INHIBITION OF HEAD AND NECK SQUAMOUS-CELL CARCINOMA GROWTH AND INVASION BY THE CALCIUM INFLUX INHIBITOR CARBOXYAMIDO-TRIAZOLE

Local invasion and lymph node metastasis are correlated with a decreas ed overall survival in head and neck cancer patients and warrant new s trategies to intervene in the metastatic cascade. One approach is to f ocus on the intracellular signaling pathways underlying the metastatic process. A common regulatory point in several signal transduction pat hways is intracellular calcium homeostasis. We assessed the effect of a novel calcium influx inhibitor, carboxyamido-triazole (CAI), on the growth and invasive phenotype of cell lines derived from head and neck squamous cell carcinoma (HNSCC). CAI inhibited the growth of FaDu and EVSCC17M cells in a dose-dependent (IC50, 13-15 mu M) and reversible manner. CAI also caused a generalized attenuation of receptor-mediated calcium elevation to several calcium mobilization agonists, including epidermal growth factor and bradykinin. The effects of CAI on the inv asive phenotype of HNSCC cell lines were assessed by a chemoinvasion a ssay. HNSCC cell lines exhibited a range of invasive potential as meas ured by the capacity of tumor cells to penetrate a reconstituted basem ent membrane of Matrigel. HNSCCs were classified as highly invasive (E VSCC14M and EVSCC17M) or weakly invasive (EVSCC18, EVSCC19M, UMSCC10A, and FaDu). Treatment of HNSCC cell lines with 10 mu M CAI for 24 h re duced invasion 2-14-fold in a dose-dependent manner. HNSCCs also exhib ited different motilities as measured by a chemotaxis assay. EVSCC14M and EVSCC17M were highly motile, whereas EVSCC18, EVSCC19M, UMSCC10A, and FaDu were less motile, CAI reduced the migration of all cell lines . Conditioned medium from HNSCC cell lines was analyzed by zymography for production of M-r 72,000 type IV collagenase [matrix metalloprotei nase (MMP)-2] and M-r 92,000 type IV collagenase (MMP-9). All HNSCC ce ll lines secreted MMP-2 and/or MMP-9 into conditioned medium. Treatmen t of cells with 10 mu M CAI for 24 h resulted in a reduction of both M MP-2 and MMP-9 production. The results demonstrate that CAI blocks cel lular proliferation, migration, chemoinvasion, and MMP production by H NSCC in vitro and identify calcium-dependent signaling as a new target for inhibition of the malignant phenotype of HNSCC.