Expression levels of the papain-like cysteine protease cathepsin B (Ctsb) have been positively correlated with mammary tumour progression and metastasis; however, its roles in the hallmark processes of tumour development remain unclear. This study aims to investigate the effects of cathepsin B on tumour cell proliferation and development. The results show that cathepsin B is involved in the regulation of tumour cell proliferation and that its deficiency leads to delayed development of mammary tumours. These findings suggest that cathepsin B plays a crucial role in tumour development and that its inhibition may be a potential therapeutic strategy for the treatment of breast cancer.