ABSTRACT Background: Esophageal adenocarcinoma (EAC) has an extremely poor prognosis. There is a need to characterize the molecular alterations in the carcinogenesis of EAC in order to improve the diagnosis and treatment. Materials and Methods: We used 7 markers to explore the changes in the cell cycle, proliferation and apoptosis in patients with EAC and Barrett’s esophagus (BE)-associated dysplasia. The protein expression of Ki-67, PCNA, pRb, p16, p53, Bcl-2 and Bax was evaluated by immunohistochemistry in archival tissue samples, collected from 15 patients with EAC and 5 patients with BE-associated dysplasia. We analyzed also lymph-node, omentum and liver metastases from the primary esophageal tumors. Results: Ki-67, PCNA, pRb, p16, p53, Bcl-2 and Bax expression was observed in 100%, 87%, 60%, 40%, 100%, 7% and 93% of tumor samples, and in 100%, 80%, 0%, 80%, 80%, 20% and 100% of dysplasia samples, respectively. Significant difference in the expression of the markers between EAC and BE-associated dysplasia was detected for pRb (p = 0.006). Ki-67 expression was associated with clinicopathological parameter T (p = 0.012; V = 0.585). Ninefold higher risk to develop EAC was established for the patient with strong p53 expression, than the lacking p53 patient. Patients with strong p53 expression survived 6.8 months longer than the patients with weak p53 expression and 8.6 months longer than the patients with moderate p53 expression. No correlation was found between the expression of the other markers and prognosis. Conclusion: The results suggest that Ki-67, PCNA, pRb, p16, p53 and Bax participate in the pathogenesis of EAC, whereas Bcl-2 does not play essential role in EAC and BE-associated dysplasia. The balance between cell proliferation and apoptosis is lost in EAC and BE-associated dysplasia. Abnormal p53 protein expression has predictive and prognostic value in EAC. Larger prospective studies are needed to confirm these findings.
Cite this paper
A. Kotzev, M. Kamenova and A. Tcherveniakov, "Immunohistochemical Expression of Ki-67, PCNA, pRb, p16, p53, Bcl-2 and Bax in Esophageal Adenocarcinoma and Barrett’s Associated Dysplasia," Journal of Cancer Therapy, Vol. 3 No. 6, 2012, pp. 1092-1110. doi: 10.4236/jct.2012.36143.
 D. A. Corley, T. R. Levin, L. A. Habel, N. S. Weiss and P. A. Buffler, “Surveillance and Survival in Barrett’s Adenocarcinomas: A Population-Based Study,” Gastroenterology, Vol. 122, No. 3, 2002, pp. 633-640.
 E. Bollschweiler, E. Wolfgarten, C. Gutschow and A. H. Holscher, “Demographic Variations in the Rising Incidence of Esophageal Adenocarcinoma in White Males,” Cancer, Vol. 92, No. 3, 2001, pp. 549-555.
 J. Lagergren, R. Bergstrom, A. Lindgren and O. Nyrén, “Symptomatic Gastroesophageal Reflux as a Risk Factor for Esophageal Adenocarcinoma,” The New England Journal of Medicine, Vol. 340, No. 11, 1999, pp. 825-831.
 A. J. Cameron, B. J. Ott and W. S. Payne, “The Incidence of Adenocarcinoma in Columnar-Lined (Barrett’s) Esophagus,” The New England Journal of Medicine, Vol. 313, No. 14, 1985, pp. 857-859.
