In the present work, using a multidisciplinary approach, we have deeply investigated the in vitro effect of the imidazobenzoxazin-5-thione MV1035 on glioblastoma aiming to demonstrate its capability to inhibit both ALKBH5 and ALKBH2 and its useful combination with TMZ as an effective approach to hinder glioma stem cell proliferation. This evidence concerns the gene ALKBH2 and glioma.