Background: PRMT5 is a major enzyme responsible for the post-translational symmetric demethylation of protein arginine residues, which has been validated as an effective therapeutic target for cancer. Thus, many nucleoside-based PRMT5 inhibitors have been reported in the past year. Objective: To discover a novel series of non-nucleoside PRMT5 inhibitors through a molecular docking-based virtual screening approach. Methods: Our in-house compound library was virtually screened using the Glide program, identifying a new PRMT5 inhibitor 1. Based on the structural similarity of hit 1, a series of structure-oriented derivatives, including 3a-3e, 7a-7g, and 12a-12f, were synthesized and selected for the inhibitory activity evaluation against PRMT5, as well as cytotoxicity against MV4-11 cell. Results: The analogs 7a-7e with benzimidazole core exhibited potent PRMT5 inhibitory activities, with 7e displaying the most potent activity with an IC50 of 6.81 +/- 0.12 mu M. In the anti-proliferative assay, compound 7e showed a strong inhibitory effect on MV4-11 cell growth. Finally, the binding mode of 7e with PRMT5 was predicted to provide insights for further structural optimization. Conclusion: The newly discovered PRMT5 inhibitors have potential antitumor activity against MV4-11 cells. This work highlighted this series of 3-(1H-benzo[d]imidazol-2-yl)aniline derivatives as novel anti-cancer lead compounds targeting PRMT5, which were worthy of further investigation.