Pathobiology 2011;78:301310 DOI: 10.

1159/000321703

Molecular Pathology of Gastric Carcinoma

Bo-Gun Jang Woo Ho Kim
Department of Pathology and Cancer Research Institute, Seoul National University College of Medicine, Seoul, Korea

Key Words Stomach neoplasms Mutation Oncogenes Tumor suppressor gene Microsatellite instability DNA methylation

Click to edit Master subtitlebeen undertaken to identify novel methylationstyle search has
silenced genes in GC, which will help us understand the overall molecular features of GC and further provide novel opportunities in the treatment of GC.
Copyright 2011 S. Karger AG,

Basel

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Gastric carcinoma (GC) is a biologically heterogeneous Abstract diseaseinvolvingnumerousgeneticandepigeneticalterations. A very small proportion of GCs can be caused by a specific germ-linemutationoftheE-cadheringene(CDH1).Sporadic GC is developed through multistep processes that begin with Helicobacter pylori-induced atrophic gastritis. EpsteinBarr virus is another infectious cause of GC, and the above two infection-associated GCs are characterized by global CpG island methylation in the promoter region of cancerrelated genes. Mutations of tumor protein p53 (TP53) and

Introduction

-catenin(CTNNB1)genesoccurearlyinthedevelopmentof GC and contribute to gastric carcinogenesis. Furthermore, significant numbers of GCs show loss of Runx3 due to hemizygous deletion and hypermethylation of the promoter region. Aberrant Cdx2 expression has been shown in precancerous lesions as well as GC. However, it remains unclear whetherCdx2playsanoncogenicroleingastriccarcinogenesis. GC with microsatellite instability is also a welldefined E-Mail karger@karger.ch subset exhibiting distinctive clinicopathologic features. www.karger.com TarAccessible online at: geted therapy against GC with ERBB2 amplification www.karger.co recently m/pat improvedtheprognosisofpatientswithadvancedGC.Inaddition, epigenetic changes in GC could be attractive targets

Gastric carcinoma (GC) is one of the most common malignanttumorsandthesecondmostcommoncauseof cancerdeathworldwidealthoughtherehasbeenasteady decline in the incidence and mortality risk of GC over severaldecadesinmostcountries[1].Therearelargegeographical variations, and the incidence and death rate of GC are more than twice as high in Asian/Pacific Islanders as in Whites [2]. Despite the tremendous improvementsindiagnosisandtreatmenttechnologies,theprognosis of advanced GC remains poor and the survival of affected patients is less than 40% even after a curative surgical resection and adjuvant therapy [3]. GC is considered as a multifactorial disease since many inherited and environmental factors play a role in gastric carcinogenesis, including the hosts genetic background, infectious agents and dietary habits. Chronic atrophic gastritis has been known to be the most imporSeoul 110-799 (Korea) Tel. +82 27 40 8269, E-Mail risk factor for GC. Helicobacter tant and the greatest woohokim @ snu.ac.kr pylori infection, classified by the WHO as a class I carcino-

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Stomach or gastric cancer incidence is secondly only to lung cancer worldwide, with an estimated 870,000 new cases and 650,00 deaths every year. The high-risk areas are East Asia, South America, and Eastern Europe. Data show that second-generation migrants from the high- to low-risk areas have lower incidence rates than their parents. Incidence rates are twice as high in males as in females. Thirty-eight percent of the worlds stomach cancer cases occur in China, and it is the most frequent cancer in males in other parts of East Asia. Agestandardized incidence rates (ASRs) are the highest in the world in Japan (77.9 per 100,000 in men; 33.3 in women). The major risk factors for stomach cancer are hipothesized to be nutritional, including inadequate intake of fresh fruits and vegetables, and high intake of salt, smoked fsh, and meat. Refrigeration of food is considered protective. Vitamin C, cotained in vegetables and fruits, may be protective for stomach cancer. Other possibly protective nutritional factors are whole-grain cereals, carotenoids, allium compounds (e.g., garlic), and green tea. Smoking carries a slightly increased risk for stomach cancer and alcohol does not affect risk other than in the gastric cardia. Smoking is quite common in countries of the Middle East Cancer Consortium (MECC). Another important risk factor is Helicobacter pylori infection. Other gastric diseases, such as ulcer and atrophic gastritis, elevate gastric pH, thus causing anaerobic bacterial colonization in the stomach. H. pylori prevalence ranges from 25% in developed countries to 80%-90% in the developing countries. It is now considered as the principal cause of chronic gastritis and peptic ulcer disease, and is a key risk factor for the development of gastric cancer. The prevalence of H. pylori is though to be hig in MECC countries. In Israel, the prevalence has been reported to be 72% in a rural population and 65% in an urban population, which is higher than in the United States and Western Europe, but lower than in developing countries. Another study has reported 33% positive for H. pylori in an elderly population in Israel, and a case-control study is Israel found 63% positivity among the healty controls. In a population a Jordanian endoscopy patients, 82% prevalence was reported, but there is no information for the general population. Stomach cancer incidence in the MECC countries Molecular Pathology of Gastric the period of 2002-2010 was low, compared during Carcinoma with the world ASR of 10.3 in females and 22.0 in males.

