Decellularized lung biomatrix models to investigate microenvironmental regulation of organ-selective tropism in metastatic breast cancer

Background: Breast cancer is the most frequently diagnosed malignancy and a leading cause of cancer-related death in women. Metastatic disease is the main cause of breast cancer mortality, with lung as a key metastatic organ. Although a variety of tumor models, ranging from conventional two-dimensional (2D) systems to in vivo models, have been used to study breast cancer lung metastasis, they do not fully recapitulate lung specific microenvironment. Methods: To study the role and impact of organ microenvironment on the tropism of breast cancer cells, we used parental and lung-metastatic variants of MDA-MB-231cells by seeding the various cell densities (25 k, 15 k, 5 k, 2.5 k, 1 k, and 0.5 k cell/cm²) on decellularized lung matrices. The decellularized lung biomatrix scaffolds were harvested from rats using perfusion protocol. To assess cell activity, ATP-based luminescence assays were performed for all conditions and normalized to the corresponding control condition (cells cultured in the absence of lung biomatrix). Results: Normalized ATP measurements showed that decellularized lung biomatrix differentially modulated the metabolic activity of parental and lung-metastatic MDA-MB-231 cells. Across almost all seeding densities and biomatrix concentrations, the lung-metastatic variant generally exhibited higher growth velocity than the parental line, suggesting that the lung-derived matrix provides a more favorable microenvironment for the metastatic cells through enhanced cell-matrix interactions that activate proliferative signaling pathways. This difference was most evident at higher cell seeding densities and higher protein concentrations of decellularized lung biomatrix. At 25 k, 15 k, and 5 k cells/cm², the lung-metastatic cells displayed a clear increase in normalized metabolic activity relative to the parental cells at 200 and 300 µg/cm². In contrast, at cell densities of 2.5 k, 1 k, and 0.5 k cells/cm², normalized ATP values for parental and metastatic cells were comparable across all matrix protein densities within 5 days culture period. Ongoing work aims to further characterize cellular activities and cell-matrix signaling pathways in the lung-mimetic scaffolds. Conclusion: Our ex vivo model based on decellularized lung biomatrix scaffolds demonstrates that lung specific microenvironmental cues selectively enhance the metabolic activity of lung-metastatic breast cancer cells relative to parental cells, supporting microenvironmental regulation of organ specific metastatic preference.