Establishment and characterization of canine mammary tumors 622: A novel luminal B CMT cell line exhibiting partial epithelial-mesenchymal transition and intermediate drug sensitivity.

BACKGROUND AND AIM: Canine mammary tumors (CMTs) serve as valuable comparative models for human breast cancer (HBC) owing to their shared biological and molecular features. However, well-defined cell lines representing the luminal B subtype remain limited. This study aimed to establish and characterize a novel CMT cell line, designated CMT-622, to expand available in vitro models for luminal B breast cancer research. MATERIALS AND METHODS: Primary tumor tissue was collected from an 11-year-old female dog diagnosed with high-grade mammary carcinoma (T3N1M0). Tumor cells were isolated using enzymatic digestion and differential adhesion. Morphological, cytogenetic, and immunophenotypic characteristics were assessed using hematoxylin-eosin staining, immunohistochemistry, and immunofluorescence. Growth kinetics, clonogenicity, and chromosomal analyses were performed, and tumorigenicity was evaluated through xenograft assays in nude mice. Drug sensitivity and apoptosis were compared with two existing CMT lines (CMT-1211 and CMT-n7) using the cell counting kit-8 (CCK)-8 assay and flow cytometry. RESULTS: CMT-622 cells maintained stable proliferation beyond 40 passages with a doubling time of 46.23 h and >15% cloning efficiency. Karyotyping revealed hyperdiploidy (80-110 chromosomes; modal = 87). Immunohistochemistry and immunofluorescence confirmed estrogen receptor (+), progesterone receptor (-), and human epidermal growth factor receptor 2 (weak +) expression, consistent with a luminal B phenotype. Co-expression of cytokeratin-18 and vimentin indicated a partial epithelial-mesenchymal transition (EMT) state. In nude mice, CMT-622 exhibited moderate tumorigenicity and pulmonary metastasis. The line showed intermediate osthole sensitivity (half-maximal inhibitory concentration = 50.48 μM) and an apoptosis rate of 21%, between CMT-1211 and CMT-n7, indicating balanced proliferative and drug-responsive behavior. CONCLUSION: CMT-622 represents a newly established luminal B CMT cell line with stable growth, EMT plasticity, and moderate drug sensitivity, reflecting clinically relevant tumor aggressiveness. Its molecular and phenotypic consistency across in vitro and in vivo models underscores its reliability for translational oncology applications. CMT-622 provides a robust preclinical platform for exploring tumorigenesis, metastasis, and therapeutic responses in both veterinary and HBC contexts, bridging comparative and translational cancer research.