Alterations of oncogenes and tumor suppressor genes in esophageal cancer in China
?.
Mutation Research4622000343–353
https://www.360docs.net/doc/6c17173324.html, r locate r reviewsmr
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Alterations of oncogenes and tumor suppressor genes in
esophageal cancer in China
Shih Hsin Lu)
Department of Chemical Etiology and Carcinogenesis,Cancer Institute,
Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing100021,China
Received30July1999;received in revised form22November1999;accepted22November1999
Keywords:Esophageal cancer;p53;Rb;MTS1r p16;Nitrosamine
1.Introduction
?.
Human esophageal cancer EC is one of the most common cancers worldwide and occurs at very high frequencies in certain areas of China,Iran,South
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Africa,Uruguay,France and Italy1.The mortality rate of EC in China is the highest in the world. Compared with cancer mortality in five continents, 50%of EC in the world occurs in China.In China, there are many high-incidence and high mortality rate areas of EC,located in the southern parts of the Taihang Mountains on the borders of Henan,Shansi and Hopei provinces.In particular,the Lin-xian county in Henen province has the highest age-ad-justed mortality rates,151r100000for males and
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115r100000for females2.The localized cluster-ing of this disease has been the focus of numerous epidemiological studies.These studies have revealed that several factors are associated with the incidence of EC.These factors include N-nitrosamines from food sources or endogenous formation and certain
)Tel.:q86-10-6778-1331;fax:q86-10-6771-3359;e-mail: shlu@https://www.360docs.net/doc/6c17173324.html,
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nutritional deficiencies in China3,and a high, combined consumption of alcohol and cigarette
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smoking in Western countries4.
Molecular studies of human esophageal tumors have revealed frequent genetic abnormalities.Some of these data were reviewed in detail by Montesano w x
et al.5.The present paper reviews recent research developments on EC in China.Special attention has been paid to the reports published in the Chinese literature and data from China will be compared with published reports from other parts of the world.
2.Epidemiological magnitude of EC
EC exists in two main forms with different etio-logical and pathological characteristics,squamous
?.?. cell carcinoma SCC and adenocarcinoma ADC. In China,primary esophageal adenocarcinoma ?.
PEAC is a rare malignant disease of esophagus, with an incidence much lower than that of SCC. According to the literature,patients with PEAC rep-
w x resented0.4%of EC patients in China6.However, the incidence of PEAC is much higher in Northern America and in some parts of Europe,and recent
1383-5742r00r$-see front matter q2000Elsevier Science B.V.All rights reserved.
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S.H.Lu r Mutation Research4622000343–353 344
data suggests that this incidence has a tendency to increase.The histological origins of PEAC have been a controversial issue for a long time.In general, there are three proposed origins for PEAC,namely: glands of the esophagus itself,aberrant mucosa of the stomach,or the columnar epithelium of the lower
?. segment of esophagus Barrett’s esophagus,BE. The prevalence of BE was0.69%among the Chinese rural population and1.6%in out-patients.The inci-dence rate of reflux esophagitis was8.9%among the Chinese rural population.Most western authors maintain that BE is the major origin of PEAC,whilst some authors from Asia argue that BE has little to do with PEAC,because of the different localization of
w x the tumors compared to the western samples7. Recently,some authors have indicated that PEAC,
?. esophageal squamous cell carcinoma ESCC and primary small-cell carcinoma of the esophagus all originated from the multipotential primitive stem cells of esophageal mucosa.This view also gives an explanation for the origin of adenoacanthoma,which may be the result of two different differentiating
w x processes of the same multipotential stem cell8. Most reports from China have suggested that PEAC and BE are uncommon diseases.The reasons for their lower frequency compared to the West are not known,and further studies are required.
