Document Type : Original Research

Authors

1 Department of Bacteriology & Virology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran

2 Department of Pathology, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran

3 Gastroenterohepathology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran

Abstract

Background & Objective: Epstein-Barr virus nuclear antigen-1 (EBNA1) is one of the most important proteins of Epstein-Barr virus (EBV) that might be mutated in various related cancers. The purpose of this study was to compare EBNA1 mutations in the C-terminal region between patients with cervical and ovarian cancer and healthy individuals.
Methods: As test and control groups, 18 EBV-positive paraffin-embedded samples of cervical and ovarian cancer and 10 age- and gender-matched healthy volunteers who did not have cancer but were EBV-positive were both used. Utilizing a commercial DNA extraction kit, total DNA was extracted following deparaffinization. The entire C-terminal region of the EBNA1 sequence was amplified using an in-house nested PCR. Phylogenetic analysis and Sanger sequencing were used to analyze the sequences using MEGA 7 software and through NJ method.
Results: Sequence analysis revealed that the P-Ala subtype of EBNA1 was present in all samples. In two and one samples, respectively, of cervical cancer patients, the mutations A1887G and G1891A were found. The G1595T mutation was also detected in four sequences taken from ovarian cancer patients. No statistically significant difference could be found between the frequency of mutations in patients and controls (P>0.05). No known amino acid substitutions were found in the USP7-binding region and the DBD/DD domain.
Conclusion: The findings showed that P-Ala is the predominant EBV subtype across all samples. Additionally, as the sequence of EBNA1's C-terminal region is so stable, it's possible that it had little impact on the pathogenesis of ovarian and cervical malignancies. It is advised to conduct additional research to verify these findings.

Keywords

Main Subjects

  1. Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, et al. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries. CA Cancer J Clin. 2021;71(3):209-49. [DOI:10.3322/caac.21660] [PMID]
  2. Ferrall L, Lin KY, Roden RBS, Hung CF, Wu TC. Cervical Cancer Immunotherapy: Facts and Hopes. Clin Cancer Res. 2021;27(18):4953-73. [DOI:10.1158/1078-0432.CCR-20-2833] [PMID] [PMCID]
  3. Castle PE, Einstein MH, Sahasrabuddhe VV. Cervical cancer prevention and control in women living with human immunodeficiency virus. CA Cancer J Clin. 2021;71(6):505-26. [DOI:10.3322/caac.21696] [PMID]
  4. Momenimovahed Z, Salehiniya H. Cervical cancer in Iran: integrative insights of epidemiological analysis. BioMedicine. 2018;8(3):18. [PMID] [DOI:10.1051/bmdcn/2018080318] [PMCID]
  5. Momenimovahed Z, Salehiniya H. Cervical cancer in Iran: integrative insights of epidemiological analysis. BioMedicine. 2018;8(3). [PMID] [DOI:10.1051/bmdcn/2018080318] [PMCID]
  6. Sipos A, Ujlaki G, Miko E, Maka E, Szabo J, Uray K, et al. The role of the microbiome in ovarian cancer: mechanistic insights into oncobiosis and to bacterial metabolite signaling. Mol Med. 2021;27(1):33. [DOI:10.1186/s10020-021-00295-2] [PMID] [PMCID]
  7. Pokhriyal R, Hariprasad R, Kumar L, Hariprasad G. Chemotherapy Resistance in Advanced Ovarian Cancer Patients. Biomark Cancer. 2019;11:1179299X19860815. [PMID] [PMCID] [DOI:10.1177/1179299X19860815]
  8. Morales-Sanchez A, Fuentes-Panana EM. Human viruses and cancer. Viruses. 2014;6(10):4047-79. [DOI:10.3390/v6104047] [PMID] [PMCID]
  9. Shi Y, Peng SL, Yang LF, Chen X, Tao YG, Cao Y. Co-infection of Epstein-Barr virus and human papillomavirus in human tumorigenesis. Chin J Cancer. 2016;35(1):16. [DOI:10.1186/s40880-016-0079-1] [PMID] [PMCID]
  10. Gupta I, Nasrallah GK, Sharma A, Jabeen A, Smatti MK, Al-Thawadi HA, et al. Co-prevalence of human Papillomaviruses (HPV) and Epstein-Barr virus (EBV) in healthy blood donors from diverse nationalities in Qatar. Cancer Cell Int. 2020;20(1):1-9. [DOI:10.1186/s12935-020-01190-2] [PMID] [PMCID]
  11. Andrei G, Trompet E, Snoeck R. Novel Therapeutics for Epstein(-)Barr Virus. Molecules. 2019;24(5). [DOI:10.3390/molecules24050997] [PMID] [PMCID]
  12. Bedri S, Sultan AA, Alkhalaf M, Al Moustafa AE, Vranic S. Epstein-Barr virus (EBV) status in colorectal cancer: a mini review. Hum Vaccin Immunother. 2019;15(3):603-10. [PMID] [PMCID] [DOI:10.1080/21645515.2018.1543525]
  13. Echelman D, Feldman S. Management of cervical precancers: a global perspective. Hematol Oncol Clin North Am. 2012;26(1):31-44. [DOI:10.1016/j.hoc.2011.11.005] [PMID]
  14. Vranic S, Cyprian FS, Akhtar S, Al Moustafa AE. The Role of Epstein-Barr Virus in Cervical Cancer: A Brief Update. Frontiers in oncology. 2018;8:113. [DOI:10.3389/fonc.2018.00113] [PMID] [PMCID]
  15. Shokouh MR, Safaei A, Moattari A, Sarvari J. Association of Human Papilloma Virus and Epstein-Barr Virus with Ovarian Cancer in Shiraz, Southwestern Iran. Iran J Pathol. 2020;15(4):292-8. [DOI:10.30699/ijp.2020.119681.2306] [PMID] [PMCID]
  16. Imai S, Nishikawa J, Takada K. Cell-to-cell contact as an efficient mode of Epstein-Barr virus infection of diverse human epithelial cells. J Virol. 1998;72(5):4371-8. [DOI:10.1128/JVI.72.5.4371-4378.1998] [PMID] [PMCID]
  17. Shinozaki-Ushiku A, Kunita A, Fukayama M. Update on Epstein-Barr virus and gastric cancer (review). Int J Oncol. 2015;46(4):1421-34. [DOI:10.3892/ijo.2015.2856] [PMID]
  18. Manivannan AC, Devaraju V, Velmurugan P, Sathiamoorthi T, Sivakumar S, Subbiah SK, et al. Tumorigenesis and diagnostic practice applied in two oncogenic viruses: Epstein Barr virus and T-cell lymphotropic virus-1-Mini review. Biomed Pharmacother. 2021;142:111974. [DOI:10.1016/j.biopha.2021.111974] [PMID]
  19. Khenchouche A, Sadouki N, Boudriche A, Houali K, Graba A, Ooka T, et al. Human papillomavirus and Epstein-Barr virus co-infection in cervical carcinoma in Algerian women. Virol J. 2013;10:340. [DOI:10.1186/1743-422X-10-340] [PMID] [PMCID]
  20. Frappier L. The Epstein-Barr Virus EBNA1 Protein. Scientifica. 2012;2012:438204. [DOI:10.6064/2012/438204] [PMID] [PMCID]
  21. Lu J, Murakami M, Verma SC, Cai Q, Haldar S, Kaul R, et al. Epstein-Barr Virus nuclear antigen 1 (EBNA1) confers resistance to apoptosis in EBV-positive B-lymphoma cells through up-regulation of survivin. Virology. 2011;410(1):64-75. [PMID] [DOI:10.1016/j.virol.2010.10.029] [PMCID]
  22. Yates JL, Warren N, Sugden B. Stable replication of plasmids derived from Epstein-Barr virus in various mammalian cells. Nature. 1985;313 (6005):812-5. [DOI:10.1038/313812a0] [PMID]
  23. Frappier L. Viral disruption of promyelocytic leukemia (PML) nuclear bodies by hijacking host PML regulators. Virulence. 2011;2(1):58-62. [DOI:10.4161/viru.2.1.14610] [PMID]
  24. Kahla S, Oueslati S, Achour M, Kochbati L, Chanoufi MB, Maalej M, et al. Correlation between ebv co-infection and HPV16 genome integrity in Tunisian cervical cancer patients. Braz J Microbiol. 2012;43(2):744-53. [PMID] [DOI:10.1590/S1517-83822012000200039] [PMCID]
  25. Bhattacharya S, Chakraborty D, Basu M, Ghosh MK. Emerging insights into HAUSP (USP7) in physiology, cancer and other diseases. Signal Transduct Target Ther. 2018;3(1):17. [DOI:10.1038/s41392-018-0012-y] [PMID] [PMCID]
  26. Sun L, Zhao Z, Liu S, Liu X, Sun Z, Luo B. Sequence Variation Analysis of Epstein-Barr Virus Nuclear Antigen 1 Gene in the Virus Associated Lymphomas of Northern China. PloS one. 2015;10(10):e0140529. [PMID] [PMCID] [DOI:10.1371/journal.pone.0140529]
  27. Mai SJ, Ooka T, Li DJ, Zeng MS, Jiang RC, Yu XJ, et al. Functional advantage of NPC-related V-val subtype of Epstein-Barr virus nuclear antigen 1 compared with prototype in epithelial cell line. Oncol Rep. 2007;17(1):141-6. [DOI:10.3892/or.17.1.141] [PMID]
  28. Mai SJ, Xie D, Huang YF, Wang FW, Liao YJ, Deng HX, et al. The enhanced transcriptional activity of the V-val subtype of Epstein-Barr virus nuclear antigen 1 in epithelial cell lines. Oncol Rep. 2010;23(5):1417-24. [DOI:10.3892/or_00000779]
  29. Mai SJ, Li DJ, Zhou XX, Chen LZ, Feng QS, Zhang RH, et al. [Functions of V-val subtype of Epstein-Barr nuclear antigen 1]. Ai zheng = Aizheng = Chinese journal of cancer. 2008;27(3):231-6.
  30. Dheekollu J, Malecka K, Wiedmer A, Delecluse HJ, Chiang AK, Altieri DC, et al. Carcinoma-risk variant of EBNA1 deregulates Epstein-Barr Virus episomal latency. Oncotarget. 2017;8(5):7248-64. [DOI:10.18632/oncotarget.14540] [PMID] [PMCID]
  31. Joharinia N, Faghihinejad S, Seyedi K, Farhadi A, Hosseini SY, Safaei A, et al. Co-existing of HSV1/2 or EBV Infection with the Presence of High-Risk HPV DNA in Cervical Lesions in the Southwest of Iran. Asian Pac J Cancer Prev. 2020;21(5):1459-64. [PMID] [PMCID] [DOI:10.31557/APJCP.2020.21.5.1459]
  32. Mahmoudvand S, Zamani K, Safaei A, Khashei R, Motamedifar M, Azizi Z, et al. No Detection of Streptococcus gallolyticus and Helicobacter pylori in Colorectal Cancer Tissue Samples in Shiraz, Iran. Iranian Journal of Cancer Prevention. 2017;In Press(In Press):e6337. [DOI:10.17795/ijcp-6337]
  33. Dowran R, Joharinia N, Safaei A, Bakhtiyarizadeh S, Alidadi Soleimani A, Alizadeh R, et al. No detection of EBV, BKV and JCV in breast cancer tissue samples in Iran. BMC Res Notes. 2019;12(1):171. [DOI:10.1186/s13104-019-4178-3] [PMID] [PMCID]
  34. Sarvari J, Mahmoudvand S, Pirbonyeh N, Safaei A, Hosseini SY. The very low frequency of Epstein-Barr JC and BK Viruses DNA in colorectal cancer tissues in Shiraz, Southwest Iran. Pol J Microbiol. 2018;67(1):73-9. [DOI:10.5604/01.3001.0011.6146] [PMID]
  35. Kwok H, Tong AH, Lin CH, Lok S, Farrell PJ, Kwong DL, et al. Genomic sequencing and comparative analysis of Epstein-Barr virus genome isolated from primary nasopharyngeal carcinoma biopsy. PloS one. 2012;7(5):e36939. [DOI:10.1371/journal.pone.0036939] [PMID] [PMCID]
  36. Karbalaie Niya MH, Safarnezhad Tameshkel F, Keyvani H, Esghaei M, Panahi M, Zamani F, et al. Epstein-Barr virus molecular epidemiology and variants identification in head and neck squamous cell carcinoma. Eur J Cancer Prev. 2020;29 (6):523-30. [DOI:10.1097/CEJ.0000000000000554] [PMID]
  37. Littman AJ, Rossing MA, Madeleine MM, Tang M-TC, Yasui Y. Association between late age at infectious mononucleosis, Epstein-Barr virus antibodies, and ovarian cancer risk. Scand J Infect Dis. 2003;35(10):728-35. [DOI:10.1080/00365540310016556] [PMID]
  38. Borozan I, Zapatka M, Frappier L, Ferretti V. Analysis of Epstein-Barr Virus Genomes and Expression Profiles in Gastric Adenocarcinoma. J Virol. 2018;92(2). [DOI:10.1128/JVI.01239-17] [PMID] [PMCID]
  39. Gaal K, Weiss LM, Chen WG, Chen YY, Arber DA. Epstein-Barr virus nuclear antigen (EBNA)-1 carboxy-terminal and EBNA-4 sequence polymorphisms in nasal natural killer/T-cell lymphoma in the United States. Lab Invest. 2002;82(7):957-62. [PMID] [DOI:10.1097/01.LAB.0000020416.66825.A0]
  40. Wang Y, Liu X, Xing X, Cui Y, Zhao C, Luo B. Variations of Epstein-Barr virus nuclear antigen 1 gene in gastric carcinomas and nasopharyngeal carcinomas from Northern China. Virus Res. 2010;147(2):258-64. [DOI:10.1016/j.virusres.2009.11.010] [PMID]
  41. Chen MR, Tsai CH, Wu FF, Kan SH, Yang CS, Chen JY. The major immunogenic epitopes of Epstein-Barr virus (EBV) nuclear antigen 1 are encoded by sequence domains which vary among nasopharyngeal carcinoma biopsies and EBV-associated cell lines. J Gen Virol. 1999;80 ( Pt 2):447-55. [DOI:10.1099/0022-1317-80-2-447] [PMID]
  42. Wrightham MN, Stewart JP, Janjua NJ, Pepper SD, Sample C, Rooney CM, et al. Antigenic and sequence variation in the C-terminal unique domain of the Epstein-Barr virus nuclear antigen EBNA-1. Virology. 1995;208(2):521-30. [DOI:10.1006/viro.1995.1183] [PMID]