A Japanese Patient with Gastric Cancer and Dihydropyrimidine Dehydrogenase Deficiency Presenting with DPYD Variants

A 63-year-old Japanese male with stomach adenocarcinoma received oral 5-fluorouracil derivative, cisplatin and trastuzumab chemotherapy. On day 8, severe diarrhea and mucositis developed; chemotherapy was stopped. On day 14, the patient developed renal dysfunction and febrile neutropenia. He also suffered from pneumonia due to Candida albicans. Systemic symptoms improved after intensive conservative treatment. Best supportive care was continued until the patient died from gastric cancer. The dihydropyrimidine dehydroge-nase protein level was low at 3.18 U/mg protein. The result of DPYD genotyping revealed three variants at posi-tions 1615 (G > A), 1627 (A > G), and 1896 (T > C) in exons 13, 13, and 14, respectively

Lansoprazole Upregulates Polyubiquitination of the TNF Receptor-Associated Factor 6 and Facilitates Runx2-mediated Osteoblastogenesis

The transcription factor, runt-related transcription factor 2 (Runx2), plays a pivotal role in the differentiation of the mesenchymal stem cells to the osteochondroblast lineages. We found by the drug repositioning strategy that a proton pump inhibitor, lansoprazole, enhances nuclear accumulation of Runx2 and induces osteoblastogenesis of human mesenchymal stromal cells. Systemic administration of lansoprazole to a rat femoral fracture model increased osteoblastogenesis. Dissection of signaling pathways revealed that lansoprazole activates a noncanonical bone morphogenic protein (BMP)-transforming growth factor-beta (TGF-β) activated kinase-1 (TAK1)–p38 mitogen-activated protein kinase (MAPK) pathway. We found by in cellulo ubiquitination studies that lansoprazole enhances polyubiquitination of the TNF receptor-associated factor 6 (TRAF6) and by in vitro ubiquitination studies that the enhanced polyubiquitination of TRAF6 is attributed to the blocking of a deubiquitination enzyme, cylindromatosis (CYLD). Structural modeling and site-directed mutagenesis of CYLD demonstrated that lansoprazole tightly fits in a pocket of CYLD where the C-terminal tail of ubiquitin lies. Lansoprazole is a potential therapeutic agent for enhancing osteoblastic differentiation

Wnt/β-catenin signaling suppresses expressions of Scx, Mkx, and Tnmd in tendon-derived cells

After tendon injuries, biomechanical properties of the injured tendon are not fully recovered in most cases. Modulation of signaling pathways, which are involved in tendon development and tendon repair, is one of attractive modalities to facilitate proper regeneration of the injured tendon. The roles of TGF-beta signaling in tendon homeostasis and tendon development have been elucidated. In contrast, the roles of Wnt/beta-catenin signaling in tendon remain mostly elusive. We found that the number of beta-catenin-positive cells was increased at the injured site, suggesting involvement of Wnt/beta-catenin signaling in tendon healing. Activation of Wnt/beta-catenin signaling suppressed expressions of tenogenic genes of Scx, Mkx, and Tnmd in rat tendon-derived cells (TDCs) isolated from the Achilles tendons of 6-week old rats. Additionally, activation of Wnt/beta-catenin reduced the amounts of Smad2 and Smad3, which are intracellular mediators for TGF-beta signaling, and antagonized upregulation of Scx induced by TGF-beta signaling in TDCs. We found that Wnt/beta-catenin decreased Mkx and Tnmd expressions without suppressing Scx expression in Scx-programmed tendon progenitors. Our studies suggest that Wnt/beta-catenin signaling is a repressor for tenogenic gene expressions

Verapamil Protects against Cartilage Degradation in Osteoarthritis by Inhibiting Wnt/β-Catenin Signaling

