Induction of potent preferential cell death, severe DNA damage and p53‑independent ROS‑mediated mitochondrial apoptosis by CaTiO3NPs in HNO‑97 tongue cancer cells

Abstract

The aggressive nature and high recurrence rate of tongue cancer, along with the severe side efects associated with current conventional treatments, underscore the urgent need for alternative therapeutic strategies that ofer improved selectivity and reduced toxicity. Calcium titanate nanoparticles (CaTiO3NPs) have previously demonstrated signifcant cytotoxic efects against MCF-7 breast cancer and A549 non-small cell lung cancer lines. However, their therapeutic potential against tongue cancer remains unexplored. Therefore, this study investigates, for the frst time, the cytotoxic efect of CaTiO3NPs on HNO- 97 tongue cancer cells in comparison to normal human skin fbroblasts (HSF) cells. Using various assays, we assessed their efects on cell viability, reactive oxygen species (ROS) generation, genomic DNA and mitochondrial integrity, and cell death pathways. Cytotoxicity assessment using the sulforhodamine B (SRB) assay demonstrated that CaTiO3NPs signifcantly reduced HNO-97 cell viability in a concentration-dependent manner, with an IC50 of 29.67 µg/ml. In contrast, normal HSF cells exhibited a much higher IC50 of 262.6 µg/ml, indicating strong selectivity for cancer cells (selectivity index=8.85). Mechanistic studies revealed that HNO-97 cell exposure CaTiO3NPs at the IC50 concentration markedly increased ROS production, inducing oxidative stress. This led to signifcant genomic DNA damage, confrmed by the comet assay, and mitochondrial dysfunction, validated through Rhodamine-123 staining. Flow cytometry analysis indicated the induction of both apoptotic and necrotic pathways. At the molecular level, qRT-PCR analysis revealed signifcant downregulation of key genes, including the pro-apoptotic p53, the anti-apoptotic Bcl-2, and the mitochondrial ND3 gene, supporting the involvement of mitochondrial dysfunction and impaired DNA repair mechanisms in the observed cytotoxic efects. Conclusion: Collectively, these fndings demonstrate that CaTiO3NPs elicit strong and selective multi-modal cytotoxic efects against HNO-97 tongue cancer cells and minimal impact on normal HSF cells, supporting their promise as a novel nanotherapeutic agent for tongue cancer. Further in vivo studies are needed to validate their therapeutic efcacy, clinical applicability and biosafety.