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The incidence was highest in Israeli Jews, followed by Cypriots (4.9), Jordanians (4.8), and Israeli Arabs (4.6). Although a study from Jordan reports that the characterisitics of gastric cancers diagnosed there resemble those in high-risk countries, it nevertheless appears to be a low-incidence country. Egypt had the lowest incidence in the region, with 2.9. US SEER incidence was 5.3 for the same years. These rates are 5 to 15 times lower than in Japan, where the overall rate is more than 50. The Middle East has better access to fruits and vegetables throughout the year than in many places, and the nutritional practices have Mediterranean influences. This may result in the low incidence rates that were observed. On the other hand, H. pylori seroprevalence may be high in these countries, but his was not reflected in the observed cancer incidence. Worldwide, the stomach cancer ASR in males is twice than in females. Cyprus did not show this pattern; the maleto-female ratio was closer to 1 (1.44). Egypts ratio was closer to the world pattern (1.64), while a ratio close to 2 was observed in Israeli Jews and Arabs, Jordanians, and in US SEER. A low male incidence rate was especially noticeable in Egypt and may be partially due to undiagnosed cases of stomach cancer, especially among the elderly. The Arab countries in the region had low rates, while Israel and Turkey had higher rates. In addition, all of the male-to-female rate ratios were lower than 2 for Cyprus and the Arab countries, while they were approximately 2 for Israel and Turkey. This may be partly due to ethnic differences and the more Western lifestyle in Israel and Turkey.
Age

The incidence of stomach cancer rises from age 50 years and is highest in the 70-and-older group. The highest incidence was in Israeli Jews and the lowest in Egyptians. The high incidence could be related to the deprived environmental and nutritional conditions they suffered during the Second World War. This is supported by data showing that Israeli Jews born in Europe have a higher stomach cancer incidence than Israeli Jews born elsewhere. The low rates in Egypt in this oldest age group, among both males and females, strongly suggests that older cases havent been diagnosed, perhaps due to elderly patients underuse of health care services.

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GC is a heterogeneous disorder that can be divided into at least two main histological types based on the Lauren classification: intestinal and diffuse type. These two types have distinct morphological, clinical and epidemiological features. Intestinal-type GC is more frequently observed in older patients and follows multifocal atrophic gastritis, which is accompanied by intestinal metaplasia or dysplasia. On the other hand, diffuse GC occurs more commonly in young patients and its links with atrophic gastritis or intestinal metaplasia are poor, or do not exist. These differences might represent adifferentmolecularmechanismoftumordevelopment and progression. For example, microsatellite instability and promoter hypermethylation are more commonly found in intestinal-type than in diffuse-type GC while specific genes, such as CDH1, are more often hypermethylated in diffuse-type GC [5]. GC can be also classified into four phenotypes according to mucin (MUC1, MUC2 and CD10) expression: G-type (gastric or foveolar phenotype), I-type (intestinal phenotype), GI-type (intestinal and gastric mixed phenotype) and Ntype (neithergastricnorintestinalphenotype)GC.Somedistinct genetic changes in GC have been revealed to be associated with the mucin phenotypic expression. For instance, TP53 mutations are more common in Itype than in G-type GC, whereas microsatellite instability is morefrequentinG-typethaninI-typeGC.Inaddition, somespecificepigeneticalterationsarealsoknowntobe involved in distinct mucin expression of GC. Indeed, methylation of hMLH1 occurred more frequently in MUC2-negative GC, whereas MGMT was more frequently methylated in MUC2-positive GC than in MUC2-negative GC [6]. Molecular Pathology of
Gastric Withtheadvancesinthemoleculargeneticsofcancer, Carcinoma