<div><p>In past years, the canonical Wnt/β-catenin signaling pathway has emerged as a critical regulator of cartilage development and homeostasis. FRZB, a soluble antagonist of Wnt signaling, has been studied in osteoarthritis (OA) animal models and OA patients as a modulator of Wnt signaling. We screened for FDA-approved drugs that induce <i>FRZB</i> expression and suppress Wnt/β-catenin signaling. We found that verapamil, a widely prescribed L-type calcium channel blocker, elevated FRZB expression and suppressed Wnt/β-catenin signaling in human OA chondrocytes. Expression and nuclear translocation of β-catenin was attenuated by verapamil in OA chondrocytes. Lack of the verapamil effects in LiCl-treated and FRZB-downregulated OA chondrocytes also suggested that verpamil suppressed Wnt signaling by inducing FRZB. Verapamil enhanced gene expressions of chondrogenic markers of <i>ACAN</i> encoding aggrecan, <i>COL2A1</i> encoding collagen type II α1, and <i>SOX9</i>, and suppressed Wnt-responsive <i>AXIN2</i> and <i>MMP3</i> in human OA chondrocytes. Verapamil ameliorated Wnt3A-induced proteoglycan loss in chondrogenically differentiated ATDC5 cells. Verapamil inhibited hypertrophic differentiation of chondrocytes in the explant culture of mouse tibiae. Intraarticular injection of verapamil inhibited OA progression as well as nuclear localizations of β-catenin in a rat OA model. We propose that verapamil holds promise as a potent therapeutic agent for OA by upregulating FRZB and subsequently downregulating Wnt/β-catenin signaling.</p></div

CTGF/CCN2 facilitates LRP4-mediated formation of the embryonic neuromuscular junction.

At the neuromuscular junction (NMJ), lipoprotein-related receptor 4 (LRP4) mediates agrin-induced MuSK phosphorylation that leads to clustering of acetylcholine receptors (AChRs) in the postsynaptic region of the skeletal muscle. Additionally, the ectodomain of LRP4 is necessary for differentiation of the presynaptic nerve terminal. However, the molecules regulating LRP4 have not been fully elucidated yet. Here, we show that the CT domain of connective tissue growth factor (CTGF/CCN2) directly binds to the third beta-propeller domain of LRP4. CTGF/CCN2 enhances the binding of LRP4 to MuSK and facilitates the localization of LRP4 on the plasma membrane. CTGF/CCN2 enhances agrin-induced MuSK phosphorylation and AChR clustering in cultured myotubes. Ctgf-deficient mouse embryos (Ctgf-/- ) have small AChR clusters and abnormal dispersion of synaptic vesicles along the motor axon. Ultrastructurally, the presynaptic nerve terminals have reduced numbers of active zones and mitochondria. Functionally, Ctgf-/- embryos exhibit impaired NMJ signal transmission. These results indicate that CTGF/CCN2 interacts with LRP4 to facilitate clustering of AChRs at the motor endplate and the maturation of the nerve terminal

CTGF/CCN2 facilitates LRP4-mediated formation of the embryonic neuromuscular junction

At the neuromuscular junction (NMJ), lipoprotein-related receptor 4 (LRP4) mediates agrin-induced MuSK phosphorylation that leads to clustering of acetylcholine receptors (AChRs) in the postsynaptic region of the skeletal muscle. Additionally, the ectodomain of LRP4 is necessary for differentiation of the presynaptic nerve terminal. However, the molecules regulating LRP4 have not been fully elucidated yet. Here, we show that the CT domain of connective tissue growth factor (CTGF/CCN2) directly binds to the third beta-propeller domain of LRP4. CTGF/CCN2 enhances the binding of LRP4 to MuSK and facilitates the localization of LRP4 on the plasma membrane. CTGF/CCN2 enhances agrin-induced MuSK phosphorylation and AChR clustering in cultured myotubes. Ctgf-deficient mouse embryos (Ctgf-/- ) have small AChR clusters and abnormal dispersion of synaptic vesicles along the motor axon. Ultrastructurally, the presynaptic nerve terminals have reduced numbers of active zones and mitochondria. Functionally, Ctgf-/- embryos exhibit impaired NMJ signal transmission. These results indicate that CTGF/CCN2 interacts with LRP4 to facilitate clustering of AChRs at the motor endplate and the maturation of the nerve terminal

Verapamil suppresses hypertrophic differentiation of chondrocytes and β-catenin staining in growth plates in explanted mouse fetal tibiae on embryonic day 17.5.

<p>(A) Tibiae are cultured with LiCl or verapamil for 10 days, and coronal slices of paraffin sections are stained with Alcian blue combined with hematoxylin and eosin staining. Three layers of proliferative (