Pathology & Oncology Research

, Volume 21, Issue 1, pp 157–166

Human Papilloma Virus Early Proteins E6 (HPV16/18-E6) and the Cell Cycle Marker P16 (INK4a) are Useful Prognostic Markers in Uterine Cervical Carcinomas in Qassim Region- Saudi Arabia



Cervical cancer is a common and an important public health problem for adult women in developing countries. In contrast, cervical cancer incidence is low in Saudi Arabia. High-risk types of human papilloma viruses (HPV16 and HPV18) are the most significant risk factors for cervical cancer. HPV16/18-E6 oncoprotein is associated with HPV etiology, viral persistence and epithelial transformation. Cell cycle protein p16 INK4a (p16) plays an important role in the pathophysiology of cervical carcinomas. The aims of this study were to investigate the expression of HPV16/18-E6 and p16 in uterine cervical carcinomas in Qassim Region - Saudi Arabia, and to relate the results to the established clinicopathological prognostic parameters (age of the patient, educational level, birth control methods, number of pregnancy, smoking status, degree of histological differentiation, clinical stage, and lymph node metastasis) The study included 40 specimens of uterine cervical squamous cell carcinomas diagnosed and confirmed by biopsy. Histopathological classification of cervical tumors cases was performed according to the International Federation of Gynecology and Obstetrics (FIGO). Immunohistochemical analysis for HPV16/18-E6 and p16 were carried out on formalin-fixed paraffin-embedded sections of cervical tissues using avidin-biotin peroxidase method. There was a significant statistical correlation between HPV16/18-E6 expression in cervical carcinoma and nationality, smoking status and size of the tumor. HPV16/18-E6 oncoprotein expression in normal lymphocytes and endothelial cells in the tumor tissues and the adjacent normal cervical tissues suggest the possibility that HPV infection might spread to other organs through blood circulation. P16 expression has been correlated with high grade, stage of cervical SCC and HPV16/18-E6 expression. The current study supports the critical function of p16 and HPV16/18-E6 as specific markers for cervical carcinoma. However the potential for usage of p16 and HPV16/18-E6 as prognostic markers will require detailed follow data for a larger group of patients.


Immunohistochemistry Cervical carcinoma HPV16/18E6 P16 (INK4a) 


