Our study investigated the relationship between MYC alterations and clinicopathological features in gastric cancers. We evaluated the effect of MYC mRNA expression and its protein immunoreactivity, as well as copy number variation, promoter DNA methylation, and point mutations, in 125 gastric adenocarcinoma and 67 paried non-neoplastic tissues. We observed that 77% of the tumors presented MYC immunoreactivity which was significantly associated with increased mRNA expression (p<0.05). These observations were associated with deeper tumor extension and the presence of metastasis (p<0.05). MYC protein expression was also more frequently observed in intestinal-type than in diffuse-type tumors (p<0.001). Additionally, MYC mRNA and protein expression were significantly associated with its copy number (p<0.05). The gain of MYC copies was associated with late-onset, intestinal-type, advanced tumor stage, and the presence of distant metastasis (p<0.05). A hypomethylated MYC promoter was detected in 86.4% of tumor samples. MYC hypomethylation was associated with diffuse-type, advanced tumor stage, deeper tumor extension, and the presence of lymph node metastasis (p<0.05). Moreover, eighteen tumor samples presented at least one known mutation. The presence of MYC mutations was associated with diffuse-type tumor (p<0.001). Our results showed that MYC deregulation was mainly associated with poor prognostic features and also reinforced the presence of different pathways involved in intestinal-type and diffuse-type gastric carcinogenesis. Thus, our findings suggest that MYC may be a useful marker for clinical stratification and prognosis.
References
[1]
Jemal A, Center MM, DeSantis C, Ward EM (2010) Global patterns of cancer incidence and mortality rates and trends. Cancer Epidemiol Biomarkers Prev 19: 1893–1907.
[2]
Nardone G (2003) Review article: molecular basis of gastric carcinogenesis. Aliment Pharmacol Ther 17 Suppl 2: 75–81.
[3]
Huppi K, Pitt JJ, Wahlberg BM, Caplen NJ (2012) The 8q24 gene desert: an oasis of non-coding transcriptional activity. Front Genet 3: 69.
[4]
Pelengaris S (2003) Khan M (2003) Arch Biochem Biophys 416 (2) 129–136.
[5]
Lutz W, Leon J, Eilers M (2002) Contributions of Myc to tumorigenesis. Biochim Biophys Acta 1602: 61–71.
[6]
Pelengaris S, Khan M, Evan GI (2002) Suppression of Myc-induced apoptosis in beta cells exposes multiple oncogenic properties of Myc and triggers carcinogenic progression. Cell 109: 321–334.
[7]
Pelengaris S, Littlewood T, Khan M, Elia G, Evan G (1999) Reversible activation of c-Myc in skin: induction of a complex neoplastic phenotype by a single oncogenic lesion. Mol Cell 3: 565–577.
[8]
Dalla-Favera R, Wong-Staal FGR (1982) Onc gene amplification in promyelocytic leukaemia cell line HL-60 and primary leukaemic cells of the same patient. Nature 299: 61–63.
[9]
Calcagno DQ, Leal MF, Assumpcao PP, Smith MAC, Burbano RR (2008) MYC and gastric adenocarcinoma carcinogenesis. World J of Gastroenterol 14: 5962–5968.
[10]
Calcagno DQ, Guimar？es AC, Leal MF, Seabra AD, Khayat AS, et al. (2009) MYC insertions in diffuse-type gastric adenocarcinoma. Anticancer Res 29: 2479–2483.
[11]
Liu Y, Gong L, Dong X, Liu H (2012) Detection of C-MYC oncogene translocation and copy number change in the normal-dysplasia-carcinoma sequence of the larynx by fluorescence in situ hybridization. Diagn Cytopathol doi: 10.1002/dc.22879 [In press].
[12]
Tuupanen S, Yan J, Turunen M, Gylfe AE, Kaasinen E, et al. (2012) Characterization of the colorectal cancer-associated enhancer MYC-335 at 8q24: the role of rs67491583. Cancer Genet 205: 25–33.
[13]
Amente S, Lania L, Majello B (2011) Epigenetic reprogramming of Myc target genes. Am J Cancer Res 1: 413–418.
