BACKGROUND Palbociclib, a specific inhibitor of CDK4/6, has been shown to provide a survival benefit in hormone receptor-positive advanced breast cancer; however, its resistance and related mechanisms are unclear. MATERIAL AND METHODS In this study, we constructed palbociclib-resistant hormone receptor-positive breast cancer cells (MCF-7-P) via culturing with palbociclib for at least 6 months. Quantitative real-time PCR (qRT-PCR) and western blot were used to detect the expression of stemness markers in MCF-7-P and MCF-7 cells. Additionally, cell spheroid formation, Transwell migration, ALDH1 activity, and flow cytometry assays were performed to detect stemness and migration ability of MCF-7-P cells, and the effects of everolimus on MCF-7-P cells stemness and migration ability. Growth inhibition assay was used to examine the effect of everolimus on the sensitivity of palbociclib in MCF-7-P and MCF-7 cells. RESULTS MCF-7-P cells had stronger stemness and higher expression of ABCG2 and MDR1. Moreover, PI3K/Akt/mTOR signaling was hyper-activated in MCF-7-P cells. Additionally, everolimus, which is a mTOR inhibitor, attenuated MCF-7-P cells stemness and re-sensitized MCF-7-P cells to palbociclib. Importantly, everolimus enhanced the antitumor effect of palbociclib in palbociclib-sensitive hormone receptor-positive cells (MCF-7 cells). CONCLUSIONS These findings provide a rationale for future clinical trials of palbociclib and everolimus combination-based therapy in hormone receptor-positive breast cancer.