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. Author manuscript; available in PMC: 2022 Sep 23.
Published in final edited form as: Science. 2022 Jul 28;377(6605):502–511. doi: 10.1126/science.abn7570

Fig. 2. KL-50 (4a) displays previously unseen MGMT-dependent, MMR-independent cytotoxicity in multiple isogenic cell models.

Fig. 2.

(A) IC50 values derived from short-term viability assays in LN229 MGMT+/−, MMR+/− cells treated with TMZ (1a) derivatives. aMGMT TI = IC50 (MGMT+/MMR+) divided by IC50 (MGMT−/MMR+). bMMR resistance index (RI) = IC50 (MGMT−/MMR−) divided by IC50 (MGMT−/MMR+). (B) Short-term viability assay curves for TMZ (1a), CCNU (14), KL-85 (4b), and KL-50 (4a) in LN229 MGMT+/−, MMR+/− cells. (C) Clonogenic survival curves for TMZ (1a) in LN229 MGMT+/−, MMR+/− cells, with representative images of wells containing 1000 plated cells treated with 30 μM TMZ (1a).(D) Clonogenic survival curves for KL-50 (4a) in LN229 MGMT+/−, MMR+/− cells, with representative images of wells containing 1000 plated cells treated with 30 μM KL-50 (4a). (E) Short-term viability assay curves for TMZ (1a) in DLD1 MSH6-deficient cells pretreated with 0.01% dimethyl sulfoxide (DMSO) control (CTR) or 10 μM O6BG (+O6BG) for 1 hour before TMZ (1a) addition to deplete MGMT. (F) Short-term viability assay curves for KL-50 (4a) in DLD1 MSH6-deficient cells pretreated with 0.01% DMSO control (CTR) or 10 μM O6BG (+O6BG) for 1 hour before KL-50 (4a) addition. (G) Short-term viability assay curves for TMZ (1a) in HCT116 MLH1−/− cells or HCT116 cells complemented with chromosome 3 carrying wild-type MLH1 (+Chr3) pretreated with 0.01% DMSO control or 10 μM O6BG (+O6BG) for 1 hour before TMZ (1a) addition. (H) Short-term viability assay curves for KL-50 (4a) in HCT116 MLH1−/− cells or HCT116 cells complemented with chromosome 3 carrying wild-type MLH1 (+Chr3) pretreated with 0.01% DMSO control or 10 μM O6BG (+O6BG) for 1 hour before KL-50 (4a) addition. For (B) to (H), points indicate the mean, and error bars indicate SD; n ≥ 3 technical replicates.