  1. 1.
    Parkin DM, Bray F, Ferlay J, Pisani P (2005) Global cancer statistics 2002. CA Cancer J Clin 55:74–108PubMedCrossRefGoogle Scholar
  2. 2.
    Sankaranarayanan R, Madhukar A, Rajkumar R (2001) Effective screening programmes for cervical cancer in low- and middle-income developing countries. Bull World Health Organ 79(10):954–962PubMedCentralPubMedGoogle Scholar
  3. 3.
    GLOBOCAN 2000. Cancer incidence, mortality and prevalence worldwide, version 10. IARC Cancer Base No 5 Lyon, IARC Press. 2001Google Scholar
  4. 4.
    Bosch FX, de Sanjose S (2003) Human papillomavirus and cervical cancer – burden and assessment of causality. J Natl Cancer Inst Monogr 31:3–13PubMedCrossRefGoogle Scholar
  5. 5.
    Walboomers JM, Jacobs MV, Manos MM, Bosch FX, Kummer JA, Shah KV, Snijders PJ, Peto J, Meijer CJ, Muñoz N (1999) Human papillomavirus is a necessary cause of invasive cervical cancer worldwide. J Pathol 189(1):12–19PubMedCrossRefGoogle Scholar
  6. 6.
    de Sanjosé S, Diaz M, Castellsagué X, Clifford G, Bruni L, Muñoz N, Bosch FX (2007) Worldwide prevalence and genotype distribution of cervical human papillomavirus DNA in women with normal cytology: a meta-analysis. Lancet Infect Dis 7(7):453–459PubMedCrossRefGoogle Scholar
  7. 7.
    Wright TC Jr, Schiffman M (2003) Adding a test for human papillomavirus DNA to cervical- cancer screening. N Engl J Med 348:489PubMedCrossRefGoogle Scholar
  8. 8.
    What Causes Cancer of the Cervix?. American Cancer Society (2014-01-31).Google Scholar
  9. 9.
    Rashed MM, Bekele A (2011) The prevalence and pattern of HPV-16 immunostaining in uterine cervical carcinomas in ethiopian women: a pilot study. Pan Afr Med J 8:21PubMedCentralPubMedCrossRefGoogle Scholar
  10. 10.
    Missaoui N, Hmissa S, Frappart L, Trabelsi A, Ben Abdelkader A, Traore C, Mokni M, Yaacoubi MT, Korbi S (2006) p16INK4A overexpression and HPV infection in uterine cervix adenocarcinoma. Virchows Arch 448(5):597–603PubMedCrossRefGoogle Scholar
  11. 11.
    Bhurgri Y, Bhurgri A, Rahim A, Bhutto K, Pinjani PK, Usman A, Hasan SH (1999) The pattern of malignancies in Karachi (1995 to 1996). J Pak Med Assoc 49:157–161PubMedGoogle Scholar
  12. 12.
    Altaf F (2001) Pattern of cervical smear cytology in western region of Saudi Arabia. Ann Saudi Med 21:94–96Google Scholar
  13. 13.
    Jamal A, Al-Maghrabi JA (2003) Profile of pap smear cytology in the Western region of Saudi Arabia. Saudi Med J 24:1225–1229PubMedGoogle Scholar
  14. 14.
    Raza SA, Franceschi S, Pallardy S, Malik FR, Avan BI, Zafar A, Ali SH, Pervez S, Serajuddaula S, Snijders PJ et al (2010) Human papillomavirus infection in women with and without cervical cancer in Karachi, Pakistan. Br J Cancer 102:1657–1660PubMedCentralPubMedCrossRefGoogle Scholar
  15. 15.
    Castellsague X, Bosch FX, Munoz N, Meijer CJ, Shah KV, de Sanjose S, Eluf-Neto J, Ngelangel CA, Chichareon S, Smith JS et al (2002) Male circumcision, penile human papillomavirus infection, and cervical cancer in female partners. N Engl J Med 346:1105–1112PubMedCrossRefGoogle Scholar
  16. 16.
    Hammouda D, Clifford GM, Pallardy S, Ayyach G, Chekiri A, Boudrich A, Snijders PJ, van Kemenade FJ, Meijer CJ, Bouhadef A et al (2011) Human papillomavirus infection in a population-based sample of women in Algiers, Algeria. Int J Cancer 128:2224–2229PubMedCrossRefGoogle Scholar
  17. 17.
    Alsbeih G, Ahmed R, Al-Harbi N, Venturina LA, Tulbah A, Balaraj K (2011) Prevalence and genotypes’ distribution of human papillomavirus in invasive cervical cancer in Saudi Arabia. Gynecol Oncol 121:522–526PubMedCrossRefGoogle Scholar
  18. 18.
    Al-Badawi IA, Al-Suwaine A, Al-Aker M, Asaad L, Alaidan A, Tulbah A, Fe Bohol M, Munkarah AR (2011) Detection and genotyping of human papiloma virus in cervical cancer specimens from Saudi patients. Int J Gynecol Cancer 21:907–910PubMedCrossRefGoogle Scholar
  19. 19.