3.Amplifications and expressions of oncogenes in EC
In previous studies on over100DNA samples obtained from Lin-xian patients who underwent surgery for EC,a significant frequency of amplifica-tion of either the human epidermal growth factor ?.
receptor HER-1gene or the C-MYC oncogene was found.These changes were found not only in tumor
?specimens,but also in adjacent non-tumor grossly .
normal tissue specimens obtained from patients with EC.RNA samples were also obtained from over30 tissues.These revealed considerable variation in the abundance of HER-1and C-MYC transcripts in both tumor and adjacent non-tumor specimens.A few samples revealed extremely high levels of these tran-w x
scripts9.In another study,the expression of HER-1 and C-MYC genes in30cases of esophageal carcino-mas and adjacent tissues were studied using hy-
?. bridization in situ.The results showed that:a the C-MYC and HER-1protooncogenes were transcrip-
?.
tionally active;b activation of C-MYC gene was observed in hyperplastic cells and carcinoma cells;?.c the degree of pathological changes of the esophageal epithelium was related to the level of C-MYC transcription,with the highest level of C-MYC expression observed in invasive carcinoma ?.
cells;d expression of HER-1gene in carcinoma cells was higher than in normal,adjacent non-tumor cells,but the frequency of expression was lower than
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that of C-MYC gene10.Thus,changes in copy numbers or level of expression of HER-1or C-MYC genes may play an important role in the pathogenesis of EC in this high-risk region.
?
The human CYCLIN D gene also referred to as .
PRAD1has been mapped to chromosome11ql3.A co-amplification of CYCLIN D and HST-1genes
?.
was found in5of2025%human squamous esophageal tumors.We also detected significant lev-els of CYCLIN D transcription in two esophageal carcinoma cell lines,even though they did not show detectable levels of HST-1transcription.These find-ings provided the first evidence for the amplification of a cyclin gene in human EC and suggested that an increase in CYCLIN D gene dosage could be an important factor in the pathogenesis of EC.Addition-ally,because the11q13locus was found to be ampli-fied in many types of human tumors,CYCLIN D gene amplification could also play an important role in the development of other forms of human cancer w x
11.
Amplification of INT-2gene in primary esophageal carcinoma and adjacent tissues,in fetal esophageal carcinoma induced by NMBzA and in fetal esophageal epithelium treated with NMBzA was detected.Amplification of INT-2gene was ob-served in23of52human esophageal carcinomas ?.?. 44.2%and in7of18adjacent tissues38.9%.We found amplification of INT-2in fetal esophageal carcinoma induced by NMBzA,and in fetal esophageal epithelium treated with NK4BzA for1 month.These results suggest that NMBzA may acti-vate INT-2gene expression in the esophageal epithe-lium to induce the esophageal carcinoma,indicating that NMBzA may play an important role in the
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development of EC in humans12.
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S.H.Lu r Mutation Research4622000343–353345
4.Alterations of TP53gene
4.1.Mutations of TP53
The TP53gene has been shown to be commonly mutated in various human cancers,and mutated p53 can act as a dominant oncogene.The mutations of TP53gene in human EC and cancer of the gastric cardia collected from Lin-xian county,Henan province,China,were analyzed.PCR-direct sequenc-ing technique was used to detect TP53point muta-tions within exons and introns5to9.Fifteen of30?.
50%of ESCCs contained a mutation of TP53.Five ?.
of1145%adjacent,non-tumor tissues also con-tained a mutation of TP53.Three of four carcinomas of gastric cardia showed TP53mutation.The muta-
?. tion spectrum in EC shows that8of22cases36.4% of mutations were G:C to A:T transition,6of22?.?. 27.3were frameshift mutations,13.6%3r22were
?.
insertions and9.1%2r22were deletions.None of the G:C?A:T mutations occurred at a CpG site. Some new sites of p53mutation in human EC were w x
identified13.In another study,carcinomas of the
?
upper digestive tract SCC of the esophagus,ADC of .
the cardia from24patients residing in Lin-xian county and nearby high-incidence areas were ana-lyzed for mutations in exons5–8of the TP53gene. Mutations were identified by PCR-direct sequencing in50%of the specimens.Eleven tumors harbored a single base-pair substitution leading to either an
?.
amino-acid substitution eight tumors or a chain-
?.
termination signal three tumors,and one tumor revealed a15-bp deletion in exon7with a silent base substitution adjacent to the deletion site.Mutations occurred in all four exons examined with a prepon-derance in exon5.Of the six mutations identified among the14adenocarcinomas examined,three were G to T transversions,a mutation that has thus far been absent from mutations in Barrett’s esophageal adenocarcinomas and dysplasia from patients in Eu-
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rope and North America14.
