Skip to main content
Log in

Activation of Wnt-β-catenin pathway in basal–parabasal layers of normal cervical epithelium comparable during development of uterine cervical carcinoma

  • Published:
Molecular and Cellular Biochemistry Aims and scope Submit manuscript


In this study, importance of Wnt-β-catenin pathway in the development of uterine cervical carcinoma was evaluated. For this purpose, the profiles (expression/methylation/deletion) of β-catenin, p-β-catenin (Y654), Wnt3a, and APC were studied in disease free normal cervical epithelium (n = 9), adjacent normal cervical epithelium of primary tumors (n = 70), CIN (n = 28), CACX (n = 102) samples, and two CACX cell lines (HeLa and SiHa). Immunohistochemical analysis revealed high/medium (74–95%) expression of β-catenin/p-β-catenin (Y654) and Wnt3a and low expression (23–26%) of APC in proliferating basal–parabasal layers contrary to differentiated spinous layer in normal cervix irrespective of HPV16 infection. The expression profile of the genes in the basal–parabasal layers did not change significantly during development of CACX. High (66%) promoter methylation of APC was seen in basal–parabasal layers and the cervical lesions (42–69%), unlike in spinous layers (25%). The promoter methylation status of APC was validated by in vitro demethylation experiments using 5-aza-dC in CACX cell lines. However, additional deletion of APC was significantly increased from CIN (12%) to stage I/II (40%) and became comparable in stage III/IV (48%) of the tumor. Patients with alterations (deletion/methylation) of APC and high/medium expression of Wnt3a/β-catenin/p-β-catenin (Y654) showed significantly poor survival. Thus our data indicate that cumulative effect of Wnt3a overexpression and APC inactivation are needed for overexpression of β-catenin during the development of CACX.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Subscribe and save

Springer+ Basic
EUR 32.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or Ebook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others


  1. Arbyn M, Castellsague X, de Sanjose S, Bruni L, Saraiya M, Bray F, Ferlay J (2008) Worldwide burden of cervical cancer in 2008. Ann Oncol 22:2675–2686

    Article  Google Scholar 

  2. WHO/ICO Information Centre on Human Papilloma Virus (HPV) and Related Diseases report in India. Summary Report. 2014

  3. Munoz N, Bosch FX, de Sanjose S, Herrero R, Castellsague X, Shah KV et al (2003) Epidemiologic classification of human papillomavirus types associated with cervical cancer. N Engl J Med 348:518–527

    Article  PubMed  Google Scholar 

  4. Snijders PJ, Steenbergen RD, Heideman DA, Meijer CJ (2006) HPV-mediated cervical carcinogenesis: concepts and clinical implications. J Pathol 208:152–164

    Article  CAS  PubMed  Google Scholar 

  5. Lopez J, Ruiz G, Organista-Nava J, Gariglio P, Garcia-Carranca A (2012) Human papillomavirus infections and cancer stem cells of tumors from the uterine cervix. Open Virol J 6:232–240

    Article  PubMed  PubMed Central  Google Scholar 

  6. Chakraborty C, Dutta S, Mukherjee N, Samadder S, Roychowdhury A, Roy A, Mondal RK, Basu P, Roychoudhury S, Panda CK (2015) Inactivation of PTCH1 is associated with the development of cervical carcinoma: clinical and prognostic implication. Tumour Biol 36:1143–1154

    Article  CAS  PubMed  Google Scholar 

  7. Uhlen M et al (2005) A human protein atlas for normal and human cancer tissues based on antibody proteomics. Mol Cell Proteom 4(12):1920–1932 (Human protein atlas: Release version 15,

    Article  CAS  Google Scholar 

  8. Zhang Yanna, Liu Bangzhong, Zhao Qingyu, Hou Teng, Huang Xin (2014) Nuclear localization of β-catenin is associated with poor survival and chemo-/radioresistance in human cervical squamous cell cancer Int J. Clin Exp Pathol 7:3908–3917

    Google Scholar 

  9. Pesterel Hadce Elf, Erdoan Gülgün, Fimfiek Tayyp, Karavel Fieyda (2006) Cadherin/catenin expression in squamous lesions of uterine cervix. Turk J Cancer 36(2):064–068

