Cathepsin D expression in pancreatic ductal adenocarcinoma (PDAC) cells increases proliferation and reduces survival of pancreatic cancer patients

AIM: A remodelling of the pancreatic cancer tumour microenvironment and extracellular matrix (ECM) has been implicated in the prognosis of PDAC. We investigated whether the aspartic protease cathepsin D contributes to proliferation, migration and fibrogenesis of pancreatic cancer cells and its expression affects the survival of pancreatic cancer patients. MATERIALS & METHODS: Mouse pancreatic stellate cells (PSC) were isolated by Nycodenz gradient. PSC and murine pancreatic cancer PD6606 cells were used for Western blotting and immunofluorescence staining. Cathepsin D activity was inhibited with pepstatin and its expression silenced with siRNA. TGFβ1 was measured using microplate sandwich ELISA and proliferation by standard MTT assay. Tumour tissue microarrays of 404 patients from the ESPAC-3 trial (all after PDAC-resection) were stained with anti-cathepsin D (C-20) antibody and patients with the highest quartile H-score (cut-off 22.35) were compared with those in the lowest quartile using Kaplan-Meier curves, log-rank tests, and Cox proportional hazards models. All statistical tests were two-sided. RESULTS: Inhibition of cathepsin D in ductal adenocarcinoma cells resulted in significantly reduced proliferation and migration. Cathepsin D expression increased in parallel with PSC trans-differentiation to myofibroblasts. Silencing (siRNA) or inhibiting cathepsin D decreased PSC proliferation by 20-30%. There was a time-dependent activation of latent TGF β1 and cathepsin D cleaved latent TGF β1, leading to its activation. Inhibition of cathepsin D decreased activated TGF β1. In 1824 tissue microarray cores from 382 patients (94.6%) median overall patients' survival was 23.85 months (95% CI 21.0-26.2). Median survival for patients in the highest quartile cathepsin D H-score was 21.2 months (95% CI 17.3-24.8) and in the lowest quartile 27.2 months (95% CI 24.1-31.2) χLR,1DF = 4.33; P = 0.0374. DISCUSSION: In pancreatic cancer, cathepsin D expression not only regulates fibrogenesis and stromal development but also ductal adenocarcinoma cell proliferation and migration. These mechanisms make cathepsin D a driver of pancreatic cancer growth and account for the poor prognosis of patients with high cathepsin D expression in PDAC.