S100A2 is considered a putative tumor suppressor due to its loss or down-regulation in several cancer types. However, no mechanism has been described for the tumor suppressor role of S100A2. In this study, ectopic expression of S100A2 in the human malignant squamous cell carcinoma cell line KB resulted in a significant inhibition of proliferation, migration, and invasion. Moreover, S100A2 significantly reduced the number of colonies (>or=0.5 mm) formed in semisolid agar and decreased tumor growth and burden in nude mice. cDNA microarray analysis was used to compare mRNA expression profiles of vector- and S100A2-expressing isogenic cells. Among the genes deregulated by S100A2, the expression of cyclooxygenase-2 (COX-2) mRNA was significantly suppressed by S100A2 (2.4-fold). Western blot analysis confirmed that S100A2 reduced the expression of COX-2 protein in stably and transiently transfected KB and RPMI-2650 cells. COX-2 is frequently overexpressed in various types of cancer and plays an important role in tumor progression. Partial restoration of COX-2 expression attenuated the antitumor effect of S100A2 both in vitro and in vivo. Although the interplay between S100A2 and COX-2 remains to be clarified, these findings first showed a potent antitumor role of S100A2 in squamous cell carcinoma partly via reduced expression of COX-2.