Unveiling the role of bone microenvironment in bone cancer progression

Bone represents a highly supportive environment for the development of primary and secondary malignancies, but ts microenvironment remains difficult to study due to its complexity. Advanced 3D microfluidic systems offer a promising strategy to mimic this complexity, but models capable of fully recapitulating bone physiology are still limited. This project investigates how cellular and acellular elements of bone contribute to the initiation and progression of bone tumors, focusing on the role of citrate in osteotropism, clonogenicity, and metabolic adaptation of bone metastases, as well as the paracrine interactions between stromal cells and neurons in osteosarcoma. A complementary goal is the development of a Bone-on-Chip platform that mimics the architecture and signaling of the Bone Remodeling Unit. Using 3D microfluidic models enriched with osteomimetic matrix composed of hydroxyapatite nanoparticles functionalized with citrate, we demonstrate that citrate enhances prostate cancer cell osteotropism and clonogenicity, promotes mitochondrial activation, and protects tumor cells from microenvironmental acidosis driven by glycolytic metabolism. Isolation of Cancer Associated Fibroblasts from human bone metastasis biopsies revealed a predominance of myofibroblastic CAFs expressing αSMA and depositing abundant ECM, with fibronectin exceeding type-I collagen. In osteosarcoma, we identified nerve fibers within the tumor microenvironment, colocalizing with stromal cells and correlating with Ki-67 expression. Mesenchymal stromal cells promoted axonogenesis through IL‑6 and BDNF, while recruited axons enhanced tumor proliferation and survival; these interactions intensified under intratumoral acidosis. Finally, a custom Bone-on-Chip device was engineered to model the BMU, enabling co-culture of osteoblasts, osteoclasts, and osteocytes and reproducing their paracrine communication. Overall, this work deepens understanding of microenvironmental drivers of bone malignancies and underscores the need for physiologically relevant models to uncover therapeutic vulnerabilities.