Citrate-functionalised hydroxyapatite mimics the microenvironment of bone metastases and demonstrates the role of citrate in prostate cancer colonization of bone

Prostate cancer (PCa) is the second leading cause of cancer-related mortality in men, with bone representing the predominant metastatic site. Progress in treating bone metastatic disease is hindered by lack of preclinical models that faithfully recapitulate the bone microenvironment while reducing animal use. Biomaterial-based platforms offer a powerful alternative, enabling controlled reconstruction of bone composition, metabolic cues, and tumour-matrix interactions. A defining feature of PCa progression is citrate-centred metabolic reprogramming. While healthy prostate epithelial cells accumulate and secrete citrate, aggressive PCa cells import and oxidise it to sustain growth. Given the citrate-rich nature of prostate tissue and bone, we hypothesised that bone-derived citrate may be exploited by metastatic PCa cells to support bone colonisation. We developed a bone-mimetic platform by functionalising hydroxyapatite nanocrystals with citrate (HA-Nc-Cit) and incorporating them into collagen-based 3D matrices within a microfluidic chip. HA-Nc-Cit were characterised and citrate release quantified. Metastatic PCa cells were analysed for migration, viability, clonogenicity, metabolic reprogramming, and citrate transporter expression. HA-Nc-Cit released physiologically relevant citrate levels. Citrate exposure enhanced migration of androgen-independent PC3 cells and, within collagen type I-enriched matrices, increased clonogenicity, upregulated plasma membrane citrate transporter, suppressed glycolysis, and promoted lactate fermentation and mitochondrial biogenesis, without affecting respiratory chain or lipid metabolism. Citrate buffering supported PC3 clonal survival under acidic stress mimicking tumour acidification. In conclusion, citrate-functionalised HA-Nc promotes bone tropism of aggressive PCa by enhancing migratory potential, modulating tumour metabolism, and buffering extracellular acidification, underscoring the value of biomaterial-based models for studying bone-tumour interactions and guiding therapeutic development. STATEMENT OF SIGNIFICANCE: PCa often spreads to bone, but current models fail to capture the complexity of bone environment, limiting progress in treatment development. In this study, we created a 3D bone-mimicking system by binding citrate, a key bone metabolite, to hydroxyapatite nanocrystals mimicking bone mineral and embedding them in collagen-based matrices. This platform shows how citrate not only fuels PCa cells but also buffers the acidic conditions they create, making bone more prone to tumour growth. Unlike traditional models, such biomaterial-based approach combines mineral chemistry, metabolism, and pH regulation in a controlled setting. This work introduces a tool to study bone-tumour interactions and guide future therapies for metastatic PCa.