Gallic Acid Enhances Carboplatin-Induced Antitumoral Responses in Cervical Cancer Cells Through Oxidative Stress-Associated Mitochondrial and Apoptotic Mechanisms

Background/Objectives: Gallic acid (GA) is a naturally occurring polyphenol with reported antioxidant and anticancer properties. This study investigated whether GA enhances carboplatin (CARB)-associated anticancer activity in HeLa cervical cancer cells through mechanisms related to oxidative stress, mitochondrial dysfunction, apoptosis, and cell cycle dysregulation, while comparatively evaluating cytotoxicity in HaCaT cells. Methods: The effects of GA and CARB, individually and in combination, were evaluated using cell viability assays, apoptosis and cell cycle analyses, intracellular reactive oxygen species (ROS) measurements, N-acetylcysteine (NAC)-mediated rescue experiments, mitochondrial membrane potential assessment, reverse transcription–quantitative polymerase chain reaction (RT-qPCR), immunocytochemistry, and three-dimensional (3D) tumor spheroid models. Bioinformatic analyses were performed to explore pathways associated with the observed molecular responses. Results: The GA + CARB combination demonstrated enhanced cytotoxicity and apoptotic activity in HeLa cells compared with either monotherapy, while exhibiting comparatively lower toxicity in HaCaT cells. Combination treatment increased intracellular ROS levels, whereas NAC pretreatment partially reversed ROS accumulation and cytotoxicity, supporting a contributory role of oxidative stress in treatment-associated responses. The combination also induced mitochondrial membrane depolarization, increased G2/M arrest and SubG1 accumulation, and modulated apoptosis- and cell cycle-related gene expression. In 3D spheroid models, GA + CARB reduced spheroid growth and viability and disrupted spheroid integrity more effectively than single-agent treatments. Bioinformatic analyses identified interconnected pathways associated with oxidative stress, apoptosis, and cell cycle regulation. Conclusions: GA may enhance CARB-associated anticancer activity through mechanisms linked to oxidative stress, mitochondrial dysfunction, apoptosis, and cell cycle dysregulation. The incorporation of ROS/NAC rescue experiments and 3D spheroid validation further supports the biological relevance of the observed effects. Nevertheless, these findings remain preliminary and require confirmation in advanced in vivo and translational cervical cancer models.