Objective ·To analyze the effect of Escherichia coli outer membrane vesicle (E.coli-OMV) on the proliferation of 4T1 breast cancer cells in vitro and the inhibition of tumor growth in BALB/c-4T1 tumor-bearing mice in vivo. Methods ·OMVs were collected from the culture supernatant of E.coli and characterized. The uptake of E.coli-OMV by 4T1 cells was detected by fluorescent label tracking method. The effect of E.coli-OMV on 4T1 cell proliferation was detected by CCK-8 method. The effect of E.coli-OMV on 4T1 cell cycle was detected by flow cytometry. The BALB/c-4T1 tumor-bearing mouse models were established by subcutaneous inoculation, and the mice were divided into E.coli-OMV group and Control group, with 10 mice in each group. The mice in the E.coli-OMV group were injected with 0.25 mg/kg E.coli-OMV every 2 d, while the mice in the Control group were injected with equal doses of PBS. The changes in body weight, 40 d survival rate, tumor volume and tumor weight of the two groups of tumor-bearing mice were observed. The pathological morphology of the tumor tissues was evaluated by hematoxylin-eosin staining (H-E staining). The expression of proliferating cell nuclear antigen (PCNA) and CyclinD1 in tumor tissues was observed by immunohistochemical staining. Results ·E.coli-OMVwas spherical membrane vesicle structure with a particle size of (216.00±18.30) nm, which expressed E.coli outer membrane protein A (OmpA) and OmpC. Fluorescence microscopy results showed that 4T1 cells could intake E.coli-OMV. CCK-8 results showed that the inhibitory effect of E.coli-OMV on 4T1 cells was positively correlated with time-dose. Flow cytometry results showed that E.coli-OMV arrested the growth cycle of 4T1 cells in G₀/G₁ phase. In vivo experiments showed that compared with the Control group, body weight of mice in the E.coli-OMV group decreased slightly after the initial injection (P=0.031), and then recovered, while 40 d survival rate increased (P=0.037). The growth of tumor volume and weight of mice in E.coli-OMV group were lower than those in the Control group (P=0.041, P=0.004). Its tumor volume inhibition rate reached 29.69%, and tumor weight inhibition rate reached 49.81%. The results of H-E staining showed that nuclear splitting images of tumor tissues of mice in the E.coli-OMV group decreased compared to the Control group (P=0.038). The results of immunohistochemical staining showed that the positive expression of PCNA and CyclinD1 in the tumor tissues of mice in the E.coli-OMV group decreased compared to the Control group (P=0.031, P=0.002). Conclusion ·Both in vitro and in vivo studies show that E.coli-OMV can significantly inhibit the proliferation of 4T1 cells.