Cancer is a major worldwide health issue, with rising prevalence and limitations to traditional chemotherapy demanding the development of innovative, selective anticancer drugs. Schiff bases have emerged as promising candidates due to their diverse biological and anticancer activities. The present study aims to emphasize the chemical identity and anticancer activity of (E)-4-(2- hydroxybenzylideneamino)-N-(pyrimidin-2-yl) benzene-sulfonamide; SB1 and 4-(5-bromo-2- hydroxybenzylideneamino)-N-(pyrimidin-2-yl)-benzenesulfonamide); SB2. The structural identity was determined by FTIR 460 PLUS and 1H NMR spectra. The cytotoxicity of SB1 and SB2 were evaluated using MTT against various human cancer cells including HCT116, MCF-7, A549, HepG2, T24 compared to WI-38 cells. Additional further studies on MCF-7 and HCT116 cells, including Annexin V-FITC apoptosis detection, ROS generation, DNA fragmentation, and Topoisomerase I and II inhibition, were carried out to clarify their modes of action. Molecular docking studies were performed to predict how the produced compounds attached to certain target proteins linked to cancer, shedding light on their putative molecular mechanisms of action. The results showed that the IC50 of MCF-7, treated with SB1, exhibited the strongest antiproliferative effect (66.27 ± 1.14 ìg/mL), followed by HCT116 (90.01 ± 0.12 ìg/mL), A549 cells (91.5 ± 2.06 ìg/mL), T24 (192.28 ± 1.12 ìg/mL), and HepG- 2 cells (240.08 ± 3.83 ìg/mL). Furthermore, SB2 had the highest antiproliferative activity of MCF-7 (59.61 ± 1.59 ìg/mL), followed by HCT116 (48.65 ± 2.12 ìg/mL), A549 cells (68.56 ± 1.6 ìg/mL), T24 (216.13 ± 0.7 ìg/mL), and HepG-2 cells (120.53 ± 2.62 ìg/mL). Moreover, SB1 and SB2 cause oxidative stress and apoptosis in cancer cells compared to the control. The molecular docking results showed that the Schiff bases have strong in vitro antineoplastic action and have high binding affinities for key proteins involved in cancer progression. These findings highlight the value of these two compounds of Schiff base derivatives as lead molecules in the development of tailored anticancer drugs.