Abstract: Background and Objective: Baicalein is an effective anti-cancer treatment of several cancer types. Although numerous studies on baicalein efficacy have been published, the specific mechanism of its cancer inhibition remains unclear. Present study was aimed to provide detailed molecular targets and signaling pathways that baicalein regulated, which may contribute to better application of baicalein in osteosarcoma cure. Materials and Methods: In this study, after treatment of human osteosarcoma cells lines SaOS-2 with baicalein, the CCK-8 assay was used to determine cell viability. The RNA sequencing was used to examine whole transcriptome variation and analyzed by Gene ontology, Kyoto Encyclopedia of Genes and Genomes and Hierarchical clustering. Apoptosis was studied by flow cytometry. Wound healing, adhesion, ATP assays followed by evaluating the expression of mRNA and protein of COL1A2, COL6A1, LAMA5, ITGA4, ITGB1, MMP2 and BINP3 via quantitative real-time PCR and western blot respectively. Results: The dose of 25 μg mL–1 of baicalein inhibited the cells proliferation in time dependent manner. The extracellular matrix receptors including the integrin (ITGA4 and ITGB1), collagen (COL1A2, COL6A1) and laminin (LAMA5) were significantly repressed in challenged cells groups at messenger RNA and protein expression level. Furthermore, the migration, adhesion as well as ATP level were statically reduced, whereas cell apoptosis was stimulated. Conclusion: The result supported that baicalein promotes the SaOS-2 apoptosis process. This study suggests that baicalein may be used alone or with other anti-cancer treatments to reverse ECM receptor-associated malignant phenotype of human cancers.