[14]
Tsai KW, Liao YL, Wu CW, Hu LY, Li SC, et al. (2011) Aberrant hypermethylation of miR-9 genes in gastric cancer. Epigenetics 6: 1189–1197.
[15]
Dang CV (2012) MYC on the path to cancer. Cell 149: 22–35.
[16]
Da Costa JDFFB, Leal MF, Silva TCR, Andrade Junior EF, Rezende AP, et al. (2011) Experimental gastric carcinogenesis in Cebus apella nonhuman primates. PloS One 6: e21988.
[17]
Lauren P (1965) The two histological main types of gastric carcinoma: diffuse and so-called intestinal-type carcinoma. An attempt at a histo-clinical classification. Acta Pathol Microbiol Scand 64: 31–49.
[18]
Sobin L, Wittekind C (2002) TNM: Classification of malignant tumours. 6th ed. New York: Wiley-Blackwell.
[19]
Stamouli MI, Ferti AD, Panani AD, Raftakis J, Consoli C, et al. (2002) Application of multiplex fluorescence in situ hybridization in the cytogenetic analysis of primary gastric carcinoma. Cancer Genet Cytogenet 135: 23–27.
[20]
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(？Delta Delta C(T)) Method. Methods 25: 402–408.
Calcagno DQ, Leal MF, Taken SS, Assump？？o PP, Demachki S, et al. (2005) Aneuploidy of chromosome 8 and C-MYC amplification in individuals from northern Brazil with gastric adenocarcinoma. Anticancer res 25: 4069–4074.
[23]
Hopman AH, Ramaekers FC, Raap AK, Beck JL, Devilee P, et al. (1988) In situ hybridization as a tool to study numerical chromosome aberrations in solid bladder tumors. Histochemistry 89: 307–316.
[24]
Herman JG, Graff JR, My？h？nen S, Nelkin BD, Baylin SB (1996) Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. Proc Natl Acad Sci USA 93: 9821–9826.
[25]
Xiong Z, Laird P (1997) COBRA: a sensitive and quantitative DNA methylation assay. Nucleic Acids Res 25: 2532–2534.
[26]
Fang JY, Cheng ZH, Chen YX, Lu R, Yang L, et al. (2004) Expression of Dnmt1, demethylase, MeCP2 and methylation of tumor-related genes in human gastric cancer. World J Gastroenterol 10: 3394–3398.
[27]
Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74: 5463–5467.
[28]
Calcagno DQ, Leal MF, Seabra AD, Khayat AS, Chen ES, et al. (2006) Interrelationship between chromosome 8 aneuploidy, C-MYC amplification and increased expression in individuals from northern Brazil with gastric adenocarcinoma. World J Gastroenterol 12: 6207–6211.
[29]
Borges B, Santos E, Bastos C, Pinto L, Anselmo N, et al. (2010) Promoter polymorphisms and methylation of E-cadherin (CDH1) and KIT in gastric cancer patients from northern Brazil. Anticancer Res 30: 2225–2233.
[30]
Fernandez PC, Frank SR, Wang L, Schroeder M, Liu S, et al. (2003) Genomic targets of the human c-Myc protein. Genes Dev 17: 1115–1129.
[31]
Li F, Zhong MZ, Li JH, Liu W, Li B (2012) Case-control Study of Single Nucleotide Polymorphisms of PSCA and MUC1 Genes with Gastric Cancer in a Chinese. Asian Pac J Cancer Prev 13: 2593–2596.
[32]
Chang MS, Uozaki H, Chong JM, Ushiku T, Sakuma K, et al. (2006) CpG island methylation status in gastric carcinoma with and without infection of Epstein-Barr virus. Clin Cancer Res 12: 2995–3002.
[33]
Shibata D, Hawes D, Stemmermann GN, Weiss LM (1993) Epstein-Barr virus-associated gastric adenocarcinoma among Japanese Americans in Hawaii. Cancer Epidemiol Biomarkers Prev 2: 213–217.
