Abstract: Objective To explore the inhibitory effect of PSB0739 on the formation of semen-derived amyloid fibrils. Methods PAP248-286 (440 μmol/L) was incubated with PSB0739 at different concentrations, and at different time points of incubation, aliquots were taken from each sample for Congo red staining to detect the formation of amyloid fibers. The morphology of amyloid fibrils incubated in the presence or absence of PSB0739 was visualized using transmission electron microscope. The effect of PSB0739 on amyloid fibril formation was determined using virus infection assays at different time points, and the surface charges of amyloid fibril incubated with PSB0739 were calculated using a Zeta potentiometer. The cytotoxicity of PSB0739 in Hela cells was determined using MTT assay. The antiviral effect of PSB0738 against HIV- 1 was evaluated by infection assay. Results PSB0739 inhibited SEVI fibril formation in a dose-dependent manner in vitro. At 48 h of incubation, 220 μmol/L of PSB0739 obviously inhibited the formation of amyloid fibrils in 440 μmol/L of SEVI. Transmission electron microscopy revealed that 220 μmol/L PSB0739 prevented PAP248- 286 (440 μmol/L) from forming amyloid fibrils. PSB0739 antagonized SEVI-mediated enhancement of HIV-1 infection, and 1760 μmol/L of PSB0739 completely reversed the positive charge of SEVI (P<0.05). PSB0739 below the concentration of 62.5 μmol/L showed no obvious cytotoxicity in Hela cells in vitro (P>0.05). PSB0739 showed a direct anti-HIV activity with an IC50 of 21.77 ± 5.15 μmol/L. Conclusion PSB0739 can inhibit the formation of semen-derived amyloid fibrils in vitro.