Wu Y, Palad A J, Wasilenko W J, Blackmore P F, Pincus W A, Schechter G L, Spoonster J R, Kohn E C, Somers K D
Departments of Microbiology, Eastern Virginia Medical School, Norfolk, Virginia 23507, USA.
Clin Cancer Res. 1997 Nov;3(11):1915-21.
Local invasion and lymph node metastasis are correlated with a decreased overall survival in head and neck cancer patients and warrant new strategies to intervene in the metastatic cascade. One approach is to focus on the intracellular signaling pathways underlying the metastatic process. A common regulatory point in several signal transduction pathways 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 microM) 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 invasive phenotype of HNSCC cell lines were assessed by a chemo-invasion assay. HNSCC cell lines exhibited a range of invasive potential as measured by the capacity of tumor cells to penetrate a reconstituted basement membrane of Matrigel. HNSCCs were classified as highly invasive (EVSCC14M and EVSCC17M) or weakly invasive (EVSCC18, EVSCC19M, UMSCC10A, and FaDu). Treatment of HNSCC cell lines with 10 microM CAI for 24 h reduced invasion 2-14-fold in a dose-dependent manner. HNSCCs also exhibited 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 Mr 72,000 type IV collagenase [matrix metalloproteinase (MMP)-2)] and Mr 92,000 type IV collagenase (MMP-9). All HNSCC cell lines secreted MMP-2 and/or MMP-9 into conditioned medium. Treatment of cells with 10 microM CAI for 24 h resulted in a reduction of both MMP-2 and MMP-9 production. The results demonstrate that CAI blocks cellular proliferation, migration, chemoinvasion, and MMP production by HNSCC in vitro and identify calcium-dependent signaling as a new target for inhibition of the malignant phenotype of HNSCC.
局部侵袭和淋巴结转移与头颈癌患者总体生存率降低相关，因此需要新的策略来干预转移过程。一种方法是关注转移过程背后的细胞内信号通路。几个信号转导通路中的一个共同调节点是细胞内钙稳态。我们评估了一种新型钙内流抑制剂羧酰胺基三唑（CAI）对源自头颈部鳞状细胞癌（HNSCC）的细胞系生长和侵袭表型的影响。CAI以剂量依赖性（IC50，13 - 15 microM）和可逆方式抑制FaDu和EVSCC17M细胞的生长。CAI还导致受体介导的钙升高对几种钙动员激动剂（包括表皮生长因子和缓激肽）普遍减弱。通过化学侵袭试验评估CAI对HNSCC细胞系侵袭表型的影响。通过肿瘤细胞穿透基质胶重组基底膜的能力来衡量，HNSCC细胞系表现出一系列侵袭潜力。HNSCC被分类为高侵袭性（EVSCC14M和EVSCC17M）或低侵袭性（EVSCC18、EVSCC19M、UMSCC10A和FaDu）。用10 microM CAI处理HNSCC细胞系24小时以剂量依赖性方式使侵袭减少2 - 14倍。通过趋化试验测量，HNSCC也表现出不同的运动性。EVSCC14M和EVSCC17M运动性高，而EVSCC18、EVSCC19M、UMSCC10A和FaDu运动性较低。CAI减少了所有细胞系的迁移。通过酶谱法分析HNSCC细胞系的条件培养基中分子量72,000的IV型胶原酶[基质金属蛋白酶（MMP）-2]和分子量92,000的IV型胶原酶（MMP-9）的产生。所有HNSCC细胞系都向条件培养基中分泌MMP-2和/或MMP-9。用10 microM CAI处理细胞24小时导致MMP-2和MMP-9产生均减少。结果表明，CAI在体外阻断HNSCC的细胞增殖、迁移、化学侵袭和MMP产生，并确定钙依赖性信号传导是抑制HNSCC恶性表型的新靶点。
