Gnas^R201C Induces Murine Pancreatic Cystic Neoplasms through Suppression of YAP1 Signaling and Transcriptional Reprogramming

Abstract

Background & Aims: Somatic "hotspot" mutations of GNAS, which encodes for the alpha subunit of stimulatory G-protein, are present in ~60% of intraductal papillary mucinous neoplasms (IPMNs) of the pancreas. There are currently no cognate animal models that recapitulate the biology of mutant Gnas-induced IPMNs, and the underlying mechanisms that lead to the cystic pathway of neoplasia in the pancreas remain unknown. Methods:We generated p48-Cre; LSL-Kras G12D ; Rosa26R-LSL-rtTA-TetO-Gnas R201C mice (Kras; Gnas mice) where pancreas-specific Gnas R201C expression was induced by doxycycline administration. In this model, mutant Kras is constitutively expressed, and control mice were produced through absence of doxycycline. Separate cohorts of mice were utilized for timed necropsies and for Kaplan-Meier survival analysis. Isogenic cell lines (with doxycycline inducible mutant Gnas expression) were propagated from the resulting pancreatic ductal adenocarcinoma (PDAC). Results: Co-expression of KrasG12D and Gnas R201C resulted in the development of pancreatic cystic lesions resembling human IPMNs in 100% of mice, with higher grades of epithelial dysplasia observed over time. Approximately one-third of Kras; Gnas mice developed PDAC at a median of 38 weeks post doxycycline induction. Gnas R201C did not accelerate oncogenic transformation with Kras G12D , but rather, reprogrammed Ras-induced neoplasms towards a welldifferentiated phenotype. Gnas R201C induction led to activation of the inhibitory Hippo kinase cascade and cytoplasmic sequestration of phosphorylated YAP1 protein, a phenomenon that was also observed in human IPMN with GNAS mutations.Conclusions: GNAS R201C functions not as a traditional oncogene, but rather as an "oncomodulator" of KRAS-mediated pancreatic neoplasia, through suppression of YAP1 and transcriptional reprogramming towards a differentiated (large ductal) phenotype.

Gαs–Protein Kinase A (PKA) Pathway Signalopathies: The Emerging Genetic Landscape and Therapeutic Potential of Human Diseases Driven by Aberrant Gαs-PKA Signaling

“…Instead, inactivation of tumor suppressors, like p53 (TP53), cyclin dependent kinase kinhibitor 2a (CDKN2A), or SMAD family member 4 (SMAD4), are needed to facilitate efficient progression to PDAC (Ideno et al, 2018;Patra et al, 2018). Interestingly, in the context of PDAC, GNAS R201C expression through activation of PKA actually attenuates aggressiveness and invasiveness due to epithelial differentiation (Pattabiraman et al, 2016;Ideno et al, 2018). This is supported by clinical evidence that patients with GNAS mutant have a better overall survival in appendix cancer (Ang et al, 2018).…”

“…Somewhat counterintuitively, GNAS R201C expression does not accelerate KRAS-driven progression to pancreatic adenocarcinoma (PDAC). Instead, inactivation of tumor suppressors, like p53 (TP53), cyclin dependent kinase kinhibitor 2a (CDKN2A), or SMAD family member 4 (SMAD4), are needed to facilitate efficient progression to PDAC (Ideno et al, 2018;Patra et al, 2018). Interestingly, in the context of PDAC, GNAS R201C expression through activation of PKA actually attenuates aggressiveness and invasiveness due to epithelial differentiation (Pattabiraman et al, 2016;Ideno et al, 2018).…”

“…This suggests that in epithelial tissues, GNAS is most important in early initiation events. Indeed, several studies have highlighted that GNAS mutation can accelerate tumorigenesis (Wilson et al, 2010;Ideno et al, 2018;Patra et al, 2018;Coles et al, 2020). Given that tumors are heterogeneous, GNAS may confer a selective advantage initially, in which context additional mutational insults, like KRAS and subsequently TP53, can drive malignant growth ultimately independent of GNAS mutation.…”

“…Mechanistically, stem cell expansion in the hair follicle is controlled by PKA-mediated repression of yesassociated protein (YAP) and glioma-associated oncogene (GLI) transcriptional activity, with no effect on other stem cell programs like Wnt (Iglesias- Bartolome et al, 2015). Of note, PKA has been shown to repress YAP activity in pancreatic cancer (in which PKA functions as an oncogene) but still induce a differentiation phenotype (Ideno et al, 2018). Much like GNAS and PKA, YAP has also been shown to behave as either an oncogene or a tumor suppressor depending on the cellular context.…”

Metabolic Reprogramming by Mutant GNAS Creates an Actionable Dependency in Intraductal Papillary Mucinous Neoplasms of the Pancreas

“…The autochthonous model of IPMN, which results from pancreas-specific co-expression of mutant Kras G12D and Gnas R201C (Kras;Gnas mice) has been described [8]. Briefly, we generated p48 Cre ; Kras LSL-G12D ; Rosa26 LSL-rtTA , Tet operon (TetO)-Gnas R201C mice.…”