Quantification of HPV-16 Viral Load in Breast Cancer Tissues and Its Association with Tumor Stage, Histological Grade, and Lymph Node Metastasis
DOI:
https://doi.org/10.64354/twyqfj30Keywords:
Breast cancer; HPV-16; Viral load; qPCR; Lymph node metastasis; Tumor stage; Histological gradeAbstract
Background: Though HPV-16 has been discovered in breast cancer tissues globally, its clinical significance in breast carcinogenesis is uncertain. Viral load can separate temporary viral presence from physiologically relevant infection that boosts tumours. Methods: This cross-sectional analytical investigation examined 120 FFPE breast cancer tissues and 40 neighbouring non-neoplastic breast tissues from Al-Hussein Teaching Hospital in Karbala, Iraq, between January 2023 and December 2024. A commercial spin-column kit extracted DNA, and type-specific primers and a TaqMan probe were used for real-time quantitative PCR (qPCR) to detect and quantify HPV-16. Normalising HPV-16 copy count per cell to human RNase P gene copies. Clinicopathological parameters (tumour stage, histological grade, and lymph node status) and viral load were assessed using non-parametric tests and binary logistic regression. Results: We found HPV-16 DNA in 38 of 120 breast cancer samples (31.7%) and 3 of 40 (7.5%) non-neoplastic tissues (P = 0.003). An HPV-16-positive tumour had a median viral load of 2.74 × 10 22 copies/cell (IQR: 4.12 × 10 22 – 1.58 × 10⁴). Tumours with lymph node metastases (N+) and advanced-stage tumours (Stage III–IV: median 8.63 × 10³ vs. Stage I–II: median 6.81 × 10³; P = 0.001) had significantly higher viral loads (N+: median 6.27 × 10³ vs. N0: median 7.94 × 10²; Viral loads were substantially higher in Grade III tumours (P = 0.008). An independent predictor of lymph node metastasis was identified using logistic regression analysis, with a viral load of ≥ 10³ copies/cell (OR = 3.42; 95% CI: 1.18–9.91; P = HPV-16 viral load was substantially related with advanced breast cancer stage, high histological grade, and lymph node metastases. Quantifying HPV-16 viral load may be a biomarker for tumour aggressiveness and warrants multi-center prospective investigations.
References
1 Sung H, Ferlay J, Siegel RL, 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):209–249.
2 Bray F, Laversanne M, Sung H, et al. Global cancer statistics 2022: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin. 2024;74(3):229–263.
3 De Villiers EM. Relationship between steroid hormone contraceptives and HPV, cervical intraepithelial neoplasia and cervical carcinoma. Int J Cancer. 2003;103(6):705–708.
4 Zur Hausen H. Papillomaviruses and cancer: from basic studies to clinical application. Nat Rev Cancer. 2002;2(5):342–350.
5 Moody CA, Laimins LA. Human papillomavirus oncoproteins: pathways to transformation. Nat Rev Cancer. 2010;10(8):550–560.
6 Scheffner M, Werness BA, Huibregtse JM, et al. The E6 oncoprotein encoded by human papillomavirus types 16 and 18 promotes the degradation of p53. Cell. 1990;63(6):1129–1136.
7 Band V, Zajchowski D, Kulesa V, Sager R. Human papilloma virus DNAs immortalize normal human mammary epithelial cells and reduce their growth factor requirements. Proc Natl Acad Sci U S A. 1990;87(1):463–467.
8 Simoes PW, Medeiros LR, Simoes Pires PD, et al. Prevalence of human papillomavirus in breast cancer: a systematic review. Int J Gynecol Cancer. 2012;22(3):343–348.
9 Bae JM, Kim EH. Human papillomavirus infection and risk of breast cancer: a meta-analysis of case-control studies. Infect Agents Cancer. 2016;11:14.
10 Li N, Bi X, Zhang Y, Zhao P, Zheng T, Dai M. Human papillomavirus infection and sporadic breast carcinoma risk: a meta-analysis. Breast Cancer Res Treat. 2011;126(2):515–520.
11 Lawson JS, Glenn WK, Salyakina D, et al. Human papilloma viruses and breast cancer. Front Oncol. 2015;5:277.
12 Glenn WK, Heng B, Delprado W, Iacopetta B, Whitaker NJ, Lawson JS. Epstein-Barr virus, human papillomavirus and mouse mammary tumour virus as multiple viruses in breast cancer. PLoS One. 2012;7(11):e48788.
13 Josefsson AM, Magnusson PK, Ylitalo N, et al. Viral load of human papilloma virus 16 as a determinant for development of cervical carcinoma in situ: a nested case-control study. Lancet. 2000;355(9222):2189–2193.
