New indole-linked 1,2,4-triazole derivatives as dual FAK inhibitors and apoptosis inducers targeting survival and migration in triple-negative breast cancer in-vitro

Focal adhesion kinase (FAK) is overexpressed and hyperactivated in Triple-Negative Breast Cancer, driving tumor aggressiveness and Cancer stem cell-mediated therapy resistance. Therefore, targeting FAK signalling represents a promising therapeutic strategy. In this study, a series of indole and bis-indole-1,2,4-triazoles were synthesized and evaluated as anti-TNBC agents targeting FAK. Compounds 3c, 4c, and 5c displayed potent cytotoxicity (IC₅₀ = 41-77 µg/mL) with minimal toxicity to normal cells, outperforming precursor compound 2. Wound-healing assay revealed significant inhibition of cell migration, particularly by 4c. Cell cycle analysis revealed that 4c induced S-phase arrest in MCF-7 cells and G1-phase arrest in MDA-MB-231 cells, accompanied by significant Apoptosis. In MDA-MB-231 cells, 4c triggered extensive total Apoptosis (90.84%) with minimal necrosis. Gene expression studies demonstrated that 4c markedly downregulated PTK2 (FAK), CCL5, and BCL2, while upregulating CASP3, highlighting its dual role as FAK inhibitor and Apoptosis inducer. Importantly, 4c efficiently suppressed FAK protein expression (61.3%) in TNBC, compared to the FAK inhibitor GSK-2256098 (70.7%). In vivo toxicity assessment confirmed good tolerability in mice without profound hepatic or renal impairments, while docking and ADMET analyses confirmed strong FAK binding affinity, and favourable pharmacokinetics of 4c. Collectively, 4c emerges as a promising FAK-targeted candidate for TNBC therapy.