Introduction: Leukemia is one of the types of cancer that its most common treatments, including radiation and chemotherapy, are associated with many problems and side effects such as drug resistance. Therefore, researchers are looking for multi-factor combined therapies with fewer side effects and more anti-cancer effects to treat leukemia. The aim of this study was to analyze the combination effect of gold nanoparticles (Au NPs) and hyperthermia on the proliferation of chronic myeloid leukemia cells.Materials and Methods: To study the nanoparticle absorption by K۵۶۲ cancer cells, various amounts of nanoparticles were used. The MTT and apoptosis assays were used to evaluate the anti-proliferation properties of NPs (۲۰, ۴۰, and ۸۰ μg/ml) and hyperthermal (۴۱ °C for an hour) on cancer cells.Results: Our results revealed a significant decrease in the proliferation index (۴۳ ± ۳.۳۱۱, p<۰.۰۱) and an increase in apoptosis (۵۴ ± ۳.۶۰۵, p<۰.۰۱) of K۵۶۲ cells in the group receiving a combination of ۸۰ µg/ml of Au NPs and hyperthermia compared to the control and other treatment groups.Conclusions: According to the results, the combination of Au NPs and hyperthermia could significantly suppress the proliferation and induce apoptosis in K۵۶۲ cells in comparison with Au NPs or hyperthermia, alone. Therefore, it confirms that the application of several anti-cancer strategies using NPs and hyperthermia is a useful strategy in cancer treatment.