    Google Scholar 

  10. MacDonald BT, Tamai K, He X (2009) Wnt/beta-catenin signaling: components, mechanisms, and disease. Dev Cell 17:9–26

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Zi David, Ren N, Yang YJ (2015) Promoter methylation of APC gene in cervical cancer and precancerous lesions and its association with APC expression. J Pract Oncol 3:320–324

    Google Scholar 

  12. Van Veelen W, Le NH, Helvensteijn W, Blonden L, Theeuwes M, Bakker ER, Franken PF, van Gurp L, Meijlink F, van der Valk MA, Kuipers EJ, Fodde R, Smits R (2011) β-catenin tyrosine 654 phosphorylation increases Wnt signalling and intestinal tumorigenesis. Gut 9:1204–1212

    Article  Google Scholar 

  13. Dasgupta S, Mukherjee N, Roy S, Roy A, Sengupta A, Roychowdhury S, Panda CK (2002) Mapping of the candidate tumor suppressor genes’ loci on human chromosome 3 in head and neck squamous cell carcinoma of an Indian patient population. Oral Oncol 38:6–15

    Article  CAS  PubMed  Google Scholar 

  14. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory, Cold Spring Harbor

    Google Scholar 

  15. Singh RK, Maulik S, Mitra S, Mondal RK, Basu PS, Roychowdhury S, Panda CK (2006) Human papillomavirus prevalence in postradiotherapy uterine cervical carcinoma patients: correlation with recurrence of the disease. Int J Gynecol Cancer 16:1048–1054

    Article  CAS  PubMed  Google Scholar 

  16. Chakraborty C, Roychowdhury A, Samadder S, Roy A, Mandal RK, Basu P, Roychoudhury S, Panda CK (2016) Association of P16-RBSP3 inactivation with phosphorylated-RB1 over-expression in basal–parabasal layers of normal-cervix unchanged during CACX development. Biochem J 473:3221–3236

    Article  CAS  PubMed  Google Scholar 

  17. Mukherjee N, Bhattacharya N, Alam N, Roy A, Roychoudhury S, Panda CK (2012) Subtype-specific alterations of the Wnt signaling pathway in breast cancer: clinical and prognostic significance. Cancer Sci 103:210–220

    Article  CAS  PubMed  Google Scholar 

  18. Perrone F, Suardi S, Pastore E, Casieri P, Orsenigo M, Caramuta S, Dagrada G, Losa M, Licitra L, Bossi P et al (2006) Molecular and cytogenetic subgroups of oropharyngeal squamous cell carcinoma. Clin Cancer Res 12:6643–6651

    Article  CAS  PubMed  Google Scholar 

  19. Loginov VI, Maliukova AV, Seregin IuA, Khodyrev DS, Kazubskaia TP, Ermilova VD, Gar’kavtseva RF, Kiselev LL, Zabarovskii ER, Braga EA (2004) Methylation of the promoter region of the RASSF1A gene, a candidate tumor suppressor, in primary epithelial tumors. Molekuliarnaia biologiia 38:654–667

    CAS  PubMed  Google Scholar 

  20. Ivanova T, Petrenko A, Gritsko T, Vinokourova S, Eshilev E, Kobzeva V, Kisseljov F, Kisseljova N (2002) Methylation and silencing of the retinoic acid receptor-beta 2 gene in cervical cancer. BMC Cancer 2:4

    Article  PubMed  PubMed Central  Google Scholar 

  21. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using realtime quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 25:402–408

    Article  CAS  PubMed  Google Scholar 

  22. Maiti GP, Mondal P, Mukherjee N, Ghosh A, Ghosh S, Dey S, Chakrabarty J, Roy A, Biswas J, Roychoudhury S, Panda CK (2013) Overexpression of EGFR in head and neck squamous cell carcinoma is associated with inactivation of SH3GL2 and CDC25A genes. PLoS ONE 8(5):e63440

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Mukherjee N, Dasgupta H, Bhattacharya R, Pal D, Roy R, Islam S, Alam N, Biswas J, Roy A, Roychoudhury S, Panda CK (2015) Frequent inactivation of MCC/CTNNBIP1 and overexpression of phospho-beta-catenin(Y654) are associated with breast carcinoma: clinical and prognostic significance. Biochim Biophys Acta 1862:1472–1484

