UNLABELLED: In pancreatic ductal adenocarcinoma cancer (PDAC) drug resistance is a severe clinical problem and patients relapse within a few months after receiving the standard-of-care chemotherapy. One contributing factor to treatment resistance is the desmoplastic nature of PDAC
the tumours are surrounded by thick layers of stroma composing up to 90% of the tumour mass. This stroma, which is mostly comprised of extracellular matrix (ECM) proteins, is secreted by cancer-associated fibroblasts (CAFs) residing in the tumour microenvironment. However, the mechanistic basis by which the tumour stroma directly contributes to chemoresistance remains unclear. Here, we show that CAF-secreted ECM proteins induce chemoresistance by blunting chemotherapy-induced DNA damage. Mechanistically, we identify N-myc downstream regulated gene 1 (NDRG1) as a key protein required for stroma-induced chemoresistance that responds to signals from the ECM and adhesion receptors. We further show that NDRG1 is a novel DNA repair protein that physically interacts with replication forks, maintains DNA replication and functions to resolve stalled forks caused by chemotherapy. More specifically, NDRG1 reduces R-loops, RNA-DNA hybrids that are known to cause genomic instability. R-loops occur during replication-transcription conflicts in S-phase and after chemotherapy treatments, thus posing a major threat to normal replication fork homeostasis. We identify NDRG1 as highly expressed in PDAC tumours, and its high expression correlates with chemoresistance and poor disease-specific survival. Importantly, knock-out of NDRG1 or inhibition of its phosphorylation restores chemotherapy-induced DNA damage and resensitizes tumour cells to treatment. In conclusion, our data reveal an unexpected role for CAF-secreted ECM proteins in enhancing DNA repair via NDRG1, a novel DNA repair protein, directly linking tumour stroma to replication fork homeostasis and R-loop biology, with important therapeutic implications for restoring DNA damage response pathways in pancreatic cancer. SUMMARY PARAGRAPH: Drug resistance is a severe clinical problem in stroma-rich tumours, such as pancreatic ductal adenocarcinoma (PDAC), and patients often relapse within a few months on chemotherapy