NANOSTRING GEOMX SPATIAL TRANSCRIPTOMIC ANALYSIS OF PANCREATIC ISLETS IN DIABETIC AND NONDIABETIC PANCREATIC CANCER PATIENTS REVEALS BETA CELL DEDIFFERENTIATION ASSOCIATED WITH ER STRESS PATHWAYS AND ALTERED LIPID METABOLISM

Background: Pancreatic ductal adenocarcinoma (PC) is a deadly cancer with a rapidly rising incidence and a low 5-year relative survival rate. A complex relationship exists between PC and diabetes, as diabetes is both a risk factor for, and a consequence of PC, the latter exemplified by the clinical observation that new-onset diabetes after age 50 can be an early sign of PC. PC patients with diabetes have poorer outcomes. However, the mechanistic links between PC and diabetes are not known. Here, we use spatial transcriptomics to obtain baseline information about tumor islet cell function to guide future mechanistic studies. Methods: We performed GeoMx Spatial Whole Transcriptome Assays (WTA) on pancreatic tumors resected from PC patients with and without diabetes. GeoMx selectively extracted signals from insulin⁺ (β cells), glucagon⁺ (α cells), and pan-cytokeratin⁺ positive (ductal/neoplastic) cells in selected tumor regions to derive transcriptomic data grouped by cell type. The Human WTA interrogates ~19,000 transcripts encompassing 99.5% of human protein-coding gene expression. Results: These studies support the finding that β cells are dysfunctional in PC tumors compared to healthy controls, which is exacerbated in diabetic patients. Evidence of β cell dysregulation in the PC cohort is provided by the expression in insulin⁺ cells of key transcription factors (TFs) that govern maturation of endocrine α and β cells and their distinct hormonal profile. β cells in both diabetic and nondiabetic patients expressed low levels of the TFs MAFA, RBPJL and PAX6 that regulate β cell maturation; and low insulin (INS) and Islet Amyloid Polypeptide (IAPP) expression. Further, β cells displayed high levels of ARX, that regulates the α-cell phenotype, and the hormones TTR and glucagon (GCG) that are normally expressed in α cells, suggesting that β cells are dedifferentiating in PC. In addition, the Endoplasmic Reticulum (ER) TF XBP1 that maintains the β cell secretory phenotype was also reduced while levels of the ER stress factor DDIT3/CHOP and downstream genes such as the TF ATF3 and the proapoptotic CASP12 were increased. Accordingly, we found that CHOP protein levels were higher in tumor than in nontumor adjacent areas, and that CHOP silencing in β cells halts PC progression in experimental models of PC. Conclusions: The data show that in advancing PC the functional status of β cells is compromised with a trend toward cell loss or dedifferentiation, and hormonal balances shift toward glucagon (in diabetic patients) and pancreatic polypeptide (PPY) production. Diabetic patients have lower expression of lipid metabolism genes including those involved in fatty acid β oxidation than nondiabetics. The data inform further mechanistic studies to investigate the unique mechanisms of PC-related diabetes and therapeutic targets to improve PC patient outcomes.