The mutations of p53gene in ESCC from Lin-xian are distributed throughout exons5–9,at codons 173,180,187,194and307.These codons are the most frequently mutated,with three to four muta-tions observed at each site in22cases of ESCC. Whether there are hostpots for mutation in EC re-
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mains to be determined13.In another study,a mutational spectrum for exons5–8of the TP53 tumor suppressor gene in209cases of SCC speci-mens obtained from five different geographical lo-
?
cales in China Zhengzhou,Taiyuan,Shantou,
.
Guangzhou,and Hong Kong was studied.This study involved Zhengzhou and Shantou,which were high-incidence regions for EC,whereas the other three regions had low or intermediate incidence of the disease.Analysis by single-strand conformation polymorphism and DNA sequencing showed that87?.
specimens41.6%contained mutations in exons 5–8of the p53gene.Point mutations accounted for ?.
80.4%87r107of all genetic changes.The speci-mens from northern China exhibited fewer p53gene aberrations and a more even distribution of muta-tions in exons5–8compared to those from southern China in which60%of all mutations were found in exon5.A major hot spot was found at codon176in exon5,where41samples from Shantou,Guangzhou, and Hong Kong had a G?T transversion.It is likely that among southern Chinese this codon is susceptible to mutagenesis by carcinogens.Codons 175,203,245,250,273,and282were also shown to be mutational hot spots,with three or more muta-
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tions observed at each site15.The p53mutational data obtained in China showed that Chinese esophageal carcinomas are often associated with some unique genetic alterations,which may be at-tributed to specific dietary or environmental carcino-gens that affect the Chinese depending geographical locales and human species.
We have compared the p53mutations in EC from
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China and France16.The data listed in Table1 show that the mutation spectrum of p53in EC from Lin-xian and France was not significantly different ?.
p)0.05.The results demonstrate that alterations of bases in TP53in EC from both Lin-xian and France were similar in frequency,but that they oc-curred at different positions,suggesting that the pa-tients were exposed to different carcinogens.This difference in the distribution of mutational specifici-ties suggests the involvement of different etiological w x
factors17.
4.2.Expression of p53protein
In normal cells,p53protein is virtually unde-tectable by immunohistochemical methods.The im-
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S.H.Lu r Mutation Research4622000343–353
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Table1
Comparison of TP53mutations in EC from China and France
?.?.?.?. Country No.of Mutation Mutation G:C?%CpG transversion%Mutation A:T%?Frameshift% A:T T:A C:G T:A G:C C:G
?.?.?.?.?.
Lin-xian22834.8627.30012 4.509.1522.7
?.?.?.?.?.?.?.
France21838.1628.6314.3314.329.51 4.71 4.7
P value)0.05)0.05)0.05)0.05)0.05)0.05)0.05
munoreactivity of p53protein is a general indicator of tumors with altered p53function.EC samples obtained from Lin-xian patients who had undergone surgery were assayed for expression of p53protein. Among18primary EC and their adjacent tissues, overexpression of p53protein were detected im-munohistochemically in seven cases of EC and five adjacent tissues.In the cases where overexpression of p53in adjacent tissues was detected,four were also positive for p53expression in the carcinomas. These results suggest that overexpression of p53 protein is a common molecular event in EC and may occur in the early stage of esophageal tumorigenesis. In addition,overexpression of the p53was found in human fetal esophageal carcinoma induced by NM-
?. BzA,indicating that p53gene mutation s might
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have occurred18.
The correlation between p53protein accumulation and mutations of the TP53gene in human EC from Lin-xian was analyzed by immunohistochemistry for p53protein accumulation,and the results were com-pared to TP53gene mutations by PCR-direct se-quence analysis.A highly significant correlation be-tween the presence of mutations and nuclear protein
?. accumulation was found.Of33tumors,2369.7%
?. demonstrated p53protein expression and1236% had TP53mutations.Of these12tumors with TP53
?.