E-Cadherin Gene (CDH1)

E-cadherin is a cell adhesion protein that is expressed at the adherence junctions of epithelial tissue and required for development, cell differentiation and maintenance of epithelial architecture. Downregulation of Ecadherin is commonly observed in many sporadic tumors, and its loss during tumor progression has led to a concept of E-cadherin acting as a suppressor of tumor invasionandmetastasis[7].Ontheotherhand,germ-line mutationsintheCDH1generesultinacancersyndrome, a hereditary diffuse gastric cancer (HDGC), which comprisesapproximately13%ofallGCs[8].Untilnow,more than 100 different germ-line mutations of CDH1 have been described in HDGC families [9]. Carriers of CDH1 germ-linemutationshavea70%lifetimeriskofadvanced diffusegastriccancer[9]andtendtopresentwithseveral hundred microscopic foci of stage T1a intramucosal signet ring cell carcinoma, which are termed early HDGC (eHDGC) [10]. The tumor cells in multifocal eHDGC show very low E-cadherin expression, implying that the wild-type CDH1 allele is also suppressed or lost in tumor cells. Studies have revealed that the second hit is caused by promoter hypermethylation of CDH1 in at least 50% of cases [11]. The methylation patterns observed in eHDGCs are monoallelic and specific for each focus, indicatingthateacheHDGChasanindependentmonoclonal origin and that the epigenetic silencing of CDH1 by promotermethylationisanearlyeventintumordevelopment [12]. Remarkably, promoter methylation has also been identified as a major mechanism underlying Ecadherin downregulation in sporadic diffuse gastric cancers [13]. Of interest, eHDGCs have unique features, i.e. they Pathobiology 3 are hypoproliferative and lack Wnt pathway 2011;78:302310 0 activation 3 [11]. Lineage-labeling experiments with gastric differen-

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it has been revealed that carcinogenesis is a multistep process involving alterations of multiple genes, such as

that begins with H. pylori-induced superficial gastritis, progressing towards chronic atrophic gastritis, intestinal metaplasia, dysplasia and finally GC [14]. The causal relationship between H. pylori and chronic gastritis has been confirmed by the results of interventional studies showingthatbacterialeradicationleadstotheregression of precursor lesions [15]. It is well known that persistent inflammation induces increased tissue turnover, which predisposes to an excessive rate of proliferation, and in manycasesresultsinmorefrequentmitoticerrorsandan increasedrateofgeneticmutation[8].Inaddition,chronic inflammation causes genetic instability through the generation of reactive oxygen species and reactive nitrogen species which can directly damage the genomic and mitochondrial DNA [16].

terleukin 1 receptor antagonist (IL1RN), IL10 and TNF genes was also identified to be associated with increased risk for GC, which makes it possible to define a specific genetic profile associated with the highest risk for GC [23]. Furthermore, when combined with bacterial genotyping, the hosts genetic profile would provide a clinical tool to identify patients at high risk for GC [24].

Epstein-Barr Virus

Epstein-Barr virus (EBV) is a ubiquitous human her-

Click to edit Master subtitle style pesvirus that has well-established associations with a
variety of malignant neoplasms, such as Burkitts lymphoma, extranodal NK/T cell lymphoma, nasopharyngeal carcinomaandGC.EBVwasthereforeclassedasagroup IcarcinogenbytheIARCandWHO[25].EBV-associated GC is the result of monoclonal proliferation of EBVinfected epithelial cells and accounts for about 5% of GC [26]. EBV stably maintains its latent infection in carcinoma cells and expresses viral latent genes which belong to the latency I program: EBV-determined nuclear antigen 1 (EBNA1), EBV-encoded small RNA (EBER), latent membrane protein 2A (LMP2A) and BamH1-A rightward transcripts (BARTs) [27]. EBV-associated GC is a specific clinicopathologic subset with characteristics of younger age, male predominance, proximal location, lower rate of lymph node involvement, marked lymphocytic infiltration, and lace pattern within the mucosa [26]. EBV-positive GC also showed a distinct protein expression profile featured by frequent loss of p16 Jang/ (CDKN2A), smad4, Kim and CD82 (KAI-1) Fhit, compared to EBV-negative carcinomas, but they retained the expres-