[34]
Meyer N, Penn LZ (2008) Reflecting on 25 years with MYC. Nat Rev Cancer 8: 976–990.
[35]
Beroukhim R, Mermel CH, Porter D, Wei G, Raychaudhuri S, et al. (2010) The landscape of somatic copy-number alteration across human cancers. Nature 463: 899–905.
[36]
Assump？？o PP, Ishak G, Chen ES, Takeno SS, Leal MF, et al. (2006) Numerical aberrations of chromosome 8 detected by conventional cytogenetics and fluorescence in situ hybridization in individuals from northern Brazil with gastric adenocarcinoma. Cancer Genet Cytogenet 107: 45–49.
[37]
Burbano RR, Assump？？o PP, Leal MF, Calcagno DQ, Guimar？es AC, et al. (2006) C-MYC locus amplification as metastasis predictor in intestinal-type gastric adenocarcinomas: CGH study in Brazil. Anticancer Res 26: 2909–2914.
[38]
Calcagno DQ, Leal MF, Demachki S, Araújo MTF, Freitas FW, et al. (2010) MYC in gastric carcinoma and intestinal metaplasia of young adults. Cancer Genet Cytogenet 202: 63–66.
[39]
Costa Raiol LC, Figueira Silva EC, Mendes da Fonseca D, Leal MF, Guimar？es AC, et al. (2008) Interrelationship between MYC gene numerical aberrations and protein expression in individuals from northern Brazil with early gastric adenocarcinoma. Cancer Genet Cytogenet 181: 31–35.
[40]
Fan B, Dachrut S, Coral H, Yuen ST, Chu KM, et al. (2012) Integration of DNA copy number alterations and transcriptional expression analysis in human gastric cancer. PloS One 7: e29824.
[41]
Zhang D, Wang Z, Luo Y, Xu Y, Liu Y, et al. (2011) Analysis of DNA copy number aberrations by multiple ligation-dependent probe amplification on 50 intestinal type gastric cancers. J Surg Oncol 103: 124–132.
[42]
Buffart TE, Van Grieken NCT, Tijssen M, Coffa J, Ylstra B, et al. (2009) High resolution analysis of DNA copy-number aberrations of chromosomes 8, 13, and 20 in gastric cancers. Virchows Archiv 455: 213–223.
[43]
Park KU, Lee HE, Park DJ, Jung EJ, Song J, et al. (2009) MYC quantitation in cell-free plasma DNA by real-time PCR for gastric cancer diagnosis. Clin Chem Lab Med 47: 530–536.
[44]
Leal MF, Martins Do Nascimento JL, Da Silva CEA, Vita Lamar？o MF, Calcagno DQ, et al. (2009) Establishment and conventional cytogenetic characterization of three gastric cancer cell lines. Cancer Genet Cytogenet 195: 85–91.
[45]
Costa Guimar？es A, Gon？alves Quintana L, Ferreira Leal M, Satomi Takeno S, Pimentel Assump？？o P, et al. (2006) Aneuploidy of chromosome 8 detected by fluorescence in situ hybridisation in ACP01 cell line gastric adenocarcinoma. Clin Exp Med 6: 129–133.
[46]
Leal MF, Calcagno DQ, Borges da Costa JDFF, Silva TCR, Khayat AS, et al. (2011) MYC, TP53, and chromosome 17 copy-number alterations in multiple gastric cancer cell lines and in their parental primary tumors. J Biomed Biotechnol 2011: 631268.
[47]
Ribeiro HF, Alcantara DFA, Matos LA, Sousa JMC, Leal MF, et al. (2010) Cytogenetic characterization and evaluation of c-MYC gene amplification in PG100, a new Brazilian gastric cancer cell line. Braz J Med Biol Res 43: 717–721.
[48]
Zhao Y, Li JS, Guo MZ, Feng BS, Zhang JP (2010) Inhibitory effect of S-adenosylmethionine on the growth of human gastric cancer cells in vivo and in vitro. Chin J Cancer 29: 752–760.
