Glioblastoma is one of the most treatment-resistant and lethal cancers, with a subset of self-renewing brain tumour stem cells (BTSCs), driving therapy resistance and relapse. Here, we report that mubritinib effectively impairs BTSC stemness and growth. Mechanistically, bioenergetic assays and rescue experiments showed that mubritinib targets complex I of the electron transport chain, thereby impairing BTSC self-renewal and proliferation. Gene expression profiling and Western blot analysis revealed that mubritinib disrupts the AMPK/p27