Design, synthesis, in vitro cytotoxic activity, and in silico studies of symmetrical chlorophenylamino-s-triazine derivatives

Abstract

Twenty symmetrical chlorophenylamino-s-triazine derivatives were designed and synthesized by reflux (RF) and microwave-assisted (MW) methods. The MW method achieved superior yields (88–95%) in less time (15–30 min) compared to RF (78–86%, 12–24 h), particularly for 3-Cl and 3,4-diCl derivatives with piperidine or diethylamine, due to rapid, uniform heating, enhancing nucleophilic substitution and minimizing side reactions. In particular, compounds 2c (IC₅₀ = 4.14 μM for MCF7, 7.87 μM for C26), 3c (IC₅₀ = 4.98 μM for MCF7, 3.05 μM for C26), and 4c (IC₅₀ = 6.85 μM for MCF7, 1.71 μM for C26) exhibited potent cytotoxic activity with 4c (2,4-diCl, pyrrolidine) surpassing paclitaxel (IC₅₀ = 2.30 μM for C26), and 3c (3,4-diCl, pyrrolidine) rivaling global analogs. Compounds 2f (IC₅₀ = 11.02 μM for MCF7, 4.62 μM for C26) and 3f (IC₅₀ = 5.11 μM for MCF7, 7.10 μM for C26) also showed strong cytotoxicity. QSAR analysis revealed that electron-withdrawing groups (chloro, dichloro) and pyrrolidine enhance C26 potency via improved lipophilicity and π–π stacking, outperforming piperazine and morpholine. Pharmacokinetically, 2c, 3c, and 4c matched Bimiralisib's absorption profiles, surpassing Gefitinib, with 4c showing superior metabolic stability. Compounds 2f and 3c emerged as promising multi-targeted kinase inhibitors, with binding affinities (−7.8 to −9.1 kcal mol⁻¹) closely rivaling Gefitinib, Pazopanib, and Bimiralisib for EGFR, VEGFR2, and PI3K, driven by balanced polar and hydrophobic interactions. Therefore, these findings underscore the potential of 2f and 3c as multi-targeted kinase inhibitors, warranting further mechanistic studies and structural optimization to enhance MCF7 efficacy and reduce toxicity for clinical advancement.