Reduced expression of connexins (Cxs), gap junction proteins, is frequently reported in malignant cell lines and tumors, whereas recent studies suggested that C x 26, a subtype of Cxs, might help tumor cells acquire malignant phenotypes. To examine this suggestion in the clinical setting, 50 lung squamous cell carcinomas (SCCs) were stained with the anti-C x 26 antibody. No C x 26-specific signals were detectable in 34 tumors (group I; 68%), whereas the remaining 16 were judged positive for C x 26 (group II; 32%). In 14 tumors of group II, C x 26-specific signals were detected not in all SCC cells but in SCC cells facing the tumor stroma or capsule, in which the signals were localized on the plasma membrane. Involved lymph nodes of group-II patients often contained metastatic foci consisting of all C x 26-positive cells. The proportion of C x 26-positive to C x 26-negative SCC cells in the metastatic nodes was larger than that in the corresponding primary tumors. C x 26-positive SCC cells seemed to be more invasive and metastatic than negative ones. Consistently, the 5-year cancer-specific survival rate of group-II patients was significantly lower than that of group-I patients (12.5 vs 38.9%; P=0.0391). Multivariate analysis demonstrated that C x 26 expression (P=0.0448) as well as pathological stage (P=0.0338) and vascular invasion (P=0.0191) were independent, significant prognostic predictors. These results suggest that C x 26 may represent an essential effector for controlling the biological aggressiveness of lung SCC tumor.