 M. Lindblad, L. A. Rodríguez and J. Lagergren, “Body mass, Tobacco and Alcohol and Risk of Esophageal, Gastric Cardia, and Gastric Non-Cardia Adenocarcinoma among Men and Women in a Nested Case-Control Study,” Cancer Causes and Control, Vol. 16, No. 3, 2005, pp. 285-294. doi: 10.1007/s10552-004-3485-7
 M. B. Cook, F. Kamangar, D. C. Whiteman, et al., “Cigarette Smoking and Adenocarcinomas of the Esophagus and Esophagogastric Junction: A Pooled Analysis from the International BEACON Consortium,” Journal of the National Cancer Institute, Vol. 102, No. 17, 2010, pp. 1344-1353. doi:10.1093/jnci/djq289
 N. D. Freedman, L. J. Murray, F. Kamangar, et al., “Alcohol Intake and Risk of Oesophageal Adenocarcinoma: A Pooled Analysis from the BEACON Consortium,” Gut, Vol. 60, No. 8, 2011, pp. 1029-1037.
 C. A. González, G. Pera, A. Agudo, et al., “Fruit and Vegetable Intake and the Risk of Stomach and Oesophagus Adenocarcinoma in the European Prospective Investigation into Cancer and Nutrition (EPIC-EURGAST),” International Journal of Cancer, Vol. 118, No. 10, 2006, pp. 2559-2566. doi:10.1002/ijc.21678
 P. Terry, J. Lagergren, H. Hansen, A. Wolk and Nyrén O, “Fruit and Vegetable Consumption in the Prevention of Oesophageal and Cardia Cancers,” European Journal of Cancer Prevention, Vol. 10, No. 4, 2001, pp. 365-369.
 N. Shaheen and D. F. Ransohoff, “Gastroesophageal Re-Flux, Barrett Esophagus, and Esophageal Cancer: Scientific Review,” Journal of the American Medical Association, Vol. 287, No. 15, 2002, pp. 1972-1981.
 J. Jankowski, D. Provenzale and P. Moayyedi, “Esophageal Adenocarcinoma Arising from Barrett’s Metaplasia Has Regional Variations in the West,” Gastroenterology, Vol. 122, No. 2, 2002, pp. 588-590.
 W. Meyer, F. Vollmar and W. B?r, “Barrett-Esophagus Following Total Gastrectomy. A Contribution to It’s Pathogenesis,” Endoscopy, Vol. 11, No. 2, 1979, pp. 121-126.
 M. T. Barrett, C. A. Sanchez, P. C. Galipeau, K. Neshat, M. Emond and B. J. Reid, “Allelic Loss of 9p21 and Mutation of the CDKN2/P16 Gene Develop as Early Lesions during Neoplastic Progression in Barrett’s Oesophagus,” Oncogene, Vol. 13, No. 9, 1996, pp. 1867-1873.
 M. T. Barrett, C. A. Sanchez, L. J. Prevo, et al., “Evolution of Neoplastic Cell Lineages in Barrett Oesophagus,” Nature Genetics, Vol. 22, No. 1, 1999, pp. 106-109.
 D. J. Wong, T. G. Paulson, L. J. Prevo, et al., “pl6INK4a Lesions Are Common, Early Abnormalities That Undergo Clonal Expansion in Barrett’s Metaplastic Epithelium,” Cancer Research, Vol. 61, No. 22, 2001, pp. 8284-8289.
 P. L. Blount, P. C. Galipeau, C. A. Sanchez, et al., “17p Allelic Losses in Diploid Cells of Patients with Barrett’s Esophagus Who Develop Aneuploidy,” Cancer Research, Vol. 54, No. 9, 1994, pp. 2292-2295.
 P. C. Galipeau, L. J. Prevo, C. A. Sanchez, G. M. Longton and B. J. Reid, “Clonal Expansion and Loss of Heterozygosity at Chromosomes 9p and 17p in Premalignant Esophageal (Barrett’s) Tissue,” Journal of the National Cancer Institute, Vol. 91, No. 24, 1999, pp. 2087-2095.
 J. A. Jankowski, N. A. Wright, S. J. Meltzer, et al., “Molecular Evolution of the Metaplasia-Dysplasia-Adeno-carcinoma Sequence in the Esophagus,” The American Journal of Pathology, Vol. 154, No. 4, 1999, pp. 965-973.
