The influence of Zr and Ti alloying on the microstructural evolution and mechanical properties of an Al-4Ni-0.4V alloy at elevated temperatures was investigated. The results showed that the coarsening rate of the eutectic Al3Ni phase in the Al-4Ni-0.4V-0.2Zr-0.2Ti alloy decreased from 21.1 nm3/s in the Al-4Ni-0.4V alloy to 7.4 nm3/s after annealing at 350 °C for 0 h to 120 h. The tensile strength, yield strength, and elongation at room temperature of the annealed Al-4Ni-0.4V-0.2Zr-0.2Ti alloy (350 °C, 8 h) were 181 MPa, 108 MPa, and 27.2 %, respectively. After tensile test at 350 °C, these values of the annealed Al-4Ni-0.4V-0.2Zr-0.2Ti alloy were 108 MPa, 72 MPa, and 29.8 %, respectively. Transmission electron microscopy results revealed the precipitation of the L12 ordered structure of the Al3M (M = Zr, V, Ti) phase, approximately 2 nm in size, in the α-Al after annealing. These phases exhibited excellent lattice matching with the α-Al and considerable precipitation strengthening. Segregation of Zr, V, and Ti elements at the α-Al/Al3Ni interface hindered Ni diffusion in the matrix, thereby reducing Al3Ni phase coarsening. Moreover, the Al3M phase precipitated at the interface exhibited favorable matching with the lattice planes of the α-Al and Al3Ni phase, which improved the thermal stability of the eutectic structure and the mechanical properties at elevated temperatures.