    Article  Google Scholar 

  24. Rigi-Ladiz MA, Kordi-Tamandani DM, Torkamanzehi A (2011) Analysis of hypermethylation and expression profiles of APC and ATM genes in patients with oral squamous cell carcinoma. Clin Epigenetics 3:6

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Karbova E, Davidson B, Metodiev K, Tropé CG, Nesland JM (2002) Adenomatous polyposis coli (APC) protein expression in primary and metastatic serous ovarian carcinoma. Int J Surg Pathol 3:175–180

    Article  Google Scholar 

  26. Song Y, Zhang C (2009) Hydralazine inhibits human cervical cancer cell growth in vitro in association with APC demethylation and re-expression. Cancer Chemother Pharmacol 63:605–613

    Article  CAS  PubMed  Google Scholar 

  27. Fang Jing-Yuan, Juan Lu, Chen Ying-Xuan, Yang Li (2003) Effects of DNA methylation on expression of tumor suppressor genes and proto-oncogene in human colon cancer cell lines. World J Gastroenterol 9:1976–1980

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Dong Seung Myung, Kim Hy-Sook, Rha Seo-Hee, Sidransky D (1982) Promoter hypermethylation of multiple genes in carcinoma of the uterine cervix. Clin Cancer Res 7:1982–1986

    Google Scholar 

  29. Gao X, Zacharek A, Grignon D, Liu H, Sakr W, Porter A, Chen Y, Honn K (1995) High-frequency of loss of expression and allelic deletion of the apc and mcc genes in human prostate-cancer. Int J Oncol 6:111

    CAS  PubMed  Google Scholar 

Download references


The authors thank the Director of Chittaranjan National Cancer Institute, Kolkata, India. We are grateful to Professor (Dr.) H. zur Hausen and Professor (Dr. Mrs.) E.M. de Villiers for their generous gift of HPV-16/18 plasmids. This work was supported by CSIR (Council of Scientific and Industrial Research, Government of India)-JRF/NET grant [File No.09/030(0059)/2010-EMR-I] to Mr C.Chakraborty, grant [Sr. No. 2121130723] from UGC (University Grants Commission, Government of India) to Mr. Sudip Samadder, grant [SR/SO/HS-116/2007] from the Department of Science and Technology (DST), Government of India and grant [No. 60(0111)/14/EMR-II of dt.03/11/2014] from CSIR (Council of Scientific and Industrial Research, Government of India) to Dr. C. K. Panda.

Author information

Authors and Affiliations


Corresponding author

Correspondence to Chinmay Kumar Panda.

Ethics declarations

Conflicts of interest

The authors declare that they have no conflict of interest.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Fig. S1 Schematic Distribution of the samples used for the study. Supplementary material 1 (TIFF 2030 kb)

Fig. S2 Laser capture microdissection of normal cervical epithelium.Supplementary material 2 (TIFF 1077 kb)


Fig. S3 Kaplan–Meier 5-year survival probability curves with cumulative survival of CACX patients by Wnt3a protein expression. Supplementary material 3 (TIFF 1396 kb)

Table S1 Primers for promoter methylation analysis APC. Supplementary material 4 (DOCX 11 kb)

Table S2 Primers for mRNA expression analysis of APC. Supplementary material 5 (DOCX 11 kb)

Table S3 Primers for deletion analysis of APC. Supplementary material 6 (DOCX 11 kb)


Table S4 Expression/methylation status of APC, β-catenin, p-β-catenin (Y654) and Wnt3a in different normal cervical epithelium. Supplementary material 7 (DOCX 33 kb)


Table S5 Relation between deletion (%) and methylation (%) of APC in CIN and CACX. Supplementary material 8 (DOCX 12 kb)

Table S6 Allelic status of APC in CIN and CACX. Supplementary material 9 (DOCX 74 kb)


Table S7 Correlation with expression/methylation/deletion status of APC, β-catenin, p-β-catenin and Wnt3a in CIN and CACX. Supplementary material 10 (DOCX 28 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Chakraborty, C., Samadder, S., Roychowdhury, A. et al. Activation of Wnt-β-catenin pathway in basal–parabasal layers of normal cervical epithelium comparable during development of uterine cervical carcinoma. Mol Cell Biochem 443, 121–130 (2018).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: