Activation of c-MET through hepatocyte growth factor (HGF) increases tumorigenesis, induces resistance, and is associated with poor prognosis in various solid tumors. However, the clinical value of serum HGF (sHGF) in patients with advanced non-small cell lung cancer (NSCLC), especially those receiving cytotoxic chemotherapy, remains unknown. Here, we show that sHGF may be useful to predict tumor response and progression-free survival (PFS) in patients with advanced NSCLC. A total of 81 patients with NSCLC were investigated. sHGF levels were evaluated using ELISA at 4 time-points: at pre-treatment, at response-evaluation (1-2 months after treatment initiation), at the best tumor response, and at disease progression. As a control biomarker, CEA was also evaluated in lung adenocarcinoma. Positive-sHGF at response-evaluation predicted poor PFS compared with Negative-sHGF in both first-line (median, 153.5 vs. 288.0; P < 0.05) and second-line treatment (87.0 vs. 219.5; P = 0.01). In 55 patients that received cytotoxic chemotherapy, multiple Cox proportional hazards models showed significant independent associations between poor PFS and Positive-sHGF at response-evaluation (hazard ratio, 4.24; 95% CI, 2.05 to 9.46; P < 0.01). Lung adenocarcinoma subgroup analysis showed that in patients receiving second cytotoxic chemotherapy, there were no significant differences in PFS between patients with low-CEA compared with those with high-CEA, but Positive-sHGF at pre-treatment or at response-evaluation predicted poor PFS (35.0 vs. 132.0; P < 0.01, 50.0 vs. 215.0; P < 0.01, respectively). These findings give a rationale for future research investigating the merit of sHGF as a potential clinical biomarker to evaluate HGF/c-MET activity, which would be useful to indicate administration of c-MET inhibitors.