Hypoxia has been shown to be related to osteosarcoma development and progression. Pseudogene MSTO2P was reported to be dysregulated in hepatocellular carcinoma and lung cancer. However, the mechanism by which MSTO2P-modulated osteosarcoma remains unclear. MSTO2P and PD-L1 expression levels were examined by RT-qPCR and westernblot. Tumor cell invasion was determined by tranwell assay. EMT process was probed by determining E-cadherin and Vimentin levels. Soft agar assay was used to examine anchorage-independent growth of osteosarcoma cells. In vivo tumor growth was measured by xenografting tumor experiment. Hypoxia treatment promoted cell growth, invasion and EMT of osteosarcoma cells. MSTO2P knockdown led to attenuated cell growth, invasion and EMT of osteosarcoma cells under hypoxia condition. More interestingly, our data revealed that MSTO2P was positively associated with tumor growth in immunodeficient mice and human clinical tissues. PD-L1 was shown to act as a key effector for MSTO2P-regulated osteosarcoma progression under hypoxia condition. In conclusion, we unravel a novel mechanism for explaining MSTO2P-involved osteosarcoma progression under hypoxia condition, which will facilitate development of potential diagnostic and therapeutical strategies for osteosarcoma. (c) 2020 Elsevier Inc. All rights reserved.