Loss of P16 Protein Expression and Its Association with Epstein-Barr Virus LMP-1 Expression in Hodgkin's Lymphoma

Fawzi Irshaid Irshaid--- Dept. of Biological Sciences, Faculty of Science, Al al-Bayt University, Al-Mafraq, Jordan,
Khaled Tarawneh--- . Dept. of Biological Sciences, Faculty of Science, Mutah University, Mutah, Jordan,
Aisha Alshdefat--- Mafraq Hospital of Children and Maternity, Al-Mafraq, Jordan,
Fatiha Dilmi--- Dept. of Biological Sciences, Faculty of Science, Al al-Bayt University, Al-Mafraq, Jordan,
Adnan Jaran--- Dept. of Biological Sciences, Faculty of Science, Al al-Bayt University, Al-Mafraq, Jordan,
Raji Al-Hadithi--- Dept. of Pathology, Faculty of Medicine, Jordan University of Sciences and Technology, Irbid, Jordan,
Ahad Al-Khatib--- Dept. of Pathology, Faculty of Medicine, Jordan University of Sciences and Technology, Irbid, Jordan



Background: Expression of Epstein-Barr virus Latent Member Protein-1 (EBV LMP-1) and loss of P16 protein expression are documented in lymphoma, indicating a relationship between them, but this relationship is not clear and sometimes contradictory. Thus, this study was conducted to examine the relationship between the loss of P16 and EBV LMP-1 expression in Jordanian patients diagnosed with lymphoma.

Methods: Sections were made from archival formalin-fixed and paraffin-embedded blocks from 55 patients diagnosed with lymphoma. P16 expression and LMP-1 expression were detected by immunohistochemistry using monoclonal antibodies.

Results: In Hodgkin's Lymphoma (HL), the loss of P16 was higher in LMP-1 positive cases (61%) than LMP-1 negative cases (25%; P = 0.072). Conversely, in Non-Hodgkin's Lymphoma (NHL), none of LMP-1 positive samples showed loss of P16. Furthermore, among LMP-1 HL positive cases, the loss of P16 was more frequent in male (75%) than female (33%). Also, there was a significantly higher proportion of LMP-1 positive cases showing loss of P16 in HL (11:18), compared to those in NHL (0:8, P < 0.001), confirming a difference between HL and NHL, concerning the LMP-1/P16 relationship.

Conclusion: A trend for an association between loss of P16 and LMP-1 expression was observed in HL but not NHL patients. These findings suggest that there are molecular and clinical differences in the pathogenesis and development of different subtypes of lymphoma.

Please cite this article as: Irshaid F, Tarawneh Kh, Alshdefat A, Dilmi F, Jaran A, Al-HadithiR, Al-Khatib A. Loss of P16 Protein Expression and Its Association with Epstein-Barr Virus LMP-1 Expression in Hodgkin's Lymphoma. Iran J Cancer Prev. 2013; 6(2):78-84.


1. Jaffe ES, Harris NL, Stein H, Isaacson PG. Classification of lymphoid neoplasms: the microscope as a tool for disease discovery. Blood. 2008; 112:4384-99.

2. Piccaluga PP, Agostinelli C, Gazzola A, Tripodo C, Bacci F, Sabattini E, et al. Pathobiology of Hodgkin Lymphoma. Advances in Hematology. 2011; 2011:18.

3. Jordanian National Cancer Registry, Cancer incidence in Jordan, Report of Ministry of Health, pp. 47-49; 2004. http://www.khcc.jo/National-cancer-statistics.aspx.

4. Cohen, JI. Epstein–Barr virus infection. N Engl J Med. 2000; 17:481-92.

5. Bechtel D, Kurth J, Unkel C, Kuppers R. Transformation of BCR-deficient germinal-center B cells by EBV supports a major role of the virus in the pathogenesis of Hodgkin and post transplantation lymphomas. Blood. 2005; 106:4345-50.

6. Kapatai G, Murray P. Contribution of the Epstein Barr virus to the molecular pathogenesis of Hodgkin lymphoma. J Clin Pathol. 2007; 60:1342-9.

7. Kamranvar SA, Gruhne B, Szeles A, Masucci MG. Epstein–Barr virus promotes genomic instability in Burkett's lymphoma. Oncogene. 2007; 26:5115-23.

8. Dessy E, Rossi E, Berenzi A, Tironi A, Benett A, Grigolato P. Chromosome 9 instability and alterations of p16 gene in squamous cell carcinoma of the lung and in adjacent normal bronchi: FISH and immunohistochemical study. Histopathology Journal. 2008; 52:475-82.

9. Serrano M, Lee H, Chin L, Cordon-Cardo C, Beach D, DePinho RA. Role of the INK4a locus in tumor suppression and cell mortality. Cell. 1996; 85:27-37.

