[Abstract] Objective:To explore the anti-tumor effect of CAR-T cells secreting PD-1 scFv on gastric cancer. Method: We selected EGFR as the target of CAR-T cells and constructed second-generation EGFR-CAR-T cells (EGFR BB-z) and fourth-generation EGFR-CAR-T cells secreting PD-1 scFv (EGFR BB-z/E30). The anti-tumor activity was examined after in vitro activation and long-term stimulation, and its tumor suppression ability was validated through a mouse gastric cell xenograft model. Results: EGFR was highly expressed in gastric cancer tissues and cells (all P<0.01). EGFR BB-z and EGFR BB-z/E30 cells were successfully obtained by lentivirus infection. In vitro experiments showed that compared with EGFR BB-Z, EGFR BB-Z/E30 had longer long-term proliferation ability and stronger tumor killing activity (all P<0.01). In vivo experiments also validated that EGFR BB-z/E30 had obvious tumor inhibitory function in subcutaneous gastric tumor cell transplanted xenograft model and patient-derived tumor xenograft model (PDX) (all P<0.01). It also significantly increased T cell infiltration in tumor site and decreased the expression level of PD-1 (P<0.01 or P<0.05) on EGFR BB-z/E30 cell surface as well as the high secretion of IFN-γ (P<0.05). Conclusion: EGFR-CAR-T cell EGFR BB-z/E30 secreting PD-1 scFv can significantly inhibit the progression of gastric cancer and provide a potential new strategy for the treatment of gastric cancer.