Targeting tumor intrinsic metabolic node in pancreatic cancer causes tumor regression, remodels extracellular matrix and sensitizes to anti-PD1 therapy.

Pancreatic ductal adenocarcinoma (PDAC) is considered to be a highly immunosuppressive and heterogenous neoplasm which has extremely poor survival. Despite improved knowledge regarding the genetic background of the tumor and better understanding of the tumor microenvironment, immune checkpoint inhibitor therapy (targeting CTLA-4, PD-1, PD-L1), which has shown effect in other solid tumors, has not been very successful against PDAC. The robust desmoplastic stroma, along with an extensive extracellular matrix (ECM) that is rich in glycosaminoglycans like hyaluronan, plays an integral role in this immune evasion. Hexosamine biosynthesis pathway (HBP), a shunt pathway of glycolysis, is a metabolic node in cancer cells that can promote survival pathways (by regulating pro-oncogenic signaling via O-GlcNAc mediated protein modifications) on one hand and influence the ECM in the tumor by regulating hyaluronan synthesis on the other. The rate-limiting enzyme of the pathway, glutamine-fructose amidotransferase (GFAT1), uses glutamine and fructose 6-phosphate to eventually synthesize UDP-GlcNAc. Our results show that enzymes of HBP including GFAT1 are overexpressed in pancreatic tumors and chronic pancreatitis patient samples. In the current manuscript, we target this glutamine-utilizing enzyme by a small molecule glutamine analog (6-diazo-5-oxo-L-norleucine or DON). DON decreases pro-survival pathways in pancreatic cancer cells and decreases the aggressive self-renewal potential and metastatic ability of tumor cells. Further, treatment with DON results in a decrease in hyaluronan and collagen in the tumor microenvironment, leading to an extensive remodeling of the ECM. Treatment with DON further increases CD68+ activated macrophages in the tumor and increases infiltration of CD8+ cytotoxic T-cells. This, in turn, makes the tumor microenvironment more conducive to cytotoxic CD8+ T-cell infiltration and makes tumors more susceptible to anti-PD1 therapy.