Loss of heterozygosity and microsatellite instability as predictive markers among Iranian esophageal cancer patients

Document Type : Original Article


1 Cellular and Molecular Research Center, Sabzevar University of Medical Sciences, Sabzevar, Iran

2 Division of Human Genetics, Immunology Research Center, Avicenna Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran

3 School of Systems Biology, George Mason University, Manassas, Virginia, U.S.A

4 Department of hematopathology, Central Laboratory, Imam Reza Hospital, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran

5 Cancer Molecular Pathology Research Center, Imam Reza Hospital, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran

6 Medical Genetics Research Center, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran


Objective(s): Variation in microsatellite sequences that are dispersed in the genome has been linked to a deficiency in cellular mismatch repair system and defects in several genes of this system are involved in carcinogenesis. Our aim in this study was to illustrate microsatellite DNA alteration in esophageal cancer.
Materials and Methods: DNA was extracted from formalin fixed paraffin embedded (FFPE) tissues from surgical and matched margin-normal samples. Microsatellite instability (MSI) and loss of heterozygosity (LOH) were studied in 50 cases of esophageal squamous cell carcinoma (ESCC) by amplifying six microsatellite markers: D13S260 (13q12.3), D13S267 (13q12.3), D9S171 (9p21), D2S123 (2p), D5S2501 (5q21) and TP53 (17p13.1) analyzed on 6% denaturing polyacrylamide gel electrophoresis.
Results: Statistical analysis indicated a near significant reverse correlation between grade and LOH (P= 0.068, correlation coefficient= -0.272). Specifically, increased LOH in tumor DNA has a significant correlation with increased differentiation from poorly differentiated to well differentiated tumors (P= 0.002 and P= 0.016 respectively). In addition, higher number of chromosomal loci with LOH showed a reverse correlation with lymph node metastasis (P= 0.026, correlation coefficient= -0.485). Furthermore, there was a positive correlation between addiction and MSI (P= 0.026, correlation coefficient= 0.465).
Conclusion: Microsatellite DNA alterations may be a prognostic tool for detection and the evolution of prognosis in patients with SCC of esophagus. It can be concluded that regional lymph node metastasis would be less likely with increased heterozygote loci and addiction with any of opium, cigarette, water pipe or alcohol can be a susceptibility factor(s) for MSI.