14 Flores R, Papenfuss M, Klimecki WT, Giuliano AR. Cross-sectional analysis of oncogenic HPV viral load and cervical intraepithelial neoplasia. Int J Cancer. 2006;118(5):1187–1193.
15 Sun CA, Liu JF, Wu DM, Nieh S, Yu CP, Chu TY. Viral load of high-risk human papillomavirus in cervical squamous intraepithelial lesions. Int J Gynaecol Obstet. 2002;76(1):41–47.
16 Moberg M, Gustavsson I, Gyllensten U. Real-time PCR-based system for simultaneous quantification of human papillomavirus types 6, 11, 16, and 18. J Clin Microbiol. 2003;41(7):3221–3228.
17 Khan NA, Castillo A, Zafar N, et al. Human papillomavirus detected in female breast carcinomas in Pakistan. Br J Cancer. 2008;98(5):812–815.
18 Heng B, Glenn WK, Ye Y, et al. Human papilloma virus is associated with breast cancer. Br J Cancer. 2009;101(8):1345–1350.
19 Peitsaro P, Johansson B, Syrjanen S. Integrated human papillomavirus type 16 is frequently found in cervical cancer precursors as demonstrated by a novel quantitative real-time PCR technique. J Clin Microbiol. 2002;40(3):886–891.
20 Haghshenas MR, Mousavi T, Kheradmand M, Afshari M, Moosazadeh M. Human papillomavirus and breast cancer in Iran: a meta-analysis. Iran J Basic Med Sci. 2016;19(3):231–237.
21 Al-Lawati Z, Khamis AH, Al-Hamdani A, et al. Prevalence of human papillomavirus in breast cancer in Oman. Gulf J Oncol. 2020;1(33):59–64.
22 Manzouri L, Salehi R, Shariatpanahi S, Rezaian M. Prevalence of human papillomavirus among women with breast cancer since 2005–2009 in Isfahan. Adv Biomed Res. 2014;3:75.
23 Williams VM, Filippova M, Soto U, Duerksen-Hughes PJ. HPV-DNA integration and carcinogenesis: putative roles for inflammation and oxidative stress. Future Virol. 2011;6(1):45–57.
24 Fernandes JV, De Araujo JM, Fernandes TA. Biology and natural history of human papillomavirus infection. Open Access J Clin Trials. 2013;5:1–12.
25 Talora C, Sgroi DC, Crum CP, Dotto GP. Specific down-modulation of Notch1 signaling in cervical cancer cells is required for sustained HPV-E6/E7 expression and late steps of malignant transformation. Genes Dev. 2002;16(17):2252–2263.
26 Cardeal LB, Boccardo E, Termini L, et al. HPV16 oncoproteins induce MMPs/RECK-TIMP-2 imbalance in primary keratinocytes: possible implications in cervical carcinogenesis. PLoS One. 2012;7(3):e33585.
27 Munger K, Basile JR, Duensing S, et al. Biological activities and molecular targets of the human papillomavirus E7 oncoprotein. Oncogene. 2001;20(54):7888–7898.
28 Akil N, Yasmeen A, Kassab A, Ghabreau L, Aboussekhra A, Al Moustafa AE. High-risk human papillomavirus infections in breast cancer in Syrian women and their association with Id-1 expression: a tissue microarray study. Br J Cancer. 2008;99(3):404–407.
29 Yasmeen A, Bismar TA, Kandouz M, Foulkes WD, Heuer T, Al Moustafa AE. E6/E7 of HPV type 16 promotes cell invasion and metastasis of human breast cancer cells. Cell Cycle. 2007;6(16):2038–2042.
30 De Cremoux P, Thioux M, Lebigot I, Sigal-Zafrani B, Salmon R, Sastre-Garau X. No evidence of human papillomavirus DNA sequences in invasive breast carcinoma. Breast Cancer Res Treat. 2008;109(1):55–58.
31 Lindel K, Forster A, Altermatt HJ, Suter P, Rieben R. Breast cancer and human papillomavirus infection: no evidence of HPV etiology of breast cancer in a group of Swiss women. Breast. 2007;16(2):172–177.
32 Salman NA, Davies G, Majidy F, et al. Association of high risk human papillomavirus and breast cancer: a UK based study. Sci Rep. 2017;7:43591.
33 Khodabandehlou N, Mostafaei S, Etemadi A, et al. Human papillomavirus and breast cancer: the role of inflammation and viral expressed proteins. BMC Cancer. 2019;19(1):61.
Downloads
Published
Issue
Section
License
Copyright (c) 2026 New Journal of Biology and Medicine
This work is licensed under a Creative Commons Attribution 4.0 International License.