10. Hayslip J, Montero A. Tumor suppressor gene methylation in follicular lymphoma: a comprehensive review. Molecular Cancer. 2006; 5: 3-7.

11. Chebel A, Chien WW, Gerland LM, Mekki Y, Bertrand Y. Does p16ink4a expression increase with the number of cell doublings in normal and malignant lymphocytes? Leuk Res. 2007; 31: 1649-58.

12. Yang X, He Z, Xin B, Cao L. LMP1 of Epstein-Barr virus suppresses cellular senescence associated with the inhibition of p16INK4a expression. Oncogene. 2000; 19: 2002-13.

13. Ohtani N, Brennan P, Gaubatz S, Sanij E, Hertzog P, Wolvetang E, et al. Epstein-Barr virus LMP1 blocks p16-RB pathway by promoting nuclear export of E2F4/5. J Cell Biol. 2003; 162:173-83.

14. Kim LH, Peh SC, Poppema S. Expression of retinoblastoma protein and p16 proteins in classic Hodgkin's lymphoma: relationship with expression of p53 and presence of Epstein-Barr virus in the regulation of cell growth and death. Journal of Human Pathology. 2006; 37:92-100.

15. Tomkinson B, Robertson E, Kieff E. Epstein-Barr virus nuclear proteins EBNA-3A and EBNA-3C are essential for B-lymphocyte growth transformation. J Virol. 1993; 67: 2014-25.

16. Irshaid F, Jaran A, Dilmi F, Tarawneh K, Hadeth R, Al-Khatib A. Prevalence of Epstein Barr virus latent membrane protein-1 in Jordanian Patients with Hodgkin's lymphoma and non-Hodgkin's lymphoma. J Biol Sci. 2010; 10:507-13.

17. Young LS, Dawson CW, Eliopoulos AG. The expression and function of Epstein-Barr virus encoded latent genes. J Clin Pathol Mol Pathol. 2000; 53:238-47.

18. Zhao P, Lu Y, Liu L, Zhong M. Aberrant expression of ID2 protein and its correlation with EBV-LMP1 and p16 (INK4A) in Classical Hodgkin Lymphoma in China. BioMed Central Journal. 2008; 8:1471-2407.

19. Skalska L, White RE, Franz M, Ruhmann M, Allday MJ. Epigenetic repression of p16INK4A by Latent Epstein-Barr Virus requires the interaction of EBNA3A and EBNA3C with CtBP. PLos Pathog. 2010; 6: e1000951.

20. Irshaid F, Dilmi F, Tarawneh K, Hadeth R, Jaran A, Al-Khatib A. Loss of P16/INK4A protein expression is a common abnormality in Hodgkin’s lymphoma. International Journal of Biological and Life Sciences. 2009; 5:215-220.

21. Villuendas R, Sanchez-Beato M, Martinez JC, Saez AI, Martinez-Delgado B, Garcia JF, et al. Loss of p16/INK4a protein expression in non-Hodgkin’s lymphoma is a frequent finding associated with tumor progression. Am J Pathol. 1998; 153:887–97.

22. Klangby U, Okan I, Magnusson KP, Wendland M, Lind P, Wiman KG. P16/INK4a and p15/INK4b gene methylation and absence of p16/INK4a mRNA and protein expression in Burkett's lymphoma. Blood. 1998; 91:1680-7.

23. Pinyol M, Cobo F, Bea S, Jares P, Nayach I, Fernandez PL, et al. P16 (INK4a) gene inactivation by deletions, mutations, and hypermethylation is associated with transformed and aggressive variants of non-Hodgkin’s lymphomas. Blood. 1998; 91:2977-84.

24. Lee HS, Chang MS, Yang HK, Lee BL, Kim WH. Epstein-Barr Virus-positive gastric carcinoma has a distinct protein expression profile in comparison with Epstein-Barr Virus-negative carcinoma. Clin Cancer Res. 2004; 10:1698-1705.

25. Kang S, Kim J, Kim HB, Shim JW, Nam E, Kim SH, et al. Methylation of p16INK4a is a non-rare event in cervical intraepithelial neoplasia. Diagn Mol Pathol. 2006; 15:74-82.

26. Wang P, Zhang Q, Yang JF, Cheng ZN, Zhang K, Yu DH. Epstein-Barr virus infection and p16(INK4a) over expression in gastric adenocarcinoma. Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi. 2008; 22:244-6.

27. Nehls K, Vinokurova S, Schmidt D, Kommoss F, Reuschenbach M, Kisseljov F, et al.. P16 methylation does not affect protein expression in cervical carcinogenesis. Eur J Cancer. 2008; 44:2496-505.


Epstein-Barr virus infections; Immunohistochemistry; Lymphoma; Monoclonal antibodies

Full Text: