Berberine (BBR) is a compound derived from Chinese herbal medicine, known for its anticancer properties through multiple signaling pathways. However, whether BBR can inhibit tumor growth by participating in ferroptosis remains unconfirmed. In this study, we demonstrated that berberine synergistically inhibited NSCLC in combination with multiple ferroptosis inducers, and this combination synergistically down-regulated the mRNA and protein expression of SLC7A11, GPX4, and NRF2, resulting in ferroptosis accompanied by significant depletion of GSH, and aberrant accumulation of reactive oxygen species and malondialdehyde. In a lung cancer allograft model, the combination treatment exhibited enhanced anticancer effects compared to using either drug alone. Notably, p53 is critical in determining the ferroptosis sensitivity. We found that the combination treatment did not elicit a synergistic anticancer effect in cells with a p53 mutation or with exogenous expression of mutant p53. These findings provide insight into the mechanism by which combination induces ferroptosis and the regulatory role of p53 in this process. It may guide the development of new strategies for treating NSCLC, offering great medical potential for personal diagnosis and treatment.