It is now clear that bacterial virulence factors are importantdeterminantsthathavebeenimplicatedinthedevelopmentofGC.ThemoststudiedvirulencefactorofH. pylori is cagA protein. Infection with a cagA-positive H. pylori strain in comparison with a cagA-negative strain increasestheriskfordevelopmentofGC[17].Oncetranslocated into host cells, cagA induces a growth factorlike response in gastric epithelial cells by forming a physical complexwiththeSrchomology2domain(SH2)-containing tyrosine phosphatase (SHP-2) in a phosphorylationdependentmanner[18].Notably,ithasalsobeenreported thatcagApositiveH.pyloriinfectionofthegastricepithelium triggered aberrant expression of activationinduced cytidine deaminase (AICDA), a gene originally linked to immunoglobulin class switching and B lymphocyte hypermutation, which resulted in the accumulation of nucleotide alterations in the TP53 tumor suppressor genes Pathobiology 3 2011;78:302310 [19]. Therefore, it was hypothesized that aberrant 0 4 AICDA expressioncausedbyH.pyloriinfectionmightbeamechanism of mutation accumulation in the gastric mucosa

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line through activation of the Ras/PI3K/Akt pathway [33]. These findings suggest that LMP2A is an oncogenic fectedepithelialcells,whichmayeventuallyleadtodevelmolecule that provides a selective advantage for opment of EBV-associated GC. EBV-in(CTNNB1) Tumor Protein p53 (TP53)

in adenomas with high-grade dysplasia, suggesting that the presence of missense mutations of TP53 in adenomas mightbeakeyindicatorofmalignanttransformation[41].
-Catenin Gene

Tumor protein p53 gene (TP53) is the most commonly mutatedgeneinhumantumors;itactsasatumorsuppressor gene that induces cell cycle arrest and apoptosis. Approximately50%ofallcancersinvolvemissensemutatio ns of one p53 allele coupled with a deletion of the second allele,whichleadtocompletelossofp53function[34].Inon e report, both loss of heterozygosity and mutations of TP53 have been demonstrated in greater than 60% of GC [35]. The incidence of TP53 mutations in invasive GC ranges from 0 to 76.9%, and the mutational spectrum of TP53 in GC is very wide [36]. There are several sites where mutation occurs more commonly than in others; they include in metastatic than in primary GC common codons 175, 248, 273, 282, 245, and 213. Of interest [36]. It is theappears that TP53 alterations occur early in the fact that they are all CpG sites. Transition of development G:C]A:T atof GC because they are frequently found in CpG sites is the most common type of mutation precancerous lesions. For example, point irrespecmutations of tiveofthehistologictypeofGC[36].MorethanonemutaTP53 have been demonstrated in 52% of H. pyloritioncanbepresentinasingletumor[37],andtherecanbe associheterogeneity of the p53 mutational status within a ated gastritis even though they were located in nongiven hot tumor[38].YoungpatientshavealowerincidenceofTP53 spotcodons[39].Indeed,ithasrecentlybeenrevealedthat mutationsthanolderones,andadvancedGCtendstohave aberrantAICDAexpressioninducedbyH.pyloriinfectio ahigherincidenceofmutations.Furthermore,TP53muta in gastric carcinogenesis [36]. TP53 mutations were n also in 37.5% of intestinal metaplasia and 58% of found cancauseTP53mutations.HelicobacterinfectioninTP5 tionsoccurmuchmorecommonlyintumorsarisinginthe the than in tumors in the antrum, and they are 3 cardia dysplasticmice resulted Interestingly, while of knockout lesions [40]. in the development silent more mutadysplastic tionstendtobeobservedinadenomaswithmildormodlesions, whereas these infections in normal mice eratedegreesofdysplasia,missensemutationswerefoun failed to d Molecularany pre-neoplastic changes, which showed produce Pathology of Gastric that Carcinoma H.pyloriinfectionandp53mayactinasynergisticfashion