    Khorasanizadeh F, Hassanloo J, Khaksar N, Taheri SM, Marzaban M, Rashidi BH, Sari AA, Zendehdel K (2012) Epidemiology of cervical cancer and human papilloma virus infection among Iranian women - analyses of national data and systematic review of the literature. Gynecol Oncol 128:277–281PubMedCrossRefGoogle Scholar
  20. 20.
    Hussain S, Bharadwaj M, Nasare V, Kumari M, Sharma S, Hedau S, Das BC (2012) Human papillomavirus infection among young adolescents in India: impact of vaccination. J Med Virol 84:298–305PubMedCrossRefGoogle Scholar
  21. 21.
    Al-Muammar T, Al-Ahdal MN, Hassan A, Kessie G, Dela Cruz DM, Mohamed GE (2007) Human papilloma virus-16/18 cervical infection among women attending a family medical clinic in Riyadh. Ann Saudi Med 27:1–5PubMedCrossRefGoogle Scholar
  22. 22.
    Duttagupta C, Sengupta S, Roy M, Sengupta D, Bhattacharya P, Laikangbam P, Roy S, Ghosh S, Das R (2004) Are Muslim women less susceptible to oncogenic human papillomavirus infection? a study from rural Eastern India. Int J Gynecol Cancer 14:293–303PubMedCrossRefGoogle Scholar
  23. 23.
    de Boer MA, Vet JN, Aziz MF, Cornain S, Purwoto G, van den Akker BE, Dijkman A, Peters AA, Fleuren GJ (2006) Human papillomavirus type 18 and other risk factors for cervicacancer in Jakarta, Indonesia. Int J Gynecol Cancer 16:1809–1814PubMedCrossRefGoogle Scholar
  24. 24.
    Alsbeih G, Al-Harbi N, El-Sebaie M, Al-Badawi I (2013) HPV prevalence and genetic predisposition to cervical cancer in Saudi Arabia. Infect Agent Cancer 8(1):15PubMedCentralPubMedCrossRefGoogle Scholar
  25. 25.
    Ren C, Cheng X, Lu B, Yang G. Activation of interleukin-6/signal transducer and activator of transcription 3 by human papillomavirus early proteins 6 induces fibroblast senescence to promote cervical tumourigenesis through autocrine and paracrine pathways in tumour microenvironment. Eur J Cancer. 2013 Aug 14.Google Scholar
  26. 26.
    Kaewprag J, Umnajvijit W, Ngamkham J, Ponglikitmongkol M (2013) HPV16 oncoproteins promote cervical cancer invasiveness by upregulating specific matrix metalloproteinases. PLoS One 8(8):e71611PubMedCentralPubMedCrossRefGoogle Scholar
  27. 27.
    Sano T, Oyama T, Kashiwabara K, Fukuda T, Nakajima T (1998) Expression status of p16 protein is associated with human papillomavirus oncogenic potential in cervical and genital lesions. Am J Pathol 153(6):1741–1748PubMedCentralPubMedCrossRefGoogle Scholar
  28. 28.
    Bringold F, Serrano M (2000) Tumor suppressors and oncogenes in cellular senescence. Exp Gerontol 35(3):317–329PubMedCrossRefGoogle Scholar
  29. 29.
    Lin HP, Wang YP, Chiang CP (2011) Expression of p53, MDM2, p21, heat shock protein 70, and HPV 16/18 E6 proteins in oral verrucous carcinoma and oral verrucous hyperplasia. Head Neck 33:334–340PubMedGoogle Scholar
  30. 30.
    Amaro-Filho SM, Golub JE, Nuovo GJ, Cunha CB, Levi JE, Villa LL, Andrade CV, Russomano FB, Tristão A, Pires A, Nicol AF (2013) A comparative analysis of clinical and molecular factors with the stage of cervical cancer in a Brazilian cohort. PLoS One 8(3):e57810PubMedCentralPubMedCrossRefGoogle Scholar
  31. 31.
    Mendoza L, Mongelos P, Paez M, Castro A, Rodriguez-Riveros I, Gimenez G, Araujo P, Echagüe G, Diaz V, Laspina F, Castro W, Jimenez R, Marecos R, Ever S, Deluca G, Picconi MA (2013) Human papillomavirus and other genital infections in indigenous women from Paraguay: a cross-sectional analytical study. BMC Infect Dis 13(1):531PubMedCentralPubMedCrossRefGoogle Scholar
  32. 32.
    Simonella LM, Lewis H, Smith M, Neal H, Bromhead C, Canfell K (2013) Type-specific oncogenic human papillomavirus infection in high grade cervical disease in New Zealand. BMC Infect Dis 13:114PubMedCentralPubMedCrossRefGoogle Scholar
  33. 33.
    Cornet I, Gheit T, Iannacone MR, Vignat J, Sylla BS, Del Mistro A, Franceschi S, Tommasino M, Clifford GM (2013) HPV16 genetic variation and the development of cervical cancer worldwide. Br J Cancer 108(1):240–244PubMedCentralPubMedCrossRefGoogle Scholar
  34. 34.
