To investigate the regulatory effect and mechanism of circular RNA PVT1 (circPVT1) in proliferation and chemoresistance of osteosarcoma (OS) cells.The expression of circPVT1 in human OS and adjacent normal tissues was detected. The correlation between circPVT1 expression and clinical features of OS was analyzed. The expressions of circPVT1 and miR-24-3p in OS cells resistant to cisplatin, doxorubicin or methotrexate and parental OS cells were detected after cell transfection. CCK-8 and colony formation assay assessed the viability and proliferative ability of OS cells. qRT-PCR and Western blotting measured the expression of KLF8. Dual-luciferase reporter and RNA pull-down assays verified the targeting relationships of circPVT1/miR-24-3p and miR-24-3p/KLF8.CircPVT1 was over-expressed in OS tissues and cells, and correlated with clinical features of OS. Over-expressed circPVT1 reduced the survival of OS patients. CircPVT1 was up-regulated in chemoresistant OS cells compared to their parental cells. CircPVT1 inhibition suppressed the proliferation and chemoresistance of OS cells. MiR-24-3p was under-expressed in OS cells and further down-regulated in chemoresistant cells. CircPVT1 could bind and down-regulate miR-24-3p. MiR-24-3p overexpression inhibited the proliferation and chemoresistance of OS cells. KLF8 was over-expressed in OS cells and further up-regulated in chemoresistant cells. MiR-24-3p negatively regulated the expression of KLF8.CircPVT1 mediates proliferation and chemoresistance of OS cells via the miR-24-3p/KLF8 axis. The findings may provide guidance for clinical treatment of OS.© 2022. The Author(s) under exclusive licence to Japan Society of Clinical Oncology.