Tae-Shin Kim

The Hippo–Salvador pathway restrains hepatic oval cell proliferation, liver size, and liver tumorigenesis

Loss of Hippo signaling in Drosophila leads to tissue overgrowth as a result of increased cell proliferation and decreased cell death. YAP (a homolog of Drosophila Yorkie and target of the Hippo pathway) was recently implicated in control of organ size, epithelial tissue development, and tumorigenesis in mammals. However, the role of the mammalian Hippo pathway in such regulation has remained unclear. We now show that mice with liver-specific ablation of WW45 (a homolog of Drosophila Salvador and adaptor for the Hippo kinase) manifest increased liver size and expansion of hepatic progenitor cells (oval cells) and eventually develop hepatomas. Moreover, ablation of WW45 increased the abundance of YAP and induced its localization to the nucleus in oval cells, likely accounting for their increased proliferative capacity, but not in hepatocytes. Liver tumors that developed in mice heterozygous for WW45 deletion or with liver-specific WW45 ablation showed a mixed pathology combining characteristics of hepatocellular carcinoma and cholangiocarcinoma and seemed to originate from oval cells. Together, our results suggest that the mammalian Hippo–Salvador pathway restricts the proliferation of hepatic oval cells and thereby controls liver size and prevents the development of oval cell–derived tumors.

Hippo-Foxa2 signaling pathway plays a role in peripheral lung maturation and surfactant homeostasis

Respiratory distress syndrome (RDS), which is induced by insufficient production of surfactant, is the leading cause of mortality in preterm babies. Although several transcription factors are known to be involved in surfactant protein expression, the molecular mechanisms and signaling pathways upstream of these transcription factors have remained elusive. Here, using mammalian Hippo kinases (Mst1/2, mammalian sterile 20-like kinase 1/2) conditional knockout mice, we demonstrate that Mst1/2 kinases are critical for orchestration of transcription factors involved in surfactant protein homeostasis and prevention of RDS. Mice lacking Mst1/2 in the respiratory epithelium exhibited perinatal mortality with respiratory failure and their lungs contained fewer type I pneumocytes and more immature type II pneumocytes lacking microvilli, lamellar bodies, and surfactant protein expression, pointing to peripheral lung immaturity and RDS. In contrast to previous findings of YAP (Yes-associated protein)-mediated canonical Hippo signaling in the liver and intestine, loss of Mst1/2 kinases induced the defects in pneumocyte differentiation independently of YAP hyperactivity. We instead found that Mst1/2 kinases stabilized and phosphorylated the transcription factor Foxa2 (forkhead box A2), which regulates pneumocyte maturation and surfactant protein expression. Taken together, our results suggest that the mammalian Hippo kinases play crucial roles in surfactant homeostasis and coordination of peripheral lung differentiation through regulation of Foxa2 rather than of YAP.

Ablation of Rassf2 induces bone defects and subsequent haematopoietic anomalies in mice

RASSF2 belongs to the Ras‐association domain family (RASSF) of proteins, which may be involved in the Hippo signalling pathway. However, the role of RASSF2 in vivo is unknown. Here, we show that Rassf2 knockout mice manifest a multisystemic phenotype including haematopoietic anomalies and defects in bone remodelling. Bone marrow (BM) transplantation showed that Rassf2−/− BM cells had a normal haematopoietic reconstitution activity, indicating no intrinsic haematopoietic defects. Notably, in vitro differentiation studies revealed that ablation of Rassf2 suppressed osteoblastogenesis but promoted osteoclastogenesis. Co‐culture experiments showed that an intrinsic defect in osteoblast differentiation from Rassf2−/− osteoblast precursors likely leads to both haematopoiesis and osteoclast defects in Rassf2−/− mice. Moreover, Rassf2 deficiency resulted in hyperactivation of nuclear factor (NF)‐κB during both osteoclast and osteoblast differentiation. RASSF2 associated with IκB kinase (IKK) α and β forms, and suppressed IKK activity. Introduction of either RASSF2 or a dominant‐negative form of IKK into Rassf2−/− osteoclast or osteoblast precursors inhibited NF‐κB hyperactivation and normalized osteoclast and osteoblast differentiation. These observations indicate that RASSF2 regulates osteoblast and osteoclast differentiation by inhibiting NF‐κB signalling.

Reversing the intractable nature of pancreatic cancer by selectively targeting ALDH-high, therapy-resistant cancer cells.

Human pancreatic ductal adenocarcinoma (PDAC) is a cancer with a dismal prognosis. The efficacy of PDAC anticancer therapies is often short-lived; however, there is little information on how this disease entity so frequently gains resistance to treatment. We adopted the concept of cancer stem cells (CSCs) to explain the mechanism of resistance and evaluated the efficacy of a candidate anticancer drug to target these therapy-resistant CSCs. We identified a subpopulation of cells in PDAC with CSC features that were enriched for aldehyde dehydrogenase (ALDH), a marker expressed in certain stem/progenitor cells. These cells were also highly resistant to, and were further enriched by, treatment with gemcitabine. Similarly, surgical specimens from PDAC patients showed that those who had undergone preoperative chemo-radiation therapy more frequently displayed cancers with ALDH strongly positive subpopulations compared with untreated patients. Importantly, these ALDH-high cancer cells were sensitive to disulfiram, an ALDH inhibitor, when tested in vitro. Furthermore, in vivo xenograft studies showed that the effect of disulfiram was additive to that of low-dose gemcitabine when applied in combination. In conclusion, human PDAC-derived cells that express high levels of ALDH show CSC features and have a key role in the development of resistance to anticancer therapies. Disulfiram can be used to suppress this therapy-resistant subpopulation.

Mammalian Sterile 20–like Kinase 1 Suppresses Lymphoma Development by Promoting Faithful Chromosome Segregation

The mammalian Hippo signaling pathway has been implicated in oncogenesis in the context of solid tumors such as hepatocellular carcinoma. Mammalian sterile 20-like kinase 1 (MST1), the core component of the Hippo signaling pathway, is highly expressed in hematopoietic cells. However, its possible impact on tumorigenesis in this setting is unknown. In this study, we provide evidence that Mst1 loss in the mouse enhances chemically and genetically induced lymphoma development by inducing chromosomal instability. Mst1 deficiency increased susceptibility to T-cell acute lymphoblastic leukemia induced by mutagen exposure. Notably, before transformation Mst1(-/-) normal thymocytes showed no changes in proliferation or apoptosis in vitro and in vivo, but they displayed elevated levels of abnormal mitotic chromosomes and aneuploidy, conditions known to promote tumorigenesis. Mst1(-/-) mice also showed accelerated formation of spontaneous lymphomas in a p53-deficient background, accompanied by severe aneuploidy. In clinical specimens of lymphoma and leukemia, we documented frequent downregulation of MST1 expression, consistent with our findings. Taken together, our findings reveal a tumor suppressive function of Mst1 based on its ability to prevent chromosomal instability in lymphocytes.

Hepatocellular Carcinoma with Metastasis to the Spleen in a Holstein Cow

Hepatocellular carcinoma (HCC) with metastasis to the spleen in a Holstein cow was studied by histopathologic and immunohistochemical methods. The tumor was characterized by a pseudoglandular (acinar) pattern with an associated fibrous stroma. Individual cells often had a “hepatoid” appearance but were interspersed with scattered cells exhibiting a clear, periodic acid-Schiff (PAS)-positive cytoplasm and small eccentric nuclei. This pattern was present in nodules found in both liver and spleen. Moreover, hepatoid tumor cells were positive for alpha-fetoprotein. Immunohistochemical studies suggest that myofibroblasts were responsible for the production of fibrous septa surrounding the pseudoglandular structures of bovine HCC. In summary, our histologic and immunohistochemical findings support a diagnosis of primary HCC with splenic metastasis. Furthermore, the associated stromal response appears to be of a myofibroblast origin. The primary etiology of bovine HCC and the significance of the intralesional, PAS-positive clear cells remain undetermined.

Hippo and Mouse Models for Cancer

Among the many signaling pathways related to cancer initiation and progression, the Hippo pathway has emerged recently as a mediator of tumor suppression that is evolutionarily conserved from flies to humans and plays a key role in normal organ development. Genetic engineering of the Hippo pathway in mice has provided important insights into its tumor suppression function. These mouse models have also revealed both canonical and noncanonical modes of action for pathway components in tumor suppression. In this chapter, we first discuss genetic and epigenetic changes identified for Hippo pathway components in human cancers. We then describe established mouse models of cancer related to the Hippo pathway, dividing them into those in which the canonical pathway functions through inhibition of the transcriptional co-activator YAP and those in which noncanonical functions of individual pathway components contribute to tumor suppression.

Mammalian sterile 20 kinase 1 and 2 are important regulators of hematopoietic stem cells in stress condition

The mammalian Hippo signaling pathway has been implicated in the self-renewal and differentiation of stem and progenitor cells. MST1 and MST2 (MST1/2) are core serine-threonine kinases in the Hippo signaling pathway, one of which, MST1, has been extensively investigated for its role in T cell and myeloid cell function. These studies have identified MST1 as a promising therapeutic target in immunological disease. However, the roles of MST1/2 in hematopoietic stem cell (HSC) function in vivo are not fully understood. Here, we report that mice with a conditional deletion of Mst1/2 exhibit impaired hematopoietic stem and progenitor cell (HSPC) function under stress condition. Furthermore, Mst1/2 deletion markedly altered mature cell output. Therefore, MST1/2 are indispensable for maintenance as well as function of stem and progenitor cells under steady state conditions and with transplantation stress.