MGMT expression and response to TMZ, propargyl and sulfoxide imidazotetrazine analogues. (A) Relative levels of MGMT expression were determined by Western blotting. DAOY, UW228-3 and MED3 cells showed high expression of MGMT while no expression was observed in MED1 cells. (B) Chemical structure of TMZ, propargyl and sulfoxide, shows different N3 groups. (C) Clonogenic survival was determined as in Figures 2, and 3, the data represent the IC50 values (mean ± SEM, n ≥ 3) TMZ inhibited survival in an MGMT-status dependent manner (IC50 at 290–540 μM in MGMT positive cells (DAOY, UW228-3 and MED3) compared to 31 μM in MGMT negative MED1 cells). Propargyl and sulfoxide derivatives of TMZ inhibited survival at 40–80 and 20–40 μM respectively in all cell lines, showing no dependence on MGMT expression (unpaired student’s t-test, ٭٭٭P < 0.0001; ٭P < 0.05).