mutations,975%showed intensive nuclear p53 reactivity.The mutations were found at different
w x codons,with no mutational hot spots19.This result is similar to that of studies in Indian populations, which have dietary habits and environmental factors that are considerably similar to those of the Chinese w x
population20.In another study,abnormalities of the p53protein were also analyzed by immunohisto-chemical techniques in15cases of esophageal squa-mous cell carcinoma.Five cases of EC were also analyzed for TP53gene mutation by PCR chain reaction and direct nucleotide sequencing.Overex-
?. pression of p53protein was found in5r1533%of ECs.Immunodetectable p53was found in10r14 cases with mutations and in none of11cases without mutations in ECs.Hence,immunohistochemical and genetic analyses gave concordant results in84%of the cases,revealing a good correlation between im-munostaining of p53and missense mutation of the TP53gene.Positive immunostaining was not ob-served in cases with frameshift or splicing mutations w x
21.In addition,similar studies have conducted in
w EC and adjacent epithelium by other scientists22–x
25.These results suggest that abnormalities of p53 may be closely associated with the pathogenesis of esophageal squamous cell carcinoma and that the immunoreactivity of p53protein is a general indica-tor of the tumors with altered p53function.
5.Mutation and expression of Rb gene
pRB,the retinoblastoma tumor suppressor gene product,negatively regulates the cell cycle at the G1r S transition.Many cell cycle regulators modu-late pRB function through its phosphorylation status. In different types of cancer,Rb gene alteration or functional inactivation of pRB has been reported.In EC,loss of heterozygosity of Rb gene was fre-
w x
quently detected26.Mutation and expression of Rb in human EC from Lin-xian were investigated by PCR-direct sequencing and Northern blot hybridiza-tion.In PCR amplification analysis,one of20cases of EC was found to have an Rb gene deletion in exon17and21;one of six the adjacent non-tumor-ous tissues was found to have deletion in exon21.In
?. PCR-direct sequencing analysis,four of1040% EC were found to contain Rb gene mutation in exons17and21at codons501,503,511and703,
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S.H.Lu r Mutation Research4622000343–353347
703,719,respectively.A hot spot was found at codon511in exon17,with two mutations observed at this site,and the mutations were an A?G transi-?.
tion GAT?GGT.In Northern blot analysis,7of ?.
1258.3%EC exhibited abnormal Rb gene expres-w x
sion27.The results suggest that Rb gene might play an important role in the carcinogenesis of EC in China.
6.Mutation of APC and MCC
Mutation and deletion of APC and MCC genes in human EC were analyzed by PCR amplification and direct-sequencing assay.In PCR amplification analysis,one of10cases of EC was found to have APC gene deletion in exon11;one of10cases of EC was found to be deleted for MCC gene in exon 12.One of adjacent,non-tumor tissue was also found to have the same deletion at exon12of MCC gene. In PCR direct-sequencing analysis,two of10cases of EC were found to contain APC gene mutations in exon11.Two of seven cases of EC were found to
?. contain MCC gene mutations in exon12Table2. The results confirmed that mutation of APC and MCC genes exists in human EC,although they are
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relatively rare28.
7.Deletion and mutation of MTS1r p16gene
?
The MTS1r p16gene also called INK4a or .
CDKN2,encodes two functionally related CDK in-hibitors,recently emerged as candidate tumor sup-pressor gene.p16binds to CDK4and inhibits the catalytic activity of the CDK4r cyclin D enzyme.
Table2
Mutations of APC and MCC genes in human EC
Gene Code Exon Codon Sequence Bases Amino acid APC111493AAT?AGT A?G Asn?Ser 211498ATT?GTT A?G Ile?Val
311del
MCC112522CGG?CAG G?A Arg?Gln 12541GCC?ACC G?A Ala?Thr 212541GCC?ACC G?A Ala?Thr
312del p16seems to act in a regulatory feedback circuit with CDK4,D-type cyclins and retinoblastoma pro-tein.
Mutations and deletions of MTS1r p16gene ?.
