Overexpression of epidermal growth factor receptor in human head and neck squamous carcinoma cell lines correlates with matrix metalloproteinase-9 expression and in vitro invasion

Numerous reports have shown an association between overexpression of the ep idermal growth factor receptor (EGFR), and poor prognosis in head and neck squamous cell carcinomas (HNSCC), however, the underlying mechanisms are st ill unclear. In the present study, we set out to determine whether EGFR exp ression was associated with in vitro invasive capacity in a panel of four e stablished and ten newly derived HNSCC lines. Ten of the cell lines express ed high levels of EGFR as determined by a ligand-binding assay and dot blot analysis, whereas the remaining four showed weak overexpression or normal levels of EGFR. The ability of cells to invade through Matrigel was found t o be higher in the EGFR overexpressing cell lines (p < 0.0001), Expression levels of matrix metalloproteinases (MMP-1, MMP-2, MMP-3, MMP-7, MMP-9, MMP -10, MMP-11, MMP-13, MTI-MMP) and tissue inhibitors of MMP (TIMP-1, TIMP-2) were evaluated by semiquantitative RT-PCR, substrate zymography and wester n blot. We found a strong positive correlation between EGFR levels and the expression of MMP-9 mRNA (r(2) = 0.95; p < 0.0001), MMP-9 enzyme activity ( r(2) = 0.8099; p < 0.0001) and an inverse correlation with TIMP-1 (r(2) 0.4 8; p = 0.0059), In six selected HNSCC lines, in vitro invasion was assayed in the presence of an anti-EGFR monoclonal antibody, ICR62, A significant r eduction of invasion in four selected EGFR-overexpressing cell lines was fo und with 30 nM ICR62 (from 50% to 70%; P < 0.001) but there was no effect i n two cell lines with normal EGFR levels. Our results show that the in vitr o invasive phenotype of HNSCC lines correlates with high EGFR and MMP-9 exp ression, and it is therefore suggested that the EGFR signaling pathway migh t play an important role in the invasive behavior of HNSCC via specific upr egulation of MMP-9 and downregulation of TIMP-1, Int. J. Cancer 86:307-317, 2000, (C) 2000 Wiley-Liss, Inc.