STAT3 inhibitor STAT-IN-12 induces apoptosis in mutant-p53 MDA-MB-435 and MDA-MB-231 breast cancer cell lines

Background: p53 restricts tumorigenesis by eliminating damaged cells, whereas constitutively active STAT3 promotes cancer cell survival and proliferation. Their opposing functions, often reinforced by p53 mutation, represent key cooperative drivers of malignant progression. Mutant p53 (mt-p53) further activates STAT3, thereby amplifying tumour aggressiveness. Mutant p53, exhibiting hundreds of different mutant forms and displaying both loss-of-function and acquired oncogenic properties, presents a therapeutic challenge that is difficult to target with classical drugs. Methods and Results: Inhibiting STAT3 by STAT-IN-12 may therefore improve anti-proliferation outcomes and therapeutic efficacy in mt-p53 MDA-MB-231 and MDA-MB-435 in vitro models. Cell viability was determined using the CVDK-8 kit and the xCELLigence real-time cell analysis system (RTCA). Colony formation ability was assessed by clonogenic assay. Intracellular reactive oxygen species (ROS) levels were measured using the DCFDA probe, and mitochondrial membrane potential (MMP-ΔΨm) changes were evaluated with JC-1 dye. Cell migratory ability was assessed by wound healing assay. Cell cycle distribution and apoptosis were analyzed by flow cytometry, while protein expression levels were determined by Immunoblotting in mt-p53 cell lines. In this study, the potential antitumor properties of the novel STAT3 inhibitor STAT3-IN-12 were evaluated against human mt-p53 cell lines. STAT3-IN-12 markedly affected tumour cell viability (IC₅₀ values for the MDA-MB-231 and MDA-MB-435 cell lines at 24 and 48 h were calculated as 31.631 and 19.6274 µM, 41.8782 and 17.1797 µM, respectively), proliferation, redox balance, mitochondrial integrity, and migration. It also induced G0–G1 cell-cycle arrest and promoted apoptosis. Conclusions: Collectively, these findings highlight STAT3-IN-12 as a promising candidate for mt-p53 cells, notably by restoring susceptibility to apoptosis despite the apoptosis resistance driven by mutant p53, a critical factor in therapeutic failure in this cancer subtype.