Figure 4

Growth inhibition curves and median-drug effect plots of two NSCLC cell lines treated with cisplatin, pimozide or their simultaneous combination. Exponentially growing SW1573 (A) and H1299 (B) NSCLC cells were seeded in 96-well plates. Twenty four hours after seeding, cells were treated with cisplatin, pimozide or a combination of both drugs. After 72 hours of drug exposure, the sulforhodamine B (SRB) chemosensitivity assay [97] was performed. Briefly, cells were fixed with ice-cold trichloroacetic acid, washed with deionized water, left to dry and stained with SRB dye. After washing with 1% acetic acid, cells were left to dry, and protein-bound dye was dissolved in 10 mM Tris base solution. OD at 492 nm was determined. Growth inhibition curves (upper panels) were generated, where each point represents mean ± SEM of at least 3 replicates. IC₅₀ values, the drug concentration that inhibits the cell growth by 50% (summarized in Table 3), were calculated by fitting the data to a sigmoid dose–response curve using GraphPad Prism. The treatment with the two drugs in combination was carried out using a constant cisplatin:pimozide ratio (as indicated in Table 3), which was established for each cell line as the ratio of the IC₅₀ values for each single drug. The drug concentrations on the X-axis of the growth inhibition curves for the combination treatment refer to cisplatin. In order to evaluate the interaction between cisplatin and pimozide we used the median-drug effect analysis method originally described by Chou and Talalay [95]. Using CalcuSyn, the effect of the combination was compared to the effect of each drug to determine the combination index (C.I.) at a range of Fa (fraction affected by the dose) values. The resulting C.I. values were plotted against the Fa to produce the median-drug effect plot for each cell line (lower panels).