chromosome9p21,are among the most frequent genetic abnormalities in human neoplasia,including
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esophageal carcinoma29–35.A total of60human esophageal tissue specimens,comprising30SCCs and30tumor adjacent tissue specimens collected from Lin-xian were examined for homozygous dele-tion of p16gene by using Southern blot hybridiza-tion and PCR method.The results showed that no deletions were detected in30tumor adjacent tissue samples.However,of30esophageal carcinoma spec-imens,seven were found negative for p16gene in Southern blot analysis,and deletion of the p16gene in five samples was confirmed by PCR,with a 16.7%p16deletion rate.Most deletion of the p16 gene were revealed in II or III stages of EC.In DNA sequencing analysis,3of12cases of EC were found to carry mutations in exon2of p16gene.One case was a CAC?CAT mutation,two cases were found to be frameshift mutations,and one case was a tandem base mutation.These data suggest that MTS1r p16gene alterations may play a role in the
w x progression of human esophageal carcinoma36. Multiple molecular analyses in other60ESCC speci-mens from Lin-xian,China,have been performed using DNA methylation assay,LOH analysis,dele-tion screening and SSCP-sequencing.The results showed that MTS-1r p16inactivation was predomi-nantly associated with aberrant methylation in the CpG island of its promoter https://www.360docs.net/doc/6c17173324.html,pared with aberrant methylation,which occurred in17of34 cases,homozygous deletion of p16and LOH at its nearby D9S942microsatellite marker were observed
?.
at a much lower frequency17%.Intragenic muta-tion in p16gene was rare.These results suggest that the alteration mode at9p21was not uniform,indicat-ing that p16gene is a frequent target for inactivation
w x
during ESCC development37.
The Transkei region of Eastern Cape Province, South Africa,is a high-incidence area for EC.DNA
?.
from human ESCCs n s76,as well as adjacent
.?.
tissue samples n s9and blood n s50from the same patients from the Transkei region were screened for somatic mutations.Exons1–2of the p16gene were examined for mutations using PCR-SSCP pro-
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S.H.Lu r Mutation Research4622000343–353 348
cedures and DNA sequence analysis.Among muta-tions found in the p16gene,nine were point muta-tions,four were deletions and three were insertions.
A novel C to T mutation,25bp upstream from the ATG start site of the p16gene,was found.The mutations are somatic in origin since none of the DNA samples from the adjacent control tissue or blood samples from the same patients contained w x
them38.The overall pattern of mutation of the p16 gene in human esophageal carcinomas from Lin-xian county,Henan province,China and from Transkei region was same,but differ from the one in Hong Kong.
8.Effect of N-nitrosamine on oncogenes in esophageal tissue
EC,as other tumors,results from the interactions between the environmental and genetic factors.In this respect,studies on the effect of nitrosamines on the genes in esophageal tissue are of important sig-nificance in the carcinogenesis.N-nitrosamine can
w x
induce tumors in many animal species39.Epidemi-ological investigation showed that N-nitroso com-pounds,particularly N-methylbenzylnitrosamine ?.
NMBzA,have been associated with increased inci-
?.
dence of esophageal cancer EC in Lin-xian county w x
40–46.In addition,molecular studies of human EC have revealed gene alterations and abnormalities, which might have been induced by environmental ?.
factors such as N-nitrosamines.
8.1.Effect of N-nitrosamine in human fetal esophageal tissues
Epidemiological studies have shown that ni-trosamine are involved in the carcinogenesis of EC w x
in China46.NMBzA was isolated and identified in diets and gastric juice collected from Lin-xian county w x
47.Molecular biological studies indicate that NM-BzA can induce amplification and over-expression of EGFR gene in human fetal esophageal epithelium ?.
HFEE treated with NMBzA for24h,as shown by Southern blot assay and immunohistochemistry.Pap-illary hyperplasias were induced in HFEEs cultured with NMBzA for1to3weeks.Amplifications of C-MYC and INT-2genes were found in HFEEs treated by NMBzA for1week and3weeks,respec-w x
tively48.
8.2.Effect of N-nitrosamine in monkeys
Mutations of ras oncogene in esophageal epithe-lium of rhesus monkey fed with one dose of NMBzA ?.
30mg r kg B.W.,which was found in high inci-dence areas of EC in China,were analyzed by PCR and direct sequencing.Mutation at codon12of H-RAS gene was not found in esophageal epithelium
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of monkey fed with NMBzA49.Molecular epi-demiological studies have shown that the mutation at codon12of H-RAS gene were also not found in the human EC from France,South Africa and China. Thus,activating mutations in RAS oncogenes appear
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to be absent or rare in human EC50–52.
8.3.Effect of N-nitrosamine in human fetal esophageal tissues
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Barch et al.53has reported that the H-RAS
?oncogene was mutated by point mutation G?A at .
codon12in NMBzA-induced esophageal tumorige-nesis in rats.These results differ from H-RAS muta-tions in esophageal tissues of human and monkey. These observations indicate that there are important differences among species in the effects of ni-trosamines on the genome of esophageal cells.
