The expression of calpain small subunit 1 (Capn4) is correlated with the invasion of several types of tumors. However, the roles of Capn4 in non-small cell lung cancer (NSCLC) remain unclear. In this study, we found that the expression of Capn4 in NSCLC tissues was much higher than that in nontumorous samples. High levels of Capn4 expression were associated with lymph node metastasis and large tumor size in NSCLC patients. The 5-year overall survival rate in the Capn4(high) group was significantly lower than that in the Capn4(low) group. In multivariate analysis, Capn4 was identified as an independent prognostic factor for overall survival. Moreover, in an in vitro analysis, downregulation of Capn4 expression by siRNA suppressed the invasive potential of lung cancer cells. Finally, we demonstrated that Capn4 enhanced the invasion ability of lung cancer cells by upregulating the expression of matrix metalloproteinase 2. Our findings indicated that Capn4 may represent a potential therapeutic target and a novel prognostic marker of NSCLC.