Identification of a Prostate Cancer Cell Proteinase Activated Receptor/MMP Signaling Axis

Abstract

HypothesisProteinase Activated Receptors (PARs) are G‐Protein‐Coupled Receptors activated by the proteolytic unmasking of a cryptic tethered ligand. We proposed that prostate cancer cells produce PAR‐regulating enzymes.MethodsWe monitored PC3, DU145 and LNCaP cell supernatant cleavage of PARs in intact cells by fluorescence imaging using N‐ and C‐ terminus dual‐fluorophore‐tagged PARs. Loss of the N‐terminal tag denotes receptor cleavage‐activation. We also used N‐terminal nano‐luciferase‐tagged PARs that, via luciferase release, document PAR‐cleaving proteinase activity in cell‐derived supernatants. MAPKinase and transwell migration assays were used to monitor PAR activation in PC3 cells.ResultsActivation of PARs 1 & 2 triggered MAPKinase signaling and cancer cell migration. Distinct PAR‐cleaving proteinases in conditioned media from all three prostate cancer cell lines were observed with the N‐Luc‐Luciferase assay, with differential inhibition by matrix metalloproteinase inhibitors and soya trypsin inhibitor. Further, when expressed in PC3 cells, the dual‐fluorophore‐tagged PAR1 was seen in a constitutively cleaved‐active state, with an absent N‐terminus.ConclusionOur data show that prostate cancer‐derived cells secrete PAR‐regulating proteinases that, via an autocrine process, can regulate PAR function in the tumour microenvironment. Since PAR activation triggers prostate cell migration and MAPKinase, PAR antagonists and inhibitors of PAR‐activating proteinases represent therapeutic targets for treating prostate cancer.FundingProstate Cancer Canada Movember Discovery grant.