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The Wnt signaling pathway, initially considered to play a crucial role in embryonic development, is also known to play an important role in cancer development, and -catenin, functioning in cadherin-based epithelial cell adhesion, is a key regulator of Wnt signaling. Elevationofcytoplasmic -cateninlevelcanoccurbythebinding with Wnt ligands or by mutations in APC, AXIN1 or CTNNB1. Interestingly, mutation of CTNNB1, which encodes -catenin, seems to be exclusive to the mutations thatinactivateApcprotein.Oncogenicmutationsinvolving the amino-terminal region of -catenin make it refractory to regulation by Apc [42]. Accumulation of catenin in the cytoplasm results in its binding to members of the Tcf/Lef family of transcription factors and its tumors of -catenin nuclear staining nuclear translocation,but did not identify any CTNNB1 mutations, where the Tcf/ -catenin with contained complex difference activates diffuse- and intestinal-typeand cyclin D1 between target genes such as MYC GC. Therefore, gene fur(CCND1) [42]. Indeed, the vast majority of colorectal therstudiesarenecessarytorevealtheexactcontribution tu-Wnt pathway activation in gastric carcinogenesis. of mors contain APC mutations, but the overall Sefrequency lective targeting of gastric cancer cells with the of CTNNB1 mutations is lower [42]. On the other activated hand, a -catenin pathway showed a synergistic effect with significant percentage of gastric tumors has either cheCTNNB1 orwhichmutations. The in practically all motherapy, APC could be used incidence of Runx3 mutations binds DNA with the core-binding transfactor subunit cells withand active -catenin/Tcf [45]. (CBFB), andiffuse-type GC remains unclear. activates or represses the in intestinal- versus formed principal Parketal.[43]reportednomutationsindiffuse-typeGC, regulators that 27% ofsurvival and differentiation a but found of growth, intestinal-type tumors carried pathmutation. Whereas, Clements et al. [44] found that Runt-Related Transcription Factor 3 (RUNX3) ways.RUNX3isnowacceptedasatumorsuppressorgene 26% in a wide range of invasive and preinvasive epithelial Pathobiology and 3
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mesenchymal tumors [46]. Its suppressive activity was first reported in gastric epithelial cells of RUNX3 knockout mice in 2002 [47]. Gastric mucosa of RUNX3null miceexhibitsenhancedproliferation,suppressedapoptosis and reduced sensitivity to transforming growth factor- (TGF- ). About 4560% of human GCs showed lossofRunx3expressionduetohemizygousdeletionand hypermethylationofthepromoterregion.H.pyloriinfecthe signaling pathway. In response to tion as well as precursor lesions such as intestinal Runx3 TGF- inhibits gastric epithelial proliferation by TGF- , metainducplasia and gastric adenoma also showed RUNX3 ing the CDKN1A (p21) gene, which indicates that the hypertumethylation, indicating a role for RUNX3 in gastric mor suppressor activity of Runx3 is at least partially carascinogenesis [47, 48]. RUNX3 is a downstream sociated with its ability to induce p21 expression effector of [49]. Furthermore, Runx3 also upregulates the expression of proapoptotic gene BCL2L11 (Bim) in gastric cancer cells treated with TGF- . Bcl2l11 was downregulated in the gastric epithelium of RUNX3 knockout mice, and the BCL2L11 promoter contained conserved Runx3 binding elements. Together, these findings suggested the critical role of Runx3 in transcriptional upregulation of Bcl2l11 in TGF- -induced apoptosis[50]. In addition to antiproliferative and apoptotic effects contributing to gastric carcinogenesis, Runx3 affected the progression and metastasisofGCsaswell.Forexample,restorationofRunx3 stronglyinhibitedperitonealmetastasesofGCinananimal model [51], and inhibited the expression of vascular endothelialgrowthfactorA(VEGFA)andsuppressedthe angiogenesis, growth, and metastasis of GCs [52].

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topic Cdx2 expression, which suggested that Cdx2 plays a role in the development of intestinal metaplasia and subsequent gastric carcinogenesis [55]. Indeed, it has been demonstrated that gastric Cdx2 expression alone wassufficienttoinduceintestinalmetaplasiainmice[56], and long-standing intestinal metaplasia induces invasive GC in CDX2 transgenic mice [57]. In addition, Cdx2 expressionwasincreasedinhigh-gradedysplasiaandintestinal-type gastric adenocarcinomas compared to tionmayrepresentanearlyevent.However,somecontralowdictory studies have shown that Cdx2 expression grade dysplasia [58]. Taken together, these findings was sugprogressivelyreducedingastricdysplasiaandcancer[59, gest that Cdx2 expression in GC may contribute to 60], and Cdx2-positive GCs had a significantly the better progression of gastric carcinogenesis and that its outcome than Cdx2-negative GCs [61]. activaFurthermore, overexpression of Cdx2 significantly inhibited cell growth and reduced the motility and invasion of cancer cellsinvitro,whichsupportedthenotionthatCdx2plays a similar tumor-suppressive role in GC as in colorectal carcinoma[62].Althoughthedifferencesincutoffvalues todefineCdx2positivityorthelackofsubtypingcriteria for gastric epithelial dysplasia may in part explain these conflicting results [60], further studies are required to clarify the exact role of Cdx2 in gastric carcinogenesis.