    Tungteakkhun SS, Duerksen-Hughes PJ (2008) Cellular binding partners of the human papillomavirus E6 protein. Arch Virol 153(3):397–408PubMedCentralPubMedCrossRefGoogle Scholar
  35. 35.
    Togtema M, Pichardo S, Jackson R, Lambert PF, Curiel L, Zehbe I (2012) Sonoporation delivery of monoclonal antibodies against human papillomavirus 16 E6 restores p53 expression in transformed cervical keratinocytes. PLoS One 7(11):e50730PubMedCentralPubMedCrossRefGoogle Scholar
  36. 36.
    Lin HP, Wang YP, Chiang CP (2011) Expression of p53, MDM2, p21, heat shock protein 70, and HPV 16/18 E6 proteins in oral verrucous carcinoma and oral verrucous hyperplasia. Head Neck 33(3):334–340PubMedGoogle Scholar
  37. 37.
    Doorbar J, Quint W, Banks L, Bravo IG, Stoler M, Broker TR, Stanley MA (2012) The biology and life-cycle of human papillomaviruses. Vaccine 30(Suppl 5):F55–F70PubMedCrossRefGoogle Scholar
  38. 38.
    Cuschieri K, Wentzensen N (2008) Human papillomavirus mRNA and p16 detection as biomarkers for the improved diagnosis of cervical neoplasia. Cancer Epidemiol Biomarkers Prev 17(10):2536–2545PubMedCentralPubMedCrossRefGoogle Scholar
  39. 39.
    Iijima N, Goodwin EC, Dimaio D, Iwasaki A (2013) High-risk human papillomavirus E6 inhibits monocyte differentiation to Langerhans cells. Virology 444(1-2):257–262PubMedCentralPubMedCrossRefGoogle Scholar
  40. 40.
    Offringa RI, de Jong A, Toes RE, van der Burg SH, Melief CJ (2003) Interplay between human papillomaviruses and dendritic cells. Curr Top Microbiol Immunol 276:215–240PubMedGoogle Scholar
  41. 41.
    Sasagawa T, Takagi H, Makinoda S (2012) Immune responses against human papillomavirus (HPV) infection and evasion of host defense in cervical cancer. J Infect Chemother 18(6):807–815PubMedCrossRefGoogle Scholar
  42. 42.
    Niebler M, Qian X, Höfler D, Kogosov V, Kaewprag J, Kaufmann AM, Ly R, Böhmer G, Zawatzky R, Rösl F, Rincon-Orozco B (2013) Post-translational control of IL-1β via the human papillomavirus type 16 E6 oncoprotein: a novel mechanism of innate immune escape mediated by the E3-ubiquitin ligase E6-AP and p53. PLoS Pathog 9(8):e1003536PubMedCentralPubMedCrossRefGoogle Scholar
  43. 43.
    Bodaghi S, Wood LV, Roby G, Ryder C, Steinberg SM, Zheng ZM (2005) Could human papilloma viruses be spread through blood? J Clin Microbiol 43(11):5428–5434PubMedCentralPubMedCrossRefGoogle Scholar
  44. 44.
    Chen AC, Keleher A, Kedda MA, Spurdle AB, McMillan NA, Antonsson A (2009) Human papillomavirus DNA detected in peripheral blood samples from healthy Australian male blood donors. J Med Virol 81(10):1792–1796PubMedCrossRefGoogle Scholar
  45. 45.
    Foresta C, Bertoldo A, Garolla A, Pizzol D, Mason S, Lenzi A, De Toni L (2013) Human papillomavirus proteins are found in peripheral blood and semen Cd20+ and Cd56+ cells during Hpv-16 semen infection. BMC Infect Dis 13:593PubMedCentralPubMedCrossRefGoogle Scholar
  46. 46.
    Reijmers RM, Spaargaren M, Pals ST (2013) Heparan sulfate proteoglycans in the control of B cell development and the pathogenesis of multiple myeloma. FEBS J 280(10):2180–2193PubMedCrossRefGoogle Scholar
  47. 47.
    Leemans CR, Braakhuis BJ, Brakenhoff RH (2011) The molecular biology of head and neck cancer. Nat Rev Cancer 11:9–22PubMedCrossRefGoogle Scholar
  48. 48.
    Carozzi F, Gillio-Tos A, Confortini M, Del Mistro A, Sani C, De Marco L, Girlando S, Rosso S, Naldoni C, Dalla Palma P, Zorzi M, Giorgi-Rossi P, Segnan N, Cuzick J, Ronco G (2013) NTCC working group. risk of high-grade cervical intraepithelial neoplasia during follow-up in HPV-positive women according to baseline p16-INK4A results: a prospective analysis of a nested substudy of the NTCC randomised controlled trial. Lancet Oncol 14(2):168–176PubMedCrossRefGoogle Scholar
  49. 49.
    Chiou HL, Wu MF, Liaw YC, Cheng YWSchiffman M, Castle PE, Jeronimo J, Rodriguez AC, Wacholder S (2007) Human papillomavirus and cervical cancer. Lancet 370(9590):890–907CrossRefGoogle Scholar
  50. 50.
