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. 2021 Jun 22;20(9):1614–1626. doi: 10.1158/1535-7163.MCT-20-0502

Figure 4.

Figure 4. Evaluation of target engagement of DDRi across cell lines. A, PAR relative light units (RLU) in no radiotherapy controls (-IR), or following a 6 Gy radiotherapy dose alone or in combination with an IC50 or IC90 concentration of olaparib (μmol/L) as indicated. 1 μmol/L was used as the maximum on-target concentration in in vitro assays. Hence, where IC50 values were higher than 1 μmol/L (NCI-H1299 and NCI-H1703), the level of PARylation inhibition of an IC50 concentration was compared with a 1μmol/L on-target concentration rather than the IC90 concentration. B–E, Cell lines were irradiated with 6 Gy in the presence or absence of an IC50 or IC90 concentration of ceralasertib (B), KU-60648 (C), adavosertib (D) and three concentrations of AZD0156 as indicated (E). Cell extracts were collected after 30 minutes and western blotted for total and phosphorylated (p-) proteins. GAPDH, Tubulin and Vinculin served as total loading controls for either phospho or total protein blots as indicated.

Evaluation of target engagement of DDRi across cell lines. A, PAR relative light units (RLU) in no radiotherapy controls (-IR), or following a 6 Gy radiotherapy dose alone or in combination with an IC50 or IC90 concentration of olaparib (μmol/L) as indicated. 1 μmol/L was used as the maximum on-target concentration in in vitro assays. Hence, where IC50 values were higher than 1 μmol/L (NCI-H1299 and NCI-H1703), the level of PARylation inhibition of an IC50 concentration was compared with a 1μmol/L on-target concentration rather than the IC90 concentration. B–E, Cell lines were irradiated with 6 Gy in the presence or absence of an IC50 or IC90 concentration of ceralasertib (B), KU-60648 (C), adavosertib (D) and three concentrations of AZD0156 as indicated (E). Cell extracts were collected after 30 minutes and western blotted for total and phosphorylated (p-) proteins. GAPDH, Tubulin and Vinculin served as total loading controls for either phospho or total protein blots as indicated.