We found plasmic OPRT (P = 0.024), TS (P = 0.044) expression significantly inversely, while DPD (P = 0.073), TP (P = 0.080) borderline inversely correlated with AUC0–24h of 5-FU. The number of CYP2A6 gene variants (P = 0.889) did not correlate with AUC0–24h. This was consistent with that CYP2A6 gene correlated with tegafur pharmacokinetics, but not with 5-FU pharmacokinetics . AUC0–24h of 5-FU, not OPRT/TS, was predictive of PFS revealed that other factors may also influence PFS by AUC0–24h of 5-FU. We did not demonstrate the number of CYP2A6 gene variants correlated with efficacy. Although some studies demonstrated patients having fewer CYP2A6 variants had better PFS in AGC patients with S-1 plus cisplatin, or S-1 plus docetaxel [6, 46], divergences on the relation between CYP2A6 genetic polymorphisms and response existed for both gastric and colorectal cancer patients with S-1 or S-1-based chemotherapy [47, 48]. In our study, second-line treatment excluded S-1 and tegafur. CYP2A6 converts enzymatically tegafur, the effector molecule of S-1, to 5-FU, and this role produces no meaning on second-line treatment.