Intracellular calprotectin (S100A8/A9) controls epithelial differentiation and caspase-mediated cleavage of EGFR in head and neck squamous cell carcinoma

Oral Oncol. 2019 Aug:95:1-10. doi: 10.1016/j.oraloncology.2019.05.027. Epub 2019 Jun 4.

Abstract

Objectives: Calprotectin (S100A8/A9) appears to function as a tumor suppressor in head and neck squamous cell carcinoma (HNSCC) and expression in the carcinoma cells and patient survival rates are directly related. We seek to characterize the suppressive role of calprotectin in HNSCC.

Aims: (1) Investigate changes in S100A8/A9 expression as oral carcinogenesis progresses and (2) determine whether intracellular calprotectin can regulate epidermal growth factor receptor (EGFR), a negative prognostic factor, in HNSCC.

Materials and methods: Using immunohistochemistry (IHC), S100A8/A9 was analyzed in HNSCC specimens (N = 46), including well-differentiated (WD, N = 19), moderately-differentiated (MD, N = 14), poorly-differentiated (PD, N = 5) and non-keratinizing/basaloid (NK/BAS, N = 8), and premalignant epithelial dysplasias (PED, N = 16). Similarly, EGFR was analyzed in HNSCCs (N = 21). To determine whether calprotectin and EGFR expression are mechanistically linked, TR146 HNSCC cells that are S100A8/A9-expressing or silenced (shRNA) were compared for EGFR levels and caspase-3/7 activity using western blotting and immunofluorescence microscopy.

Results: In normal oral mucosal epithelium, S100A8/A9 stained strongly in the cytoplasm and nucleus of suprabasal cells; basal cells were consistently S100A8/A9 negative. In PED and HNSCC, S100A8/A9 expression was lower than in adjacent normal epithelial tissues (NAT) and declined progressively in WD, MD, PD and NK/BAS HNSCCs. S100A8/A9 and EGFR levels appeared inversely related, which was simulated in vitro when S100A8/A9 was silenced in TR146 cells. Silencing S100A8/A9 significantly reduced caspase-3/7 activity, whereas EGFR levels increased.

Conclusions: In HNSCC, S100A8/A9 is directly associated with cellular differentiation and appears to promote caspase-3/7-mediated cleavage of EGFR, which could explain why patients with S100A8/A9-high tumors survive longer.

Keywords: Calprotectin; Caspase-3/7; EGFR; Epithelial differentiation; Head and neck squamous cell carcinoma; NGF; P16+ squamous cell carcinoma; Premalignant epithelial dysplasia; S100A8/A9; VEGF-A; VEGF-C.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adult
  • Aged
  • Aged, 80 and over
  • Apoptosis
  • Calgranulin A / metabolism
  • Calgranulin B / metabolism
  • Caspase 3 / metabolism
  • Caspase 7 / metabolism
  • Cell Differentiation
  • Cell Line, Tumor
  • Epithelial Cells / pathology
  • ErbB Receptors / metabolism
  • Female
  • Gene Knockdown Techniques
  • Head and Neck Neoplasms / pathology*
  • Humans
  • Leukocyte L1 Antigen Complex / genetics
  • Leukocyte L1 Antigen Complex / metabolism*
  • Male
  • Middle Aged
  • Mouth Mucosa / cytology
  • Mouth Mucosa / pathology*
  • Proteolysis
  • RNA, Small Interfering / metabolism
  • Squamous Cell Carcinoma of Head and Neck / mortality
  • Squamous Cell Carcinoma of Head and Neck / pathology*
  • Survival Rate
  • Young Adult

Substances

  • Calgranulin A
  • Calgranulin B
  • Leukocyte L1 Antigen Complex
  • RNA, Small Interfering
  • S100A8 protein, human
  • S100A9 protein, human
  • EGFR protein, human
  • ErbB Receptors
  • CASP3 protein, human
  • CASP7 protein, human
  • Caspase 3
  • Caspase 7