Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with poor survival rates and frequently carries oncogenic KRAS mutation. However, KRAS has thus far not been a viable therapeutic target. We found that the abundance of YAP mRNA, which encodes Yes-associated protein (YAP), a protein regulated by the Hippo pathway during tissue development and homeostasis, was increased in human PDAC tissue compared with that in normal pancreatic epithelia. In genetically engineered KrasG12D and KrasG12D: Trp53R172H mouse models, pancreas-specific deletion of Yap halted the progression of early neoplastic lesions to PDAC without affecting normal pancreatic development and endocrine function. Although Yap was dispensable for acinar to ductal metaplasia (ADM), an initial step in the progression to PDAC, Yap was critically required for the proliferation of mutant Kras or Kras:Trp53 neoplastic pancreatic ductal cells in culture and for their growth and progression to invasive PDAC in mice. Yap functioned as a critical transcriptional switch downstream of the oncogenic KRAS–mitogen-activated protein kinase (MAPK) pathway, promoting the expression of genes encoding secretory factors that cumulatively sustained neoplastic proliferation, a tumorigenic stromal response in the tumor microenvironment, and PDAC progression in Kras and Kras: Trp53 mutant pancreas tissue. Together, our findings identified Yap as a critical oncogenic KRAS effector and a promising therapeutic target for PDAC and possibly other types of KRAS-mutant cancers.

No abstract

Summary The Merlin/NF2 tumor suppressor restrains cell growth and tumorigenesis by controlling contact-dependent inhibition of proliferation. We have identified a tight-junction-associated protein complex comprising Merlin, Angiomotin, Patj, and Pals1. We demonstrate that Angiomotin functions downstream of Merlin and upstream of Rich1, a small GTPase Activating Protein, as a positive regulator of Rac1. Merlin, through competitive binding to Angiomotin, releases Rich1 from the Angiomotin-inhibitory complex, allowing Rich1 to inactivate Rac1, ultimately leading to attenuation of Rac1 and Ras-MAPK pathways. Patient-derived Merlin mutants show diminished binding capacities to Angiomotin and are unable to dissociate Rich1 from Angiomotin or inhibit MAPK signaling. Depletion of Angiomotin in Nf2−/− Schwann cells attenuates the Ras-MAPK signaling pathway, impedes cellular proliferation in vitro and tumorigenesis in vivo.

Drug resistance is a major challenge in cancer treatment. Emerging evidence indicates that deregulation of YAP/TAZ signaling may be a major mechanism of intrinsic and acquired resistance to various targeted and chemotherapies. Moreover, YAP/TAZ-mediated expression of PD-L1 and multiple cytokines is pivotal for tumor immune evasion. While direct inhibitors of YAP/TAZ are still under development, FDA-approved drugs that indirectly block YAP/TAZ activation or critical downstream targets of YAP/TAZ have shown promise in the clinic in reducing therapy resistance. Finally, BET inhibitors, which reportedly block YAP/TAZ-mediated transcription, present another potential venue to overcome YAP/TAZ-induced drug resistance.

The Hippo-Yap signaling pathway regulates a number of developmental and adult cellular processes, including cell fate determination, tissue growth, and tumorigenesis. Members of the scaffold protein angiomotin (Amot) family interact with several Hippo pathway components, including Yap (Yes-associated protein), and either stimulate or inhibit Yap activity. We used a combination of genetic, biochemical, and transcriptional approaches to assess the functional consequences of the Amot-Yap interaction in mice and in human cells. Mice with a liver-specific Amot knockout exhibited reduced hepatic “oval cell” proliferation and tumorigenesis in response to toxin-induced injury or when crossed with mice lacking the tumor suppressor Nf2. Biochemical examination of the Amot-Yap interaction revealed that the p130 splicing isoform of Amot (Amot-p130) and Yap interacted in both the cytoplasm and nucleus, which involved binding of PPxY and LPxY motifs in Amot-p130 to WW domains of Yap. In the cytoplasm, Amot-p130 prevented the phosphorylation of Yap by blocking access of the WW domains to the kinase Lats1. Within the nucleus, Amot-p130 was associated with the transcriptional complex containing Yap and Teads (TEA domain family members) and contributed to the regulation of a subset of Yap target genes, many of which are associated with tumorigenesis. These findings indicated that Amot acts as a Yap cofactor, preventing Yap phosphorylation and augmenting its activity toward a specific set of genes that facilitate tumorigenesis.

A strategy for targeting protein kinases with large ATP-binding sites by using bulky and rigid octahedral ruthenium complexes as structural scaffolds is presented. A highly potent and selective GSK3 and Pim1 half-sandwich complex NP309 was successfully converted into a PAK1 inhibitor by making use of the large octahedral compounds Λ-FL172 and Λ-FL411 in which the cyclopentadienyl moiety of NP309 is replaced by a chloride and sterically demanding diimine ligands. A 1.65 Å cocrystal structure of PAK1 with Λ-FL172 reveals how the large coordination sphere of the ruthenium complex matches the size of the active site and serves as a yard stick to discriminate between otherwise closely related binding sites.

Pancreatic ductal adenocarcinoma (PDAC) is characterized by a high degree of inflammation and profound immune suppression. Here we identify Yes-associated protein (Yap) as a critical regulator of the immunosuppressive microenvironment in both mouse and human PDAC. Within Kras:p53 mutant pancreatic ductal cells, Yap drives the expression and secretion of multiple cytokines/chemokines, which in turn promote the differentiation and accumulation of Myeloid-derived suppressor cells (MDSCs) both in vitro and in vivo. Pancreas-specific knockout of Yap or antibody-mediated depletion of MDSCs promoted macrophage reprogramming, reactivation of T cells, apoptosis of Kras mutant neoplastic ductal cells, and pancreatic regeneration after acute pancreatitis. In primary human PDAC, YAP expression levels strongly correlate with a MDSC gene signature, and high expression of YAP or MDSC-related genes predicts decreased survival in PDAC patients. These results reveal multifaceted roles YAP in PDAC pathogenesis and underscore its promise as a therapeutic target for this deadly disease.

The development of the embryonic vascular system into a highly ordered network requires precise control over the migration and branching of endothelial cells (ECs). We have previously identified angiomotin (Amot) as a receptor for the angiogenesis inhibitor angiostatin. Furthermore, DNA vaccination targeting Amot inhibits angiogenesis and tumor growth. However, little is known regarding the role of Amot in physiological angiogenesis. We therefore investigated the role of Amot in embryonic neovascularization during zebrafish and mouse embryogenesis. Here we report that knockdown of Amot in zebrafish reduced the number of filopodia of endothelial tip cells and severely impaired the migration of intersegmental vessels. We further show that 75% of Amot knockout mice die between embryonic day 11 (E11) and E11.5 and exhibit severe vascular insufficiency in the intersomitic region as well as dilated vessels in the brain. Furthermore, using ECs differentiated from embryonic stem (ES) cells, we demonstrate that Amot-deficient cells have intact response to vascular endothelial growth factor (VEGF) in regard to differentiation and proliferation. However, the chemotactic response to VEGF was abolished in Amot-deficient cells. We provide evidence that Amot is important for endothelial polarization during migration and that Amot controls Rac1 activity in endothelial and epithelial cells. Our data demonstrate a critical role for Amot during vascular patterning and endothelial polarization.[Keywords: Angiostatin; neovascularization; embryogenesis; GTPase; chemotaxis; transgenic; zebrafish] Supplemental material is available at http://www.genesdev.org.