteristics and surgical management. Arch Surg

1999;134:1131—6.

45] Kimura W, Sasahira N, Yoshikawa T, Muto T, Makuuchi M.Duct-ectatic type of mucin producing tumor of the pancreas–new concept of pancreatic neoplasia. Hepatogastroenterology1996;43:692—709.

[

O. Turrini et al.

46] Furukawa T, Kloppel G, Volkan AN, Albores-Saavedra J,Fukushima N, Horii A, et al. Classification of types of intraduc-tal papillary-mucinous neoplasm of the pancreas: a consensusstudy. Virchows Arch 2005;447:794—9.

47] Satoh K, Shimosegawa T, Moriizumi S, Koizumi M, Toyota T.K-ras mutation and p53 protein accumulation in intraductalmucin-hypersecreting neoplasms of the pancreas. Pancreas1996;12:362—8.

48] Yoshizawa K, Nagai H, Sakurai S, Hironaka M, Morinaga S,Saitoh K, et al. Clonality and K-ras mutation analyses ofepithelia in intraductal papillary mucinous tumor and muci-nous cystic tumor of the pancreas. Virchows Arch 2002;441:437—43.

49] Sessa F, Solcia E, Capella C, Bonato M, Scarpa A, Zamboni G, etal. Intraductal papillary-mucinous tumours represent a distinctgroup of pancreatic neoplasms: an investigation of tumour celldifferentiation and K-ras, p53 and c-erbB-2 abnormalities in26 patients. Virchows Arch 1994;425:357—67.

50] Semba S, Moriya T, Kimura W, Yamakawa M. PhosphorylatedAkt/PKB controls cell growth and apoptosis in intraductalpapillary-mucinous tumor and invasive ductal adenocarcinomaof the pancreas. Pancreas 2003;26:250—7.

51] Furukawa T, Fujisaki R, Yoshida Y, Kanai N, Sunamura M, AbeT, et al. Distinct progression pathways involving the dysfunc-tion of DUSP6/MKP-3 in pancreatic intraepithelial neoplasiaand intraductal papillary-mucinous neoplasms of the pancreas.Mod Pathol 2005;18:1034—42.

52] House MG, Guo M, Iacobuzio-Donahue C, Herman JG. Molecularprogression of promoter methylation in intraductal papillarymucinous neoplasms (IPMN) of the pancreas. Carcinogenesis2003;24:193—8.

53] Hahn SA, Seymour AB, Hoque AT, Schutte M, da Costa LT, Red-ston MS, et al. Allelotype of pancreatic adenocarcinoma usingxenograft enrichment. Cancer Res 1995;55:4670—5.

54] Inoue H, Furukawa T, Sunamura M, Takeda K, Matsuno S, HoriiA. Exclusion of SMAD4 mutation as an early genetic change inhuman pancreatic ductal tumorigenesis. Genes ChromosomesCancer 2001;31:295—9.

55] Kimura M, Abe T, Sunamura M, Matsuno S, Horii A. Detaileddeletion mapping on chromosome arm 12q in human pancre-atic adenocarcinoma: identification of a I-cM region of commonallelic loss. Genes Chromosomes Cancer 1996;17:88—93.

56] Iacobuzio-Donahue CA, Klimstra DS, Adsay NV, Wilentz RE,Argani P, Sohn TA, et al. Dpc-4 protein is expressed in virtuallyall human intraductal papillary mucinous neoplasms of the pan-creas: comparison with conventional ductal adenocarcinomas.Am J Pathol 2000;157:755—61.

57] Forcet C, Étienne-Manneville S, Gaude H, Fournier L, Debilly S,Salmi M, et al. Functional analysis of Peutz-Jeghers mutationsreveals that the LKB1 C-terminal region exerts a crucial role inregulating both the AMPK pathway and the cell polarity. HumMol Genet 2005;14:1283—92.

58] Sato N, Rosty C, Jansen M, Fukushima N, Ueki T, Yeo CJ, etal. STK11/LKB1 Peutz-Jeghers gene inactivation in intraduc-tal papillary-mucinous neoplasms of the pancreas. Am J Pathol2001;159:2017—22.

59] Michaels PJ, Brachtel EF, Bounds BC, Brugge WR, PitmanMB. Intraductal papillary mucinous neoplasm of the pan-creas: cytologic features predict histologic grade. Cancer2006;108:163—73.

60] Layfield LJ, Cramer H. Fine-needle aspiration cytology of intra-ductal papillary-mucinous tumors: a retrospective analysis.Diagn Cytopathol 2005;32:16—20.

