The Presence of SFRP1 Reduces the High Risk of Metastasis in RANKL-Expressing Canine Mammary Cancers

Canine mammary cancers (CMCs) are one of the most prevalent types of neoplasm in dogs, are frequently malignant, and display high tumour heterogeneity, making evaluating prognosis and predicting successful treatment outcomes difficult. In a previous pilot study, overexpression of the Wnt pathway-associated protein SFRP1 was found to correlate with negative metastasis status in CMCs at both mRNA and protein levels. To establish SFRP1 as a potential biomarker for CMC progression, additional verification of these results in an independent dataset is required, as well as an investigation as to whether SFRP1 expression in CMCs is associated with altered Wnt- or RANKL signalling pathways. In an independent verification cohort of 122 cases of archival CMC FFPE material, expression of SFRP1 was assessed by RT-qPCR and immunohistochemistry. The same tumours were further assessed for RANKL, phosphoROCK2, and NFkB-p65 protein expression. Our data verified that SFRP1 mRNA (p = 0.025) was negatively associated with metastasis status; however, differences in protein expression did not reach statistical significance (p = 0.139). Neither did SFRP1 significantly correlate with expression of any of the other proteins tested. Instead, a strong association was found for RANKL positivity with increased metastasis status (p < 0.001). Co-expression of SFRP1 significantly lowered the higher risk of metastatic spread when compared to RANKLpos/SFRP1neg CMCs (p = 0.033). Noticeably, all vascular-invasive cell clusters observed in tissue section vessels stained positive for RANKL. Our study identified RANKL expression as a strong marker for cancer progression with a strong link to vascular-invasive cells. However, SFRP1 expression may potentially suppress the pro-metastatic nature of RANKLpos CMCs.