1. Ferlay J, Soerjomataram I, Dikshit R, Eser S, Mathers C, Rebelo M, et al. Cancer incidence and mortality worldwide: sources, methods and major patterns in GLOBOCAN 2012. Int J Cancer 2015; 136:E359-386.
2. Forghanifard MM, Taleb Sh, Abbaszadegan MR. Notch signaling target genes are directly correlated to esophageal squamous cell carcinoma tumorigenesis. Pathol Oncol Res 2015; 21:463-467.
3. Harirchi I, Kolahdoozan S, Hajizadeh S, Safari F, Sedighi Z, Nahvijou A, et al. Esophageal cancer in Iran; a population-based study regarding adequacy of cancer surgery and overall survival. Eur J Surg Oncol 2014; 40:352-357.
4. Sadjadi A, Nouraie M, Mohagheghi MA, Mousavi-Jarrahi A, Malekezadeh R, Parkin DM. Cancer occurrence in Iran in 2002, an international perspective. Asian Pac J Cancer Prev 2005; 6:359-363.
5. Forghanifard MM, Gholamin M, Moaven O, Farshchian M, Ghahraman M, Aledavood A, et al. Neoantigen in esophageal squamous cell carcinoma for dendritic cell-based cancer vaccine development. Med Oncol 2014; 31:191.
6.  An JY, Fan ZM, Gao SS, Zhuang ZH, Qin YR, Li JL, et al. Loss of heterozygosity in multistage carcinogenesis of esophageal carcinoma at high-incidence area in Henan Province, China. World J Gastroenterol 2005; 11:2055-2060.
7. Hayden JD, Martin IG, Cawkwell L, Quirke P. The role of microsatellite instability in gastric carcinoma. Gut 1998; 42:300-303.
8. Hung CM, Yu AY, Lai YT, Shaner PJ. Developing informative microsatellite makers for non-model species using reference mapping against a model species' genome. Sci Rep 2016; 6:23087.
9. Musulen E, Moreno V, Reyes G, Sancho FJ, Peinado MA, Esteller M, et al. Standardized approach for microsatellite instability detection in gastric carcinomas. Hum Pathol 2004; 35:335-342.
10. Sturzeneker R, Bevilacqua RA, Haddad LA, Simpson AJ, Pena SD. Microsatellite instability in tumors as a model to study the process of microsatellite mutations. Hum Mol Genet 2000; 9:347-352.
11. Eisenberger CF, Knoefel WT, Peiper M, Merkert P, Yekebas EF, Scheunemann P, et al. Squamous cell carcinoma of the esophagus can be detected by microsatellite analysis in tumor and serum. Clin Cancer Res 2003; 9:4178-4183.
12. Matsumoto Y, Nagasaka T, Kambara T, Hoshizima N, Murakami J, Sasamoto H, et al. Microsatellite instability and clinicopathological features in esophageal squamous cell cancer. Oncol Rep 2007; 18:1123-1127.
13. Attaran-Bandarabadi F, Ziaee AA, Yazdanbod M, Shahpanah M, Setayeshgar A, Nassiri M. Loss of heterozygosity on chromosome 5 in Iranian esophageal cancer patients. Genet Mol Res 2011; 10:2316-2325.
14. Cai YC, So CK, Nie AY, Song Y, Yang GY, Wang LD, et al. Characterization of genetic alteration patterns in human esophageal squamous cell carcinoma using selected microsatellite markers spanning multiple loci. Int J Oncol 2007; 30:1059-1067.
15. Liu FX, Huang XP, Xu X, Cai Y, Han YL, Wu RL, et al. Alterations of MLH1 and microsatellite instability in esophageal squamous cell carcinomas. Yi Chuan Xue Bao 2005; 32:234-242.
16. Uchida A, Tachibana M, Miyakawa A, Nakamura K, Murai M. Microsatellite analysis in multiple chromosomal regions as a prognostic indicator of primary bladder cancer. Urol Res 2000; 28:297-303.
17. Grundei T, Vogelsang H, Ott K, Mueller J, Scholz M, Becker K, et al. Loss of heterozygosity and microsatellite instability as predictive markers for neoadjuvant treatment in gastric carcinoma. Clin Cancer Res 2000; 6:4782-4788.
18. Oda S, Oki E, Maehara Y, Sugimachi K. Precise assessment of microsatellite instability using high resolution fluorescent microsatellite analysis. Nucleic Acids Res 1997; 25:3415-3420.
19. Liu M, Zhang F, Liu S, Zhao W, Zhu J, Zhang X. Loss of heterozygosity analysis of microsatellites on multiple chromosome regions in dysplasia and squamous cell carcinoma of the esophagus. Exp Ther Med 2011; 2:997-1001.
20. Eisenberger CF, Schoenberg M, Enger C, Hortopan S, Shah S, Chow NH, et al. Diagnosis of renal cancer by molecular urinalysis. J Natl Cancer Inst 1999; 91:2028-2032.
21. Boland CR, Thibodeau SN, Hamilton SR, Sidransky D, Eshleman JR, Burt RW, et al. A National Cancer Institute Workshop on Microsatellite Instability for cancer detection and familial predisposition: development of international criteria for the determination of microsatellite instability in colorectal cancer. Cancer Res 1998; 58:5248-5257.