Microsatellite Instability

Microsatellites are repeating sequences of 16 base pairs of DNA throughout the genome, and microsatellite instability is defined as length changes of these microsatellitescausedbyimpairmentoftheDNAmismatchrepair system. Microsatellite instability has been found in many sporadiccarcinomasaswellasinhereditarynonpolyposis colorectal cancer, a syndrome in which germ-line mutationsoftheDNAmismatchrepairgenes,MSH2orMLH1, are present [63]. Whereas microsatellite instability in Jang/ paKim tients with hereditary nonpolyposis colorectal cancer mainlyresultsfromgerm-linemutationsofoneorseveral DNAmismatchrepairgenes,somaticmutationsofMSH2

Caudal Type Homeobox 2 (CDX2)

CDX2 is a gene for an intestine-specific

Pathobiology 3 transcription 2011;78:302310 0 factor to direct intestinal development, 6 differentiation

and maintenance of the intestinal phenotype. Cdx2 not

Notably, GC associated with a gastric adenoma showed a higher frequency of the microsatellite instability phenotype compared to those carcinomas without an adenoma [72]. GC with microsatellite instability is a welldefined subsetofGCsexhibitingdistinctiveclinicopathologicfea shift mutations of type II receptor (90.3%), BAX (61.3%), hMSH3 (38.7%), and E2F4 TGF(TGFBRII) tures, such as antral location, intestinal type, (61.3%) expanding which have microsatellite sequences genes [70], growthpattern,lowerprevalenceoflymphnodemetastain the ses, and improved survival. Theyevents occur in one coding region. These mutational display frequent frameor two alleles of these genes, and accumulation of mutation mayalsoextendtogenesinvolvedingenomeintegrityincluding the mismatch repair genes, suggesting that these mutations result in malignant transformation [73].

positive advanced GC (ToGA, protocol BO18255). In addition, new therapeutic agents with multiple targets that include blocking the Erbb2 pathway are emerging [84].

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ERBB2 Gene

Erbb2 alteration plays an important role in the development and progression of many epithelial tumors. ERBB2 amplification is particularly frequent, especially in breast cancer, and treatment with trastuzumab, a monoclonal antibody to Erbb2 protein, has been shown to be highly effective in ERBB2-amplified breast cancer [74]. ERBB2 gene amplification and overexpression of Erbb2 protein in GC were also found in a large number of studies. However, the results are inconsistent and ranged from 4 to 16% when performed by FISH [75 77]. Interestingly, the amplification of ERBB2 is more common in intestinal-type GC than in diffuse-type GC [75]. In some studies, GC with ERBB2 amplification was significantly associated with poor outcome [77, 78]. On the Molecular Pathology of Gastric other hand, there are also several studies Carcinoma documenting that amplification was unrelated to prognosis [75, 76].

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histone modifications that result in altered gene Epigenetic Alterations sion without changing the coding sequence of the expresgene. Epigenetic mechanisms supports that epigenetic Growing evidence stronglyrefer to DNA methylation dysand regulation plays an essential role both independently and cooperatively in tumor initiation and progression. The bestcharacterizedmechanismsaretranscriptionalsilencing events that are associated with DNA hypermethylationatpromoterregionsofgenesthatregulateimportant cell functions. Promoter methylation affects virtually all of the pathways in the cellular network, such as DNA repair, cell cycle and apoptosis. It is well established that numeroustumorsuppressorgenescanbesilencedthroug h promoter CpG island methylation during carcinogenesis, and aberrant DNA methylation is the most common molecular lesion of cancer cells. Neither gene mutations nor cytogenetic abnormalities are as common in human tumorsasDNAmethylationalterations[85].Thus,aberrant CpG island methylation can be used as a biomarker of malignant cells and as a predictor of their prognosis. In particular, the reversibility of epigenetic changes has made them attractive targets for cancer treatment with modulators that demethylate DNA and inhibit histone deacetylases, leading to reactivation of silenced genes. Gastric cancer is one of the tumors with a high frequency of aberrant methylation, and it frequently shows the CpG island methylator phenotype [86]. A large Pathobiology 3 num2011;78:302310 0 ber of genes that are suppressed by CpG island 7 hypermethylation have been reported in GC, involving tumor

morinitiation.Indeed,aberrantmethylationwasfrequent ly detected in gastric intestinal metaplasia of both cancer andnoncancerpatients.Andthemethylationfrequencyof severalgeneswashigherinGCthaninintestinalmetaplasia[88].Onestudywithalargesamplecollectionofchronic gastritis, intestinal metaplasia, gastric adenoma and GC demonstrated that aberrant methylation occurred in early stages and tended to accumulate along the multistep gastric carcinogenesis [89]. The above findings suggest that progressiveepigeneticchangesplayanimportantroledur ing the progression of a premalignant lesion to cancer.