[49]
Mehndiratta M, Palanichamy JK, Pal A, Bhagat M, Singh A, et al. (2011) CpG hypermethylation of the C-myc promoter by dsRNA results in growth suppression. Mol Pharm 8: 2302–2309.
[50]
Liu X, Cai H, Huang H, Long Z, Shi Y, et al. (2011) The prognostic significance of apoptosis-related biological markers in Chinese gastric cancer patients. PloS One 6: e29670.
[51]
Shah M, Ajani J (2010) Gastric cancer-an enigmatic and heterogeneous disease. JAMA 303: 1753–1754.
[52]
De Smet C, Loriot A (2013) DNA hypomethylation and activation of germline-specific genes in cancer. Adv Exp Med Biol 754: 149–166.
[53]
Portela A, Esteller M (2010) Epigenetic modifications and human disease. Nat Biotechnol 28: 1057–1068.
[54]
Du Y, Peng J, Sun A, Tang Z, Ling W, et al. (2009) Assessment of the effect of betaine on p16 and c-myc DNA methylation and mRNA expression in a chemical induced rat liver cancer model. BMC Cancer 9: 261.
[55]
Sharrard RM, Royds JA, Rogers S, Shorthouse AJ (1992) Patterns of methylation of the c-myc gene in human colorectal cancer progression. Br J Cancer 65: 667–672.
[56]
Fang JY, Zhu SS, Xiao SD, Jiang SJ, Shi Y, et al. (1996) Studies on the hypomethylation of c-myc, c-Ha-ras oncogenes and histopathological changes in human gastric carcinoma. J Gastroenterol Hepatol 11: 1079–1082.
[57]
Fang JY, Xiao SD, Zhu SS, Yuan JM, Qiu DK, et al. (1997) Relationship of plasma folic acid and status of DNA methylation in human gastric cancer. J Gastroenterol 32: 171–175.
[58]
Weng YR, Sun DF, Fang JY, Gu WQ, Zhu HY (2006) Folate levels in mucosal tissue but not methylenetetrahydrofolate reductase polymorphisms are associated with gastric carcinogenesis. World J Gastroenterol 12: 7591–7597.
[59]
Suzuki K, Suzuki I, Leodolter A, Alonso S, Horiuchi S, et al. (2006) Global DNA demethylation in gastrointestinal cancer is age dependent and precedes genomic damage. Cancer Cell 9: 199–207.
[60]
Frigola J, Solé X, Paz MF, Moreno V, Esteller M, et al. (2005) Differential DNA hypermethylation and hypomethylation signatures in colorectal cancer. Hum Mol Genet 14: 319–326.
[61]
Ehrlich M (2002) DNA methylation in cancer: too much, but also too little. Oncogene 21: 5400–5413.
[62]
Papaggeli PC, Kortsaris AC, Matsouka PT (2003) Aberrant methylation of c-myc and c-fos protooncogenes and p53 tumor suppressor gene in myelodysplastic syndromes and acute non-lymphocytic leukemia. J BUON 8: 341–350.
[63]
Shen L, Qui D, Fang J (1997) Correlation between hypomethylation of c-myc and c-N-ras oncogenes and pathological changes in human hepatocellular carcinoma. Zhonghua Zhong Liu Za Zhi 19: 173–176.
[64]
Li M, Chen WD, Papadopoulos N, Goodman SN, Bjerregaard NC, et al. (2009) Sensitive digital quantification of DNA methylation in clinical samples. Nat Biotechnol 27: 858–863.
[65]
Ryan KM, Birnie GD (1996) Myc oncogenes: the enigmatic family. Biochem J 314 (Pt 3) 713–721.
[66]
Correa P (1992) Human gastric carcinogenesis: a multistep and multifactorial process–First American Cancer Society Award Lecture on Cancer Epidemiology and Prevention. Cancer Res 52: 6735–6740.
[67]
Tahara E (2004) Genetic pathways of two types of gastric cancer. IARC scientific publications 327–349.
[68]
Smith MG, Hold GL, Tahara E, El-Omar EM (2006) Cellular and molecular aspects of gastric cancer. World J Gastroenterol 12: 2979–2990.
