Expression of Estrogen Receptor Alpha and Ki-67 in Synchronous Canine Mammary Tumors.

Mammary tumors are the most common cancer in female dogs. Many dogs develop synchronous tumors, i.e. tumors occurring simultaneously or within a short time period in the same or different mammary gland. Whether synchronous canine mammary tumors are more biologically similar than random tumors due to shared genetic and environmental factors remains uncertain. Estrogen receptor alpha (ER) and Ki-67 are important biomarkers in human breast cancer but less studied in canine mammary tumors (CMTs). To assess tumor similarity and intratumoral heterogeneity, this study examined ER and Ki-67 expression in 72 synchronous CMTs from 36 dogs using immunohistochemistry (IHC), and the expression of the PAM50 gene set in a subset of tumors. Across all tumors, benign tumors exhibited higher ER Allred scores, whereas malignant tumors had higher Ki-67 indices. Malignant tumors displayed greater intratumoral Ki-67 heterogeneity, while intratumoral ER heterogeneity remained consistent across tumor types. For synchronous tumors, Ki-67 index demonstrated moderate to high concordance across most comparisons, with highest concordance in malignant pairs. In contrast, ER Allred score exhibited a moderate similarity only in pairs with identical histopathological diagnosis. Malignant-benign pairs showed poor similarity for both markers. The poorer intradog correlation of the ER Allred score compared to the Ki‑67 index suggests that ER expression is largely tumor‑specific. Analysis of PAM50 gene expression revealed molecular patterns resembling human breast cancer subtypes across the cohort. Expression of PAM50 genes was highly correlated in benign-benign pairs, while variable in pairs with malignant tumors. These findings highlight the complexity of CMTs, emphasizing the need for individualized tumor evaluation and complete tumor removal in dogs with multiple tumors.