    Chen TH, Huang CC, Yeh KT, Chang SH, Chang SW, Sung WW, Cheng YW, Lee H (2012) Human papilloma virus 16 E6 oncoprotein associated with p53 inactivation in colorectal cancer. World J Gastroenterol 18(30):4051–8.14PubMedCentralPubMedCrossRefGoogle Scholar
  51. 51.
    Roncaglia MT, Fregnani JH, Tacla M, DE Campos SG, Caiaffa HH, Ab’saber A, DA Motta EV, Alves VA, Baracat EC, Longatto Filho A (2013) Characterization of p16 and E6 HPV-related proteins in uterine cervix high-grade lesions of patients treated by conization with large loop excision. Oncol Lett 6(1):63–68PubMedCentralPubMedGoogle Scholar
  52. 52.
    Giarnieri E, Mancini R, Pisani T, Alderisio M, Vecchione A (2000) Msh2, Mlh1, Fhit, p53, Bcl-2, and Bax expression in invasive and in situ squamous cell carcinoma of the uterine cervix. Clin Cancer Res 6(9):3600–3606PubMedGoogle Scholar
  53. 53.
    Volgareva G, Zavalishina L, Andreeva Y, Frank G, Krutikova E, Golovina D, Bliev A, Spitkovsky D, Ermilova V, Kisseljov F (2004) Protein p16 as a marker of dysplastic and neoplastic alterations in cervical epithelial cells. BMC Cancer 4:58PubMedCentralPubMedCrossRefGoogle Scholar
  54. 54.
    Hampl M, Wentzensen N, Vinokurova S, von Knebel-Doeberitz M, Poremba C, Bender HG, Kueppers V (2007) Comprehensive analysis of 130 multicentric intraepithelial female lower genital tract lesions by HPV typing and p16 expression profile. J Cancer Res Clin Oncol 133(4):235–245PubMedCrossRefGoogle Scholar
  55. 55.
    Mimica M, Tomić S, Kardum G, Hofman ID, Kaliterna V, Pejković L (2010) Ki-67 quantitative evaluation as a marker of cervical intraepithelial neoplasia and human papillomavirus infection. Int J Gynecol Cancer 20(1):116–119PubMedCrossRefGoogle Scholar
  56. 56.
    Wang JL, Zheng BY, Li XD, Nokelainen K, Angström T, Lindström MS, Wallin KL (2005) p16INK4A and p14ARF expression pattern by immunohistochemistry in human papillomavirus-related cervical neoplasia. Mod Pathol 18(5):629–637PubMedCrossRefGoogle Scholar
  57. 57.
    Murphy N, Ring M, Killalea AG et al (2003) p16INK4A as a marker for cervical dyskaryosis: CIN and cGIN in cervical biopsies and ThinPrep smears. J Clin Pathol 56:56–63PubMedCentralPubMedCrossRefGoogle Scholar
  58. 58.
    Jung YS, Kato I, Kim HR (2013) A novel function of HPV16-E6/E7 in epithelial-mesenchymal transition. Biochem Biophys Res Commun 435(3):339–344PubMedCrossRefGoogle Scholar
  59. 59.
    Qi ZL, Huo X, Xu XJ, Zhang B, Du MG, Yang HW, Zheng LK, Li J, Shen ZY (2006) Relationship between HPV16/18 E6 and 53, 21WAF1, MDM2, Ki67 and cyclin D1 expression in esophageal squamous cell carcinoma: comparative study by using tissue microarray technology. Exp Oncol 28(3):235–240PubMedGoogle Scholar
  60. 60.
    Galgano MT, Castle PE, Atkins KA, Brix WK, Nassau SR, Stoler MH (2010) Using biomarkers as objective standards in the diagnosis of cervical biopsies. Am J Surg Pathol 34(8):1077–1087PubMedCentralPubMedCrossRefGoogle Scholar
  61. 61.
    Nicol AF, Golub JE, e Silva JR, Cunha CB, Amaro-Filho SM, Oliveira NS, Menezes W, Andrade CV, Russomano F, Tristão A, Grinsztejn B, Friedman RK, Oliveira MP, Pires A, Nuovo GJ (2012) An evaluation of p16 (INK4a) expression in cervical intraepithelial neoplasia specimens, including women with HIV-1. Mem Inst Oswaldo Cruz 107(5):571–577PubMedCrossRefGoogle Scholar
  62. 62.
    Cheah PL, Looi LM, Teoh KH, Mun KS, Nazarina AR (2012) p16 (INK4a) is a useful marker of human papillomavirus integration allowing risk stratification for cervical malignancies. Asian Pac J Cancer Prev 13(2):469–472PubMedCrossRefGoogle Scholar

Copyright information

© Arányi Lajos Foundation 2014

Authors and Affiliations

  1. 1.Department of Pathology, College of MedicineAssiut UniversityAssiutEgypt
  2. 2.Department of Obstetrics and Gynacology, College of MedicineQassim UniversityBuridahKingdom of Saudi Arabia
  3. 3.Department of Pathology, College of MedicineQassim UniversityBuridahKingdom of Saudi Arabia

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