p16(INK4a) /Ki-67 co-expression specifically identifies transformed cells in the head and neck region

Int J Cancer. 2015 Apr 1;136(7):1589-99. doi: 10.1002/ijc.29130. Epub 2014 Aug 19.


p16(INK4a) immunohistochemical overexpression is an overall reliable surrogate marker of human papillomavirus (HPV)-associated head and neck squamous cell carcinomas (HNSCC). However, cases of ambiguous p16(INK4a) overexpression are regularly detected in the head and neck: p16(INK4a) expression can be observed in non-malignant tissue, such as tonsillar crypt epithelium and a proportion of branchial cleft cysts. Additionally, diverse patterns of p16(INK4) expression can complicate interpretation of "p16(INK4a) -positivity". These aspects impede the unrestricted application of p16(INK4a) as a diagnostic marker in the head and neck. We hypothesized that combined detection of p16(INK4a) and the proliferation marker Ki-67 could support clarification of ambiguous p16(INK4a) expression in the head and neck by specifically indicating p16(INK4a) -expressing cells with proliferative activity. p16(INK4a) /Ki-67 co-expression in a combined staining procedure was correlated to distinct p16(INK4a) expression patterns and HPV status (HPV DNA followed by E6*I oncogene mRNA detection) in 147 HNSCC and 50 non-malignant head and neck samples. p16(INK4a) /Ki-67 co-expression only occurred in transformed cells of the head and neck. Co-expression was never detected in non-transformed cells. Combined p16(INK4a) /Ki-67 expression was stringently associated with a diffuse p16(INK4a) expression pattern. All HPV oncogene-expressing HNSCC showed p16(INK4a) /Ki-67 co-expression. We demonstrate that p16(INK4a) /Ki-67 co-expression occurs exclusively in transformed cells of the head and neck. Our findings indicate a substantial impact of combined p16(INK4a) /Ki-67 expression in the assessment of ambiguous p16(INK4a) expression in the head and neck by specifically identifying p16(INK4a) -expressing cells with proliferative activity. This property will be of considerable significance for head and neck histo- and cytopathology.

Keywords: E6*I RNA; HPV; Ki-67; head and neck squamous cell carcinoma; p16.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biomarkers, Tumor / genetics
  • Carcinoma, Squamous Cell / diagnosis
  • Carcinoma, Squamous Cell / genetics
  • Carcinoma, Squamous Cell / virology
  • Cell Line, Transformed
  • Cyclin-Dependent Kinase Inhibitor p16 / genetics*
  • Cyclin-Dependent Kinase Inhibitor p16 / metabolism
  • DNA, Viral / genetics
  • Gene Expression*
  • Head and Neck Neoplasms / diagnosis
  • Head and Neck Neoplasms / genetics
  • Head and Neck Neoplasms / virology
  • Humans
  • Immunohistochemistry
  • Ki-67 Antigen / genetics*
  • Ki-67 Antigen / metabolism
  • Oncogene Proteins, Viral / genetics
  • Oncogene Proteins, Viral / metabolism
  • Papillomaviridae / genetics
  • Papillomavirus Infections / complications
  • Repressor Proteins / genetics
  • Repressor Proteins / metabolism
  • Squamous Cell Carcinoma of Head and Neck


  • Biomarkers, Tumor
  • Cyclin-Dependent Kinase Inhibitor p16
  • DNA, Viral
  • E6 protein, Human papillomavirus type 16
  • Ki-67 Antigen
  • Oncogene Proteins, Viral
  • Repressor Proteins