Abstract: Stage separation of two-stage-to-orbit (TSTO) vehicle under hypersonic inflow conditions will produce complex unsteady aerodynamic interference between the two stages, which directly increases the risk of TSTO separation. This complex aerodynamic interference is accompanied by the combined action of shock wave and boundary layer interaction, horseshoe vortex, shock wave, and wake interference between the two stages. In the current study, the complex models of the TSTO booster and orbiter are simplified into two three-dimensional (3-D) wedges. Based on the overset dynamic grid technology, the Navier-Stokes equations coupled with six-degree-of-freedom rigid body dynamic equations are solved to simulate and analyze the stage separation process, to explore the flow characteristics and physical mechanism of stage separation. In the numerical analysis, static and dynamic numerical simulations were carried out for the three-dimensional flowfield of TSTO under different orbiter’s lifting angles (β), and the flow patterns and interference characteristics with varying β were analyzed and discussed in detail. Combined with the flowfield structure and wall pressure distribution, as well as separated flow, the mechanism of the aerodynamic interference between the two stages during the TSTO separation, were clarified, and the effects of the lifting angle condition of the orbiter on the safe separation for the current TSTO model was discussed. The numerical results show that the interstage aerodynamic interference increases with the increase of the lifting angle of the orbiter, and decreases with the increase of the interstage clearance during stage separation. Before the release of the orbiter, the interstage aerodynamic interference and 3-D separated flow topology become more complex with the increase of β, and separation area increases, as well as the number of critical points. In the process of stage separation, the variation amplitude of the aerodynamic characteristics of both stages increases with the increase of β, and the separation time decreases. Moreover, the lifting angle of the orbiter is 6° ~ 8° would be conducive to the safe stage separation for the current TSTO model.