 C. C. Maley, P. C. Galipeau, J. C. Finley, et al., “Genetic Clonal Diversity Predicts Progression to Esophageal Adenocarcinoma,” Nature Genetics, Vol. 38, No. 4, 2006, pp. 468-473. doi:10.1038/ng1768
 M. K. Hong, W. B. Laskin, B. E. Herman, et al., “Expansion of the Ki-67 Proliferative Compartment Correlates with Degree of Dysplasia in Barrett’s Esophagus,” Cancer, Vol. 75, No. 2, 1995, pp. 423-429.
 L. Yacoub, H. Goldman and R. Odze, “Transforming Growth Factor-Alpha, Epidermal Growth Factor Receptor, and MiB-1 Expression in Barrett’s-Associated Neoplasia: Correlation with Prognosis,” Modern Pathology, Vol. 10, No. 2, 1997, pp. 105-112.
 M. Binato, R. R. Gurski, R. B. Fagundes, L. Meurer and M. I. Edelweiss, “p53 and Ki-67 Overexpression in Gastroesophageal Reflux Disease—Barrett’s Esophagus and Adenocarcinoma Sequence,” Diseases of the Esophagus, Vol. 22, No. 7, 2009, pp. 588-595.
 N. Rioux-Leclercq, B. Turlin, F. Sutherland, et al., “Analysis of Ki-67, p53 and Bcl-2 Expression in the Dysplasia-Carcinoma Sequence of Barrett’s Esophagus,” Oncology Reports, Vol. 6, No. 4, 1999, pp. 877-882.
 M. Feith, H. J. Stein, J. Mueller and J. R. Siewert, “Malignant Degeneration of Barrett’s Esophagus: The Role of the Ki-67 Proliferation Fraction, Expression of E-Cadherin and p53,” Diseases of the Esophagus, Vol. 17, No. 4, 2004, pp. 322-327.
 J. Jankowski, R. McMenemin, C. Yu, D. Hopwood and K. G. Wormsley, “Proliferating Cell Nuclear Antigen in Oesophageal Diseases; Correlation with Transforming Growth Factor Alpha Expression,” Gut, Vol. 33, No. 5, 1992, pp. 587-591. doi:10.1136/gut.33.5.587
 R. Kim, M. R. Clarke, M. F. Melhem, et al., “Expression of p53, PCNA, and C-erbB-2 in Barrett’s Metaplasia and Adenocarcinoma,” Digestive Diseases and Sciences, Vol. 42, No. 12, 1997, pp. 2453-2462.
 P. Gillen, M. McDermott, D. Grehan, D. O. Hourihane and T. P. Hennessy, “Proliferating Cell Nuclear Antigen in the Assessment of Barrett’s Mucosa,” British Journal of Surgery, Vol. 81, No. 12, 1994, pp. 1766-1768.
 Y. Huang, R. F. Boynton, P. L. Blount, et al., “Loss of Heterozygosity Involves Multiple Tumor Suppressor Genes in Human Esophageal Cancers,” Cancer Research, Vol. 52, No. 23, 1992, pp. 6525-6530.
 R. F. Boynton, Y. Huang, P. L. Blount, et al., “Frequent Loss of Heterozygosity at the Retinoblastoma Locus in Human Esophageal Cancer,” Cancer Research, Vol. 51, No. 20, 1991, pp. 5766-5769.
 S. H. Doak, G. J. Jenkins, E. M. Parry, et al., “Chromosome 4 Hyperploidy Represents an Early Genetic Aberration in Premalignant Barrett’s Oesophagus,” Gut, Vol. 52, No. 5, 2003, pp. 623-628. doi:10.1136/gut.52.5.623
 M. Sarbia, U. Tekin, M. Zeriouh, A. Donner and H. E. Gabbert, “Expression of the Rb Protein, Allelic Imbalance of the RB Gene and Amplification of the CDK4 Gene in Metaplasias, Dysplasias and Carcinomas in Barrett’s Oesophagus,” Anticancer Research, Vol. 21, No. 1A, 2001, pp. 387-392.