22. Kagawa Y, Yoshida K, Hirai T, Toge T, Yokozaki H, Yasui W, et al. Microsatellite instability in squamous cell carcinomas and dysplasias of the esophagus. Anticancer Res 2000; 20:213-217.
23. Sugai T, Uesugi N, Habano W, Suzuki K. [Microsatellite instability]. Nihon Rinsho 2011; 69:84-93.
24. Meltzer SJ, Yin J, Manin B, Rhyu MG, Cottrell J, Hudson E, et al. Microsatellite instability occurs frequently and in both diploid and aneuploid cell populations of Barrett's-associated esophageal adenocarcinomas. Cancer Res 1994; 54:3379-3382.
25. Muzeau F, Flejou JF, Belghiti J, Thomas G, Hamelin R. Infrequent microsatellite instability in oesophageal cancers. Br J Cancer 1997; 75:1336-1339.
26. Mansouri A, Foroughmand AM, Abbaszadegan MR, Memar B, Mahmoudian RA, Gholamin M. Expression analysis of CD44 isoforms S and V3, in patients with esophageal squamous cell carcinoma. Iran J Basic Med Sci 2015; 18:380-384.
27. Mironov NM, Aguelon AM, Hollams E, Lozano JC, Yamasaki H. Microsatellite alterations in human and rat esophageal tumors at selective loci. Mol Carcinog 1995; 13:1-5.
28. Mathew R, Arora S, Mathur M, Chattopadhyay TK, Ralhan R. Esophageal squamous cell carcinomas with DNA replication errors (RER+) are associated with p16/pRb loss and wild-type p53. J Cancer Res Clin Oncol 2001; 127:603-612.
29. Diaz LA Jr. The current clinical value of genomic instability. Semin Cancer Biol 2005; 15:67-71.
30. Lynch HT, Kaul K. Microsatellite instability, clinical implications, and new methodologies. J Natl Cancer Inst 2000; 92:511-512.
31. Li G, Hu N, Goldstein AM, Tang ZZ, Roth MJ, Wang QH, et al. Allelic loss on chromosome bands 13q11-q13 in esophageal squamous cell carcinoma. Genes Chromosomes Cancer 2001; 31:390-397.
32. Tarmin L, Yin J, Zhou X, Suzuki H, Jiang HY, Rhyu MG, et al. Frequent loss of heterozygosity on chromosome 9 in adenocarcinoma and squamous cell carcinoma of the esophagus. Cancer Res 1994; 54:6094-6096.
33. Xing EP, Nie Y, Wang LD, Yang GY, Yang CS. Aberrant methylation of p16INK4a and deletion of p15INK4b are frequent events in human esophageal cancer in Linxian, China. Carcinogenesis 1999; 20:77-84.
34. Taghavi N, Biramijamal F, Sotoudeh M, Khademi H, Malekzadeh R, Moaven O, et al. p16INK4a hypermethylation and p53, p16 and MDM2 protein expression in esophageal squamous cell carcinoma. BMC Cancer 2010; 10:138.
35. Lu N, Hu N, Li WJ, Roth MJ, Wang C, Su H, et al. Microsatellite alterations in esophageal dysplasia and squamous cell carcinoma from laser capture microdissected endoscopic biopsies. Cancer Lett 2003; 189:137-145.
36. Ikeguchi M, Unate H, Maeta M, Kaibara N. Detection of loss of heterozygosityat microsatellite loci in esophageal squamous-cell carcinoma. Oncology 1999; 56:164-168.
37. Liu M, Zeng HC, Zhang XL, Zhao W, Zhu J, Huang  JF, et al. [Loss of heterozygosity analysis of microsatellites on multiple chromosome regions in dysplasia and squamous cell carcinoma of esophagus]. Zhonghua Wai Ke Za Zhi 2008; 46:1337-1339.
38. Lichun Y, Ching Tang CM, Wai Lau K, Lung ML. Frequent loss of heterozygosity on chromosome 9 in Chinese esophageal squamous cell carcinomas. Cancer Lett 2004; 203:71-77.
39. Yamano M, Fujii H, Takagaki T, Kadowaki N, Watanabe H, Shirai T. Genetic progression and divergence in pancreatic carcinoma. Am J Pathol 2000; 156:2123-2133.
40. Liu M, Zhang F, Liu S, Zhao W, Zhu J, Zhang X. Microsatellite analysis in multistage carcinogenesis of esophageal squamous cell carcinoma from Chongqing in Southern China. Int J Mol Sci 2011; 12:7401-7409.
41. Hayashi M, Tamura G, Jin Z, Kato I, Sato M, Shibuya Y, et al. Microsatellite instability in esophageal squamous cell carcinoma is not associated with hMLH1 promoter hypermethylation. Pathol Int 2003; 53:270-276.
42. Shimada M, Horii A, Sasaki S, Yanagisawa A, Kato Y, Yamashita K, et al. Infrequent replication errors at microsatellite loci in tumors of patients with multiple primary cancers of the esophagus and various other tissues. Jpn J Cancer Res 1995; 86:511-515.
43. Moghbeli M, Moaven O, Dadkhah E, Farzadnia M, Roshan NM, Asadzadeh-Aghdaee H, et al. High frequency of microsatellite instability in sporadic colorectal cancer patients in Iran. Genet Mol Res 2011; 10:3520-3529.