Conclusion

Cancer is widely considered as a heterogeneous

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group of diseases with markedly different biological properties that are attributed to a series of clonally selected genetic andepigeneticalterationsintumorsuppressorgenesand oncogenes [93]. Thus, identifying the characteristics of individual cancers from the point of view of molecular pathology has a great potential for future diagnosis and targeted therapy of cancer. Multiple genetic and epigenetic changes in oncogenes and tumor suppressor genes, Most epigenetic studies of GC have focused on cell cycle regulators, cell adhesion molecules and aberDNA rant methylation in a single or in several genes, but repair genes have been demonstrated to be implicated rein cently a genome-wide search has been tried to gastric carcinogenesis. Based on this emerging identify undernovel methylation-silenced genes in gastric cancer standing of the molecular pathways, several [90, targeted 91]. After treatment with a demethylating and/or therapies,suchassmall-molecularinhibitorsandantiandegiogenic agents, are currently being evaluated in acetyling agent of the GC cell line, upregulated genes GC were screened for epigenetically silenced genes using treatment [94]. In addition, the analysis of oli1 Parkin D, Bray F, Ferlay J, Pisani P: 13 Machado JC, Oliveira C, Carvalho R, hypermethgonucleotide microarrays. TFPI2 was found to be U, Christofori G: Cell Soares Global 7 Cavallaro ylation of cell cycle genes or DNA repair genes such highly adhesion as P, Berx G, Caldas C, Seruca R, Carneiro cancer statistics, 2002. CA and its methylated (81%) in GC, Cancer J methylation was a F, Clin andsignallingbycadherinsandIgCDKN2A or hMLH1 in nonneoplastic gastric sigCAMsin mucosa nificant and independent prognostic indicator in 2005;55:74108. Sobrinho-Simoes M: E-cadherin could be used to predict the risk of malignancy. Even GC gene 2 Jemal A, Siegel R, Ward E, Murray T, cancer. Nature Rev Cancer 2004;4:118 though the complex inherent molecular heterogeneity Xu J, [90]. Most recently, a genome-wide DNA 132. (CDH1) promoter methylation as the of methylation Smigal C, Thun M: Cancer statistics, second 8 Milne A, Carneiro F, OMorain C, GC still remains to be clarified, genome-wide analysisusingamethylationmicroarrayhasbeenapplied 2006. Offerhaus analysis to directly measure the methylation level of the CpG hit in sporadic diffuse gastric techniques will also carcinoma.understand the molecular help us CA Cancer J Clin 2006;56:106130. isfeatures of GC, which would provide further novel 3 Macdonald J, Smalley genome J, lands throughout theS, Benedetti and it has been G:Naturemeetsnurture:moleculargeneticsof HunOncogene 2001;20:15251528. opsuggested 14 Matysiak-Budnik T, Megraud F: gastriccancer.HumGenet2009;126:615 the treatment of GC. portunities in thatthishigh-throughputmethodwouldbegreatlyhelpdahl S, Estes N, Stemmermann G, Helico628. ful to find the novel methylation markers [92]. Haller D,

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AjaniJ,GundersonL,JessupJ:Chemoradio-

9 BlairV,MartinI,ShawD,WinshipI,KerrD, Arnold J, Harawira P, McLeod M, Parry S, Charlton A: Hereditary diffuse gastric cancer: diagnosis and management. Clin Gastroenterol Hepatol 2006;4:262275. 10 Huntsman DG, Carneiro F, Lewis FR,

bacter pylori infection and gastric cancer. Eur J Cancer 2006;42:708716. 15 Correa P, Fontham E, Bravo J, Bravo L, Ruiz

therapyaftersurgerycomparedwithsurgery alone for adenocarcinoma of the Pathobiology 3 stomach or 2011;78:302310 0

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