Pyrazole-triazole hybrids as kinase-triad inhibitors: a triple-target strategy for synergistic anticancer therapy

The ongoing issue of drug resistance and the lack of specificity in existing cancer treatments highlight the necessity for innovative multi-target agents. This study presents the design and synthesis of two series of pyrazolyl-1,2,3-triazole hybrids: pyrazole-1,2,3-triazole series (7a-i) and pyrazole-di(1,2,3-triazole) series (10a-i). These hybrids incorporate strategic substitutions aimed at targeting the ATP-binding sites of EGFR, VEGFR-2, and AURKA. Antiproliferative screening against MCF-7 (breast) and A549 (lung) cancer cell lines demonstrated that di(1,2,3-triazole) derivatives exhibited increased activity in MCF-7 (IC₅₀: 18.7-21.9 μM), while mono-derivatives, particularly 7i, displayed greater potency in A549 (IC₅₀: 3.56 μM). Flow cytometry revealed S-phase arrest in A549 cells induced by 7i and 10a, while the clonogenic assay validated reduced colony formation and elevated cell mortality. The combination treatment involving doxorubicin demonstrated synergistic effects, with 7i/Dox IC₅₀ at 0.29 μM and 10a/Dox IC₅₀ at 0.95 μM. Biochemical markers such as Ki-67, PCNA, p21, Bax, Bcl-2, caspase-3, VEGF, P-gp, and E-cadherin exhibited improved antiproliferative, pro-apoptotic, anti-angiogenic, and anti-metastatic effects when subjected to combination therapy. Enzyme assays demonstrated low-nanomolar inhibition by 7i (EGFR = 73 nM; VEGFR-2 = 176 nM; AURKA = 89 nM) and moderate potency for 10a. Molecular docking confirmed essential interactions in the active site of each kinase, while in silico ADME profiling indicated favorable drug-likeness, especially for series 7. These findings identify compounds 7i and 10a as potential dual-scaffold leads for subsequent preclinical development as multi-target anticancer agents.