Epidithiodiketopiperazines (ETPs) are a class of fungal secondary metabolites (SMs) featuring a transannular disulfide bridge at the diketopiperazine (DKP) core. The complex structures and biological activities have attracted widespread attention from biologists and chemists. In this study, we identified five novel ETP derivatives 4–7 and 4′ from three gene deletion mutants of Trichoderma hypoxylon CGMCC 3.17906, including ΔtdaP, ΔtdaQ, and ΔtdaQΔtdaI. Their structures were characterized through NMR and HR-ESI-MS data analysis. Compounds 4 and 4′ have unique heteroatom substitutions at the α and α′ positions, compound 5 possesses a unique α, β′-disulfide bridge, and compounds 6 and 7 contain a C3′-(thio)carbonyl group. Based on structural elucidation and biosynthetic pathway of α, β′-disulfide bridged ETPs, we also proposed the formation of 4–7 and 4′. This study not only expands the chemical diversity of ETPs, but also offers new mechanistic insights into the biosynthetic pathways of fungal ETPs.