Targeting PI3K/AKT/mTOR Signaling with Novel Andrographolide Analogues Potentially Induces Apoptosis and Inhibits the Metastatic Ability of Human Breast Cancer Cells: Rational Design, Synthesis, and In Vitro Studies

Objective: This study aimed to design and synthesize novel andrographolide derivatives to enhance their pharmacokinetic properties and anticancer efficacy by incorporating lipophilic moieties and triazole rings. Methods: In vitro, cytotoxicity assays were conducted against various breast cancer cell lines to identify potent derivatives. Structure-activity relationship analysis was derived based on in vitro cytotoxicity and in silico ADME properties. In vitro studies were performed to evaluate the mechanisms underlying the cytotoxic and other anticancer capabilities, including morphological assessments, apoptosis assays, caspase assays, and cell migration/invasion assays. Western blot analysis examined the modulation of the PI3K/AKT/mTOR pathway. PI3K kinase inhibition assays and docking studies confirmed the inhibitory effects and binding affinity. Results and Discussion: Compounds (XIf) and (XIi) demonstrated significant cytotoxic potency against breast cancer cell lines, with compound (XIi) showing superior capabilities. Compound (XIi) induced a dose-dependent increase in apoptosis, particularly at 2 × IC₅₀, and activated both intrinsic and extrinsic apoptotic pathways. It also suppresses cell migration and invasion, which are key processes in cancer progression. Western blot analysis revealed significant inhibition of the PI3K/AKT/mTOR pathway by compound (XIi), especially in the presence of estrogen, suggesting potential synergy with tamoxifen in estrogen receptor-positive breast cancer. The PI3K kinase inhibition assay showed dose-dependent inhibition of PI3K-α by compound (XIi), comparable to PI-103 at higher concentrations. Docking studies indicated a strong binding affinity of compound (XIi) to the PI3Kα isoform. Conclusions: The study identified andrographolide derivatives, especially compound (XIi), as promising agents against breast cancer, warranting further exploration of their therapeutic potential.