9.Effect of N-nitrosamine on tumor-suppressor genes in esophageal tissue
() 9.1.In human fetal esophageal epithelium HFEE
?
Several tumor suppressor genes Rb,p53,APC .
and MCC were analysed in HFEE treated with ?.
NMBzA in vitro for24h or3weeks and in esophageal carcinoma induced by NMBzA.In PCR amplification analysis,Rb,p53,APC and MCC gene deletions in esophageal carcinoma of human fetus induced by NMBzA were found,but no dele-tions of these genes were revealed in HFEE treated by NMBzA in vitro.In PCR direct sequencing analy-sis,mutations of TP53gene were found in HFEE treated with NMBzA for3weeks,including G:C?
?. T:A transversions in codon146TGG?TTG and
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S.H.Lu r Mutation Research4622000343–353349
?.X
173GTG?GTT,and at the5end of intron8?.
C?A.Mutations in MCC gene in HFEE treated with NMBzA for3weeks were G:C?A:T transi-
?.?
tions in codon522CGG?CAG and541GCC?.w x?. ACC49.These results confirmed in vitro that nitrosamine can cause mutations and deletions of multiple tumor suppressor genes in human esophageal epithelium.The mutations of tumor suppressor genes in nitrosamine-induced EC may occur in the early stage of carcinogenesis,while deletions may occur at later stages.
Overexpression of p53protein in human fetal esophageal carcinomas was detected by immunohis-tochemical methods and indicated that TP53gene ?.
mutation s may have occurred.HFEE explants treated in vitro with NMBzA for3weeks were inoculated subcutaneously into Balb r c nude mice. No tumor was found within5months after inocula-
?. tion,suggesting that the changes in oncogene s observed in vitro are insufficient to induce full trans-
?formation.Other genetic alterations such as addi-tional,functional inactivation of Rb or r and p53
.
tumor suppressor genes may be necessary for fur-ther progression of malignant lesions.
9.2.In monkeys
Mutations of multiple tumor suppressor genes p53,Rb and APC were found in the esophageal epithelium of rhesus monkey fed with one dose of ?.
NMBzA30mg r kg, B.W..Some mutations of TP53gene were found in esophageal epithelium of monkey after feeding with NMBzA for24–48h. They included a G:C?T:A transversion at codon ?.
173GTG?GTT and a G:C?A:T transition at ?.
codon180GAG?AAG.In addition,a C to T mutation was also found in intron8near to the splice
?. acceptor site tctttcctagCA?tttttcctagCA.A muta-
?
tion of Rb gene was also found C?T transition in
.
codon503,TCT?TTT,as well as in APC gene ?.
A?G transition in codon498;ATT?GTT.The mutation fingerprints of these genes disappeared in esophageal epithelium of monkey after feeding with NMBzA for5days.These results demonstrated that chemical carcinogen NMBzA can induce mutations
?of multiple tumor suppressor genes in monkey in .
vivo and indicated that alteration of tumor suppres-sor genes in the initial stage of carcinogenesis needs many hits by chemical carcinogen.These alterations of p53,Rb and APC genes were similar to the changes of these genes reported previously in some
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primary EC54.Overall,the results suggest that NMBzA may be an esophageal etiological factor for human EC in China.
9.3.In rats
In a study of exon5–8and exon–intron junctions of the TP53gene from rat esophageal papillomas
w x?.
induced by NMBzA55,9of3030%esophageal papillomas contained SSCP mobility shifts,princi-pally within exons5and7.These positive SSCP findings were further validated by direct DNA se-quencing.Eight of the nine mutations were G:C?
?.
A:T transitions in codons131,149,153,2422,243, 248,and at the5X end of intron7.None of these G:C?A:T mutations occurred at CpG sites.The other mutation was a frameshift mutation at condon 176.The G:C?A:T transitions observed in the human fetal tissues,monkey and rats are consistent with the documented formation of O6-methylguanine adducts in DNA induced by nitroso compounds. These results suggest that point mutations of the p53 gene are involved in the development of EC.
10.Allelic losses in human EC
?.
