ALKBH2 overexpression increases resistance to methylating agents. (A) Clones stably overexpressing ALKBH2 were established by transfection of U87 and U251 cells with a His-tagged plasmid encoding ALKBH2. The upper band in the western blot represents the His-tagged plasmid, whereas the lower band represents endogenous ALKBH2 expression. (B) U87 and (C) U251 ALKBH2-overexpressing clones and their respective controls were treated with temozolomide (TMZ), methyl methanesulfonate (MMS), or doxorubicin (DOX) for 96h before drug-induced cytotoxicity was determined using an MTS assay. Cell viability (%) was calculated relative to corresponding control cells receiving drug vehicle only. The results shown were obtained using U87 clone #1 and U251 clone #1. Equally significant data were obtained using U87 clone #2 and U251 clone #2 (data not shown). *P < .05, **P < .01. (E) Colony formation assay using U251 ALKBH2-overexpressing cells and their respective controls. *P < .05, **P < .01.