61] Salla C, Chatzipantelis P, Konstantinou P, Karoumpalis I, Sakel-lariou S, Pantazopoulou A, et al. Endoscopic ultrasound-guidedfine-needle aspiration cytology in the diagnosis of intraduc-tal papillary mucinous neoplasms of the pancreas. A study of8 cases. JOP 2007;8:715—24.

Page 8: Genetic alterations in precancerous pancreatic lesions and their clinical implications

heir

[

[

[

[

[

Genetic alterations in precancerous pancreatic lesions and t

[62] Maire F, Voitot H, Aubert A, Palazzo L, O’Toole D, Couvelard A,et al. Intraductal papillary mucinous neoplasms of the pan-creas: performance of pancreatic fluid analysis for positivediagnosis and the prediction of malignancy. Am J Gastroenterol2008;103:2871—7.

[63] Kondo H, Sugano K, Fukayama N, Hosokawa K, Ohkura H, OhtsuA, et al. Detection of K-ras gene mutations at codon 12 in thepancreatic juice of patients with intraductal papillary muci-nous tumors of the pancreas. Cancer 1997;79:900—5.

[64] Lowenfels AB, Maisonneuve P, Cavallini G, Ammann RW,Lankisch PG, Andersen JR, et al. Pancreatitis and the risk ofpancreatic cancer. N Engl J Med 1993;328:1433—7.

[65] Rebours V, Boutron-Ruault MC, Schnee M, Ferec C, Maire F,Hammel P, et al. Risk of pancreatic adenocarcinoma in patientswith hereditary pancreatitis: a national exhaustive series. AmJ Gastroenterol 2008;103:111—9.

[66] Rosty C, Geradts J, Sato N, Wilentz RE, Roberts H, Sohn T,et al. p16 inactivation in pancreatic intraepithelial neoplasias(PanINs) arising in patients with chronic pancreatitis. Am J SurgPathol 2003;27:1495—501.

[67] Cylwik B, Nowak HF, Puchalski Z, Barczyk J. Epithelial anoma-lies in chronic pancreatitis as a risk factor of pancreatic cancer.Hepatogastroenterology 1998;45:528—32.

[68] Ardengh JC, Lopes CV, Campos AD, Pereira de Lima LF, Venco F,

Modena JL. Endoscopic ultrasound and fine needle aspirationin chronic pancreatitis: differential diagnosis between pseudo-tumoral masses and pancreatic cancer. JOP 2007;8:413—21.

[69] Iordache S, Saftoiu A, Cazacu S, Gheonea DI, Dumitrescu D,Popescu C, et al. Endoscopic ultrasound approach of pancreatic

[

clinical implications 1035

cancer in chronic pancreatitis patients in a tertiary referralcentre. J Gastrointestin Liver Dis 2008;17:279—84.

70] Varadarajulu S, Tamhane A, Eloubeidi MA. Yield of EUS-guided FNA of pancreatic masses in the presence or theabsence of chronic pancreatitis. Gastrointest Endosc 2005;62:728—36.

71] Talar-Wojnarowska R, Gasiorowska A, Smolarz B, Romanowicz-Makowska H, Strzelczyk J, Janiak A, et al. Clinical significanceof K-ras and c-erbB-2 mutations in pancreatic adenocarci-noma and chronic pancreatitis. Int J Gastrointest Cancer2005;35:33—41.

72] Maire F, Micard S, Hammel P, Voitot H, Levy P, Cugnenc PH,et al. Differential diagnosis between chronic pancreatitis andpancreatic cancer: value of the detection of KRAS2 mutationsin circulating DNA. Br J Cancer 2002;87:551—4.

73] Urgell E, Puig P, Boadas J, Capella G, Queralto JM, BoludaR, et al. Prospective evaluation of the contribution of K-rasmutational analysis and CA 19.9 measurement to cytologicaldiagnosis in patients with clinical suspicion of pancreatic can-cer. Eur J Cancer 2000;36:2069—75.

74] Queneau PE, Adessi GL, Thibault P, Cleau D, Heyd B, Man-tion G, et al. Early detection of pancreatic cancer in patientswith chronic pancreatitis: diagnostic utility of a K-ras pointmutation in the pancreatic juice. Am J Gastroenterol 2001;96:

700—4.

75] Uehara H, Nakaizumi A, Tatsuta M, Baba M, Takenaka A, Uedo N,et al. Diagnosis of pancreatic cancer by detecting telomeraseactivity in pancreatic juice: comparison with K-ras mutations.Am J Gastroenterol 1999;94:2513—8.