Studies on loss of heterozygosity LOH have been utilized to search for potential regions in the genome harboring putative tumor suppressor genes
w x
that are important in tumor pathogenesis56.In EC, frequent LOH has been detected at several chromo-somal locations,particularly chromosomes3,4,5,9,
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13,17and184,57.However,whether other candi-date tumor suppressor genes mapped to different chromosomal arms,except for chromosome17,has not been clearly defined.Recently,PCR-based LOH analysis using microsatellite and RFLP markers has prove to be an effective strategy in detecting tumor-specific involvement of chromosomal regions rele-vant in the genesis and r or progression of EC.LOH of some chromosomal regions in EC has been re-
w x
vealed and are listed in Table358–66.This ap-proach has been an important element for elucidating the tumor-suppressor function of the retinoblastoma
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S.H.Lu r Mutation Research4622000343–353
350
Table3
Allelic loss in squamous cell carcinoma of human esophagus
Chromosome No.of cases Area of loss Candidate gene Mutation Reference
?.w x
1p2r258%1p36p73no Nimura et al.58
?.w x
3p11r5022%3p14.2FHIT Zou et al.59
?.w x
4p5r1533.3%Moskaluk and Rumpel60
?.w x
4q5r1533.3%4q10–24Moskaluk and Rumpel60?.w x
5p44%5p15.2Peralta et al.61
?.
5q62%5q31.1
?.w x
35r6157%5q31.1IRF-1no Ogasawara et al.62
?.w x 8r1553.3%5q12–21Moskaluk and Rumpel60
?.w x w x 9p17r3450%9p21p16,p150–25Muzeau,et al.29;Xing et al.37?.w x
11p53.3%11p15Moskaluk and Rumpel60?.
14.3%11p22–23.3
11q11q13Cyclin D1
Exp1,EMS1
?.w x
17q37r5271%17q25Iwaya et al.63
?.w x
18q5r1435%18q21.1Smad2no Maesawa et al.64
w x
Smad4Lei et al.65
w x
21q50%Mayama et al.66
Rb and p53gene localized on chromosomes13q14
w x
and17p13,respectively67–69.
11.Conclusion:mechanism of carcinogenesis in EC
The occurrence and development of tumors is a result of interaction between the environmental fac-tors and genetic factors,in the case of EC as well as in the case of other cancers.Environmental carcino-gens have repeatedly been shown to affect the ge-netic material of host cells,inducing uncontrolled growth and,ultimately cancer.Thus,human EC carcinogenesis is a multi-factorial,multi-step and multi-gene process.
Molecular studies of human EC have revealed gene alterations and chromosomal abnormalities in-?.
cluding:a amplification of the cellular oncogene C-MYC,the epidermal growth factor receptor gene w x w x?.
9and INT-2gene12.b Amplification and ex-
w x?. pression of the human CYCLIN D10.c Mutations and deletions of Rb,p53,p16,APC,MCC and DCC in EC.These alterations of oncogenes and tumor suppressor genes play a role in the different steps of carcinogenic process in the esophageal ep-ithelia.NMBzA isolated and identified from Lin-xian county induced alterations of oncogenes and tumor suppressor genes in esophageal epithelia of human fetuses and of monkeys,with a pattern of alterations similar to changes of genes in primary human EC. Cytogenetic studies have revealed loss of heterozy-gosity at the retinoblastoma gene locus and at the TP53gene locus on chromosome13and17,respec-tively.In addition,cytogenetic studies have also revealed breakpoints and deletions at1pter-21,3p21
w x
and9p21in EC57.These molecular and cytoge-netic studies suggest that many oncogenes and tumor suppressor genes are involved in the initiation and
?.
development of EC Fig.1.However,no gene directly associated with EC has been found so far. Searching for such genes,whose activation or inacti-vation will induce the carcinogenesis in the
esopha-
Fig. 1.Activation of oncogenes on the multistep process in carcinogenesis of human esophageal carcinoma.
()
S.H.Lu r Mutation Research4622000343–353351
gus,is of great significance in the basic and clinical study on EC.In view of the above findings,we consider that cloning and identifying the genes re-lated to human EC between normal and cancerous epithelia of esophagus using differential display method is a significant and necessary approach. Acknowledgements
The research in my laboratory is funded by the National‘973’Program on Cancer Research G1998051204and was supported by the National Key Program on Cancer Research.
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