# Identification and Characterisation of Canine Osteosarcoma Biomarkers and Therapeutic Targets

## TL;DR

This study identifies potential biomarkers and drug targets for canine osteosarcoma, a deadly bone cancer in dogs, using protein expression analysis and genetic data.

## Contribution

The study identifies novel canine osteosarcoma biomarkers and explores their potential as therapeutic targets.

## Key findings

- GPR64, TOX3, MMP-12, and FOXF1 are overexpressed in canine osteosarcoma tissues.
- Differential sex expression was observed for GPR64 and TOX3.
- Potential diagnostic biomarkers and therapeutic targets were identified.

## Abstract

Osteosarcoma is a bone cancer that is commonly observed in dogs and also affects people. The cancer often spreads, metastasizes, before the original cancer is even diagnosed, making treatment more difficult, less efficient, and resulting in more patient deaths. Current therapeutic protocols, in both dogs and people, produce low survival rates, and hence new therapeutic regimens are required. Studying tumour tissue is the ‘gold standard’ technique for diagnosing many cancer types. In medical research, techniques including immunohistochemistry, which studies patient tumour specimens, help us understand how much protein cancer expresses and show its location within the tumour. The proteins investigated are termed ‘biomarkers’ if they help detect and monitor the disease, and can indicate whether treatments work or enable us to understand the disease better. The use of these techniques in veterinary medicine has not been implemented for osteosarcoma, as biomarkers have not yet been identified. This study, therefore, investigated the proteins from genes that were overexpressed in canine osteosarcoma tissue. This has helped us to understand bone cancer in dogs better and identify potential drug targets and diagnostic biomarkers.

Background: Osteosarcoma (OSA) is the most common type of bone cancer in canines. Novel therapies are required to prevent the growth, survival, and metastatic progression of this cancer, to increase life expectancy of patients. Immunohistochemical (IHC) studies and RNA sequencing help us gain a deeper understanding into the molecular mechanisms of the disease. Methods: We previously compared canine OSA tissues with patient matched non-tumour tissues, revealing 442 overexpressed genes within the samples. The present research used IHC staining for four of these genes in OSA tissues: G protein-coupled receptor 64 (GPR64), TOX High Mobility Group Box Family Member 3 (TOX3), Matrix Metallopeptidase 12 (MMP-12), and Forkhead Box F1 (FOXF1). H-scoring was performed to quantitatively assess protein expression and qualitatively contextualise staining locations. Additional analyses addressed whether gender or anatomical location of lesions (axial or appendicular tumours) affected protein expression. cBioPortal was employed to analyse expression and genetic alterations in patients. Results: GPR64, TOX3, MMP-12, and FOXF1 showed high mRNA expression and genetic alterations in people with OSA. GPR64, TOX3, MMP-12, and FOXF1 were all expressed in canine OSA with novel findings regarding cellular expression. Additionally, differential sex expression was revealed for GPR64 and TOX3. Potential biomarkers or therapeutic targets were identified. Conclusions: These studies, and subsequent analysis, have provided insights into the molecular mechanisms associated with OSA progression and revealed potential biomarkers for diagnostic and prognostic purposes. A deeper understanding of genetic and protein interactions will support and progress novel pathways towards diagnostic, prognostic, and treatment interventions for OSA in both veterinary and human medicine.