Immunohistochemical Investigation into Protein Expression Patterns of FOXO4, IRF8 and LEF1 in Canine Osteosarcoma

Simple Summary There have been limited advances in the diagnosis, prognosis, and treatment of both canine and human osteosarcoma (OSA), the most common type of primary bone cancer. OSA has an aggressive nature, with incidence rates ranging from 13.9 to 27.2 cases per 100,000 dogs, yet there have been limited advances in patient outcomes in recent decades. Recent developments have identified similarities between human and canine OSA; therefore, researching naturally occurring canine bone cancer may help inform research into OSA in people. The present research investigated three proteins, FOXO4, IRF8, and LEF1, to visualise their expression in OSA tissue. This research helps us understand where the proteins are being expressed in the tumours, which genetic pathways are changing, and may help us identify potentially informative diagnostic, prognostic, and treatment avenues for this cancer in dogs and people.Abstract Osteosarcoma (OSA) is the most common type of primary bone malignancy in people and dogs. Our previous molecular comparisons of canine OSA against healthy bone resulted in the identification of differentially expressed protein-expressing genes (forkhead box protein O4 (FOXO4), interferon regulatory factor 8 (IRF8), and lymphoid enhancer binding factor 1 (LEF1)). Immunohistochemistry (IHC) and H-scoring provided semi-quantitative assessment of nuclear and cytoplasmic staining alongside qualitative data to contextualise staining (n = 26 patients). FOXO4 was expressed predominantly in the cytoplasm with significantly lower nuclear H-scores. IRF8 H-scores ranged from 0 to 3 throughout the cohort in the nucleus and cytoplasm. LEF1 was expressed in all patients with significantly lower cytoplasmic staining compared to nuclear. No sex or anatomical location differences were observed. While reduced levels of FOXO4 might indicate malignancy, the weak or absent protein expression limits its primary use as diagnostic tumour marker. IRF8 and LEF1 have more potential for prognostic and diagnostic uses and facilitate further understanding of their roles within their respective molecular pathways, including Wnt/beta-catenin/LEF1 signalling and differential regulation of tumour suppressor genes. Deeper understanding of the mechanisms involved in OSA are essential contributions towards the development of novel diagnostic, prognostic, and treatment options in human and veterinary medicine contexts.