IADR Abstract Archives
Calprotectin and the Development of Head and Neck Cancers (HNSCC)
Objectives: During cancer development, exogenous carcinogens, endogenous reactive species, and cellular deaminases can induce DNA damage. Unrepaired DNA damage can cause accumulating mutations critical to carcinogenesis. Cigarette smoke-induced mutagenesis dominates the HNSCC mutational landscape. Calprotectin (S100A8/A9) acts as a tumor suppressor in HNSCC, controlling the G2/M cell cycle checkpoint and associated DNA damage responses (DDR) in vitro as we reported. We hypothesize, therefore, that oral premalignant lesions, likely cigarette smoking-associated, down-regulate the tumor suppressor S100A8/A9 and diminish the DDR.
Methods: Oral premalignant dysplasias (N=16) and invasive HNSCCs (N=38) were analyzed using immunochemistry for S100A8, S100A9 and EGFR; and DDR-related markers 53BP1 and γH2AX in S100A8/A9(+) and S100A8/A9-KD TR146 HNSCC lines using immunofluorescence microscopy. DNA fragmentation was visualized using comet assays and S100A8 and S100A9 methylation patterns were characterized by interrogating The Cancer Genome Atlas (TCGA).
Results: Strongly expressed in normal adjacent epithelium, S100A8/A9 protein levels progressively decline as oral epithelial dysplasia transforms into invasive carcinoma. Loss of S100A8/A9 associates with poorer tumor differentiation (higher grade) and increased membranous and cytoplasmic EGFR expression, a key negative prognosticator of HNSCC in vitro and ex vivo. Based on TCGA data, the S100A8 and S100A9 promoter regions become heavily methylated in HNSCC, suggesting that epigenetic changes driven initially by cigarette smoke cause down-regulation of S100A8/A9. In S100A8/A9(+) TR146 cells, silencing S100A8/A9 significantly compromised DDR; recruitment of 53BP1 and γH2AX is abolished. S100A8/A9(+) TR146 cells appear more radio-sensitive and show greater DNA fragmentation than S100A8/A9-KD cells, which are radio-resistant and evade nuclear fragmentation.
Conclusions:
In oral epithelium, S100A8/A9 down-regulation can lead to increased EGFR expression and defective DDR. While not itself mutated by smoking-related oxidative stress, S100A8/A9 down-regulation may create a permissive environment for mutation of other tumor suppressors and oncogenes in smoking-related dysplasias and cancers. Increased mutations are likely to facilitate progression from dysplasia to HNSCC.
Methods: Oral premalignant dysplasias (N=16) and invasive HNSCCs (N=38) were analyzed using immunochemistry for S100A8, S100A9 and EGFR; and DDR-related markers 53BP1 and γH2AX in S100A8/A9(+) and S100A8/A9-KD TR146 HNSCC lines using immunofluorescence microscopy. DNA fragmentation was visualized using comet assays and S100A8 and S100A9 methylation patterns were characterized by interrogating The Cancer Genome Atlas (TCGA).
Results: Strongly expressed in normal adjacent epithelium, S100A8/A9 protein levels progressively decline as oral epithelial dysplasia transforms into invasive carcinoma. Loss of S100A8/A9 associates with poorer tumor differentiation (higher grade) and increased membranous and cytoplasmic EGFR expression, a key negative prognosticator of HNSCC in vitro and ex vivo. Based on TCGA data, the S100A8 and S100A9 promoter regions become heavily methylated in HNSCC, suggesting that epigenetic changes driven initially by cigarette smoke cause down-regulation of S100A8/A9. In S100A8/A9(+) TR146 cells, silencing S100A8/A9 significantly compromised DDR; recruitment of 53BP1 and γH2AX is abolished. S100A8/A9(+) TR146 cells appear more radio-sensitive and show greater DNA fragmentation than S100A8/A9-KD cells, which are radio-resistant and evade nuclear fragmentation.
Conclusions:
In oral epithelium, S100A8/A9 down-regulation can lead to increased EGFR expression and defective DDR. While not itself mutated by smoking-related oxidative stress, S100A8/A9 down-regulation may create a permissive environment for mutation of other tumor suppressors and oncogenes in smoking-related dysplasias and cancers. Increased mutations are likely to facilitate progression from dysplasia to HNSCC.