Molecular triggers for neuropathy and neural invasion in genetically engineered mouse models of pancreatic ductal adenocarcinoma

• Applicant: Professor Dr. Ihsan Ekin Demir
• Subject Area: General and Visceral Surgery
• Term: since 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 522706533

Neural invasion (NI) is an independent prognostic factor and one of the main drivers of local recurrence and intractable cancer pain in PDAC, but its pathomechanism is barely understood. In the first funding period, we aimed modelling NI in innovative models of PDAC for understanding its underlying mechanisms. We screened phenotypes of over 250 allele combinations and discovered novel human-like, truly neuro-invasive GEMM of PDAC. In bulk and spatial transcriptome analyses, we identified overexpression of neuropeptide Y (Npy) in cancer cells, and the "inflammatory" subtype of pancreatic stellate cells (that upregulate Il6) to be associated with NI. In genetic screens, we found a novel link between NI and inactivating mutations in O-gylcosylation-associated genes Galnt2 and St3Gal6. Finally, we generated a novel "innervated organoid" setup, which combines cancer organoids with neural crest cells for inducing cancer innervation. In this project, we will build upon these findings to delve deeper into the underpinnings of NI by pursuing the following aims: 1) Understand the mechanism behind NPY-mediated neural invasion in PDAC at the cellular level; 2) Delve into the impact of IL6 signaling during NI in PDAC; 3) Understand how O-glycosylation is linked to neural invasion in PDAC; and 4) Generate and screen for novel models of neural invasion and neuroplasticity in PDAC. In WP1, we will study the mechanisms of Npy-mediated NI in PDAC. We will isolate the cancer cells from our Npy-overexpressing mouse mutants and will test their biological traits, their phosphoproteome and interactions with neurons and Schwann cells in our 3D culture models. In WP2, we will investigate how neuron-derived IL6, and stellate cells of neuro-invasive PDAC models augment NI. We will analyze NI in mice with constitutive IL6 signalling (KPC;L-gp130) and in co-cultures of DRG neurons and cancer cells from this model. We will co-implant murine cancer cells with stellate cells of mice with/without NI and subsequently analyze NI. In WP3, we will analyze how O-glycosylation is linked to NI. We will compare the "glycome" of Galnt2- and St3Gal6-silenced PDAC cells for identifying the differentially glycosylated proteins, and analyze the in vitro and in vivo neuro-invasiveness of PDAC cells that have been edited for these proteins. In the final WP4, we will explore novel models of NI in PDAC. Here, we will orthotopically implant murine PDAC organoids in mixture with murine neural crest cells and analyze innervation and NI. Collectively, these investigations will provide a novel insight into the pathomechanisms of NI and point out novel, potentially actionable targets.

• DFG Programme: Research Grants