 D. Coppola, R. H. Schreiber, L. Mora, W. Dalton and R. C. Karl, “Significance of Fas and Retinoblastoma Protein Expression during the Progression of Barrett’s Metaplasia to Adenocarcinoma,” Annals of Surgical Oncology, Vol. 6, No. 3, 1999, pp. 298-304.
 R. Langer, B. H. A. Von Rahden, J. Nahrig, et al., “Prognostic Significance of Expression Patterns of cerbB-2, p53, p16 INK4A, p27KIP1, cyclin D1 and Epidermal Growth Factor Receptor in Oesophageal Adenocarcinoma: A Tissue Microarray Study,” Journal of Clinical Pathology, Vol. 59, No. 6, 2006, pp. 631-634.
 L. J. Hardie, S. J. Darnton, Y. L. Wallis, et al., “p16 Expression in Barrett’s Esophagus and Esophageal Adenocarcinoma: Association with Genetic and Epigenetic Alterations,” Cancer Letters, Vol. 217, No. 2, 2005, pp. 221-230. doi:10.1016/j.canlet.2004.06.025
 Y. S. Bian, M. C. Osterheld, C. Fontolliet, F. T. Bosman and J. Benhattar, “p16 Inactivation by Methylation of the CDKN2A Promoter Occurs Early during Neoplastic Progression in Barrett’s Esophagus,” Gastroenterology, Vol. 122, No. 4, 2002, pp. 1113-1121.
 R. Hamelin, J. F. Flejou, F. Muzeau, et al., “TP53 Gene Mutations and p53 Protein Immunoreactivity in Malignant and Premalignant Barrett’s Esophagus,” Gastroenterology, Vol. 107, No. 4, 1994, pp. 1012-1018.
 P. L. Blount, S. J. Meltzer, J. Yin, Y. Huang, M. J. Krasna and B. J. Reid, “Clonal Ordering of 17p and 5q Allelic Losses in Barrett Dysplasia and Adenocarcinoma,” Proceedings of the National Academy of Sciences of the United States of America, Vol. 90, No. 8, 1993, pp. 3221-3225.
 A. G. Casson, S. C. Evans, A. Gillis, et al., “Clinical Implications of p53 Tumor Suppressor Gene Mutation and Protein Expression in Esophageal Adenocarcinomas: Results of a Ten-Year Prospective Study,” Journal of Thoracic and Cardiovascular Surgery, Vol. 125, No. 5, 2003, pp. 1121-1131. doi:10.1067/mtc.2003.176
 M. Younes, R. M. Lebovitz, L. V. Lechago and J. Lechago, “p53 Accumulation in Barrett’s Metaplasia, Dysplasia and Carcinoma: A Follow-Up Study,” Gastroenterology, Vol. 105, No. 6, 1993, pp. 1637-1642.
 K. K. Krishnadath, H. W. Tilanus, M. van Blankenstein, F. T. Bosman and A. H. Mulder, “Accumulation of p53 Protein in Normal, Dysplastic and Neoplastic Barrett’s Esophagus,” Journal of Pathology, Vol. 175, No. 2, 1995, pp. 175-180. doi:10.1002/path.1711750204
 G. Coggi, S. Bosari, M. Roncalli, et al., “p53 Protein Accumulation and p53 Gene Mutation in Esophageal Carcinoma. A Molecular and Immunohistochemical Study with Clinicopathologic Correlations,” Cancer, Vol. 79, No. 3, 1997, pp. 425-432.
 D. Shimizu, D. Vallbohmer, H. Kuramochi, et al., “Increasing Cyclooxygenase-2 (cox-2) Gene Expression in the Progression of Barrett’s Esophagus to Adenocarcinoma Correlates with That of Bcl-2,” International Journal of Cancer, Vol. 119, No. 4, 2006, pp. 765-770.
