Canine osteosarcoma (OS) is an aggressive malignancy associated with poor outcomes. Therapeutic improvements are likely to develop from an improved understanding of signalling pathways contributing to OS development and progression. The Wnt signalling pathway is of interest for its role in osteoblast differentiation, its dysregulation in numerous cancer types, and the relative frequency of cytoplasmic accumulation of β-catenin in canine OS. This study aimed to determine the biological impact of inhibiting canonical Wnt signalling in canine OS, by utilizing either β-catenin siRNA or a dominant-negative T-cell factor (TCF) construct. There were no consistent, significant changes in cell line behaviour with either method compared to parental cell lines. Interestingly, β-catenin transcriptional activity was three-fold higher in normal canine primary osteoblasts compared to canine OS cell lines. These results suggest canonical Wnt signalling is minimally active in canine OS and its targeted inhibition is not a relevant therapeutic strategy.