Prostate cancer (PCa) bone metastases are associated with mixed osteolytic/osteoblastic lesions. Identifying novel mechanisms that underlie this debilitating bone disease will ultimately reveal new therapeutic targets. Here, we identified differential expression of the key osteoblastic enzyme alkaline phosphatase (ALP) in a panel of PCa cell lines with increasing metastatic potential and aimed to elucidate ALP function and regulation in PCa in vitro and in vivo. ALP activity and mRNA expression were higher in osteoblastic ARCaPM cells than non-metastatic ARCaPE (p<0.001), and associated with in vitro mineralisation of PCa cells, indicative of functional osteomimicry. ALP expression was lower in osteolytic PC3 cells, and undetectable in most non-bone metastatic cell lines. ALP knockdown in ARCaPM cells resulted in ~50% decrease in cell viability and Mesenchymal-Epithelial Transition (MET), evidenced by a change in morphology, increased E-cadherin (p<0.001) and decreased migration. ALP knockdown in PC3 cells decreased cell viability (p<0.01). Pharmacological inhibition of ALP dose-dependently inhibited cell viability in a panel of bone metastatic PCa cells. Knockdown of the mesenchymal marker Snail, or treatment with extracellular ATP, a known growth inhibitor in PCa, resulted in ~40% decrease in ALP activity and expression (p<0.05) and induced MET, evidenced by a morphology change and altered expression of epithelial and mesenchymal markers. To investigate tumour-derived ALP in vivo, we analysed human primary and metastatic PCa samples from the MSKCC PCa dataset, demonstrating that ALP correlated positively with EMT markers (including Snail, ρ=0.5827, r=0.5120) and negatively with markers of tumour suppression (including PTEN, ρ=-0.5487, r=-0.3338) (p<0.001). Our study defines a novel mechanism whereby tumour-derived ALP induces EMT and survival of PCa cells, with ALP expression regulated by Snail and purinergic signalling. Correlation of ALP expression with known cancer genes demonstrates its importance in human disease. Hence, this study identifies a previously unsuspected role for tumour-derived ALP in PCa.
Disclosure: This work was supported by the PRO-NEST consortium (FP7 programme), the Oxford Cancer Research Centre and the University of Oxford Medical Research Fund.