 R. A. Soslow, H. Remotti, R. N. Baergen and N. K. Altorki, “Suppression of Apoptosis Does Not Foster Neoplastic Barrett’s Esophagus,” Modern Pathology, Vol. 12, No. 3, 1999, pp. 239-250.
 A. A. Raouf, D. A. Evoy, E. Carton, E. Mulligan, M. M. Griffin and J. V. Reynolds, “Loss of Bcl-2 Expression in Barrett’s Dysplasia and Adenocarcinoma Is Associated with Tumor Progression and Worse Survival but Not with Response to Neoadjuvant Chemoradiation,” Diseases of the Esophagus, Vol. 16, No. 1, 2003, pp. 17-23.
 P. Bhargava, G. M. Eisen, D. A. Holterman, et al., “Endoscopic Mapping and Surrogate Markers for Better Surveillance in Barrett Esophagus. A Study of 700 Biopsy Specimens,” American Journal of Clinical Pathology, Vol. 114, No. 4, 2000, pp. 552-563.
 G. Y. Lauwers, O. Kandemir, P. S. Kubilis and G. V. Scott, “Cellular Kinetics in Barrett’s Epithelium Carcinogenic Sequence: Role of Apoptosis, Bcl-2 Protein and Cellular Proliferation,” Modern Pathology, Vol. 10, No. 12, 1997, pp. 1201-1208.
 J. R. Goldblum and T. W. Rice, “Bcl-2 Protein Expression in the Barrett’s Metaplasia-Dysplasia-Carcinoma Sequence,” Modern Pathology, Vol. 8, No. 8, 1995, pp. 866-869.
 C. J. van Der Woude, P. L. Jansen, A. T. Tiebosch, et al., “Barrett’s Metaplasia-Dysplasia-Carcinoma Sequence: A Switch to a More Resistant Phenotype,” Human Pathology, Vol. 33, No. 7, 2002, pp. 686-692.
 T. A. Woodward, P. D. Klinger, P. V. Genko and J. T. Wolfe, “Barrett’s Esophagus, Apoptosis and Cell Cycle Regulation: Correlation of p53 with Bax, Bcl-2, and p21 Protein Expression,” Anticancer Research, Vol. 20, No. 4, 2000, pp. 2427-2432.
 N. Katada, R. A. Hinder, T. C. Smyrk, et al., “Apoptosis Is Inhibited Early in the Dysplasia-Carcinoma Sequence of Barrett’s Oesophagus,” Archives of Surgery, Vol. 132, No. 7, 1997, pp. 728-733.
 D. A. Bax, J. Haringsma, A. W. Einerhand, et al., “MUC4 Is Increased in High Grade Intraepithelial Neoplasia in Barrett’s Oesophagus and Is Associated with a Proapoptotic Bax to Bcl-2 Ratio,” Journal of Clinical Pathology, Vol. 57, No. 12, 2004, pp. 1267-1272.
 I. Sturm, H. Petrowsky, R. Volz, et al., “Analysis of p53/BAX/pl6ink4a/CDKN2 in Esophageal Squamous Cell Carcinoma: High BAX and p16 ink4a/CDKN2 Identifies Patients with Good Prognosis,” Journal of Clinical Oncology, Vol. 19, No. 8, 2001, pp. 2272-2281.
 A. Kurabayashi, M. Furihata, M. Matsumoto, Y. Ohtsuki, S. Sasaquri and S. Ogoshi, “Expression of Bax and Apoptosis-Related Proteins in Human Esophageal Squamous Cell Carcinoma Including Dysplasia,” Modern Pathology, Vol. 14, No. 8, 2001, pp. 741-747.
 T. Miyashita, S. Krajewski, M. Krajewska, et al., “Tumor Suppressor p53 Is a Regulator of Bcl-2 and Bax Gene Expression in Vitro and in Vivo,” Oncogene, Vol. 9, No. 6, 1994, pp. 1799-1805