Figure 4.

In spheroids of OvCa cell lines, GW280264X plus cisplatin led to a higher cytotoxic effect in HEY and OVCAR-8 cells compared with cisplatin alone. HEY, SKOV-3, or OVCAR-8 cells were seeded in ULA plates with pre-investigated cell densities of 300, 6500, and 4000 cells, respectively, to reach similar spheroid diameters on the day of treatment. On day four after seeding, cells were treated with the indicated concentrations of cisplatin (cis) or NaCl solvent control. For ADAM17 inhibition, GW280264X was used. GI254023X was applied to inhibit ADAM10 and DMSO was used as the solvent control. Curve fitting was performed using GraphPad Prism and EC₅₀ values were calculated from each curve of DMSO, GI254023X (GI), and GW280264X (GW), setting the response of each individual curve (0 µM cis) to the lower (EC₅₀) baseline value for calculation. (a) Quantification of viable cells by the RealTime-Glo™ Assay 48 h after treatment (means of technical replicates ±SD are displayed). (b) Apoptosis was measured using the Caspase-Glo^® 3/7 reagent. Data of at least three biological replicates were normalized to the control (DMSO, NaCl) and are presented as mean ± SEM. Graphs are displayed until the concentration of cisplatin reached the upper plateau. (c) EC₅₀ values of caspase3/7; (d) drug reduction index (DRI₅₀) at 50% effectiveness: DRI₅₀ represents, e.g., the cisplatin concentration required in combination with GW compared with cisplatin alone (to reach 50% effectiveness): synergistic (DRI₅₀ < 1), additive (DRI₅₀ = 1), and antagonistic (DRI₅₀ > 1) effects. ns: not significant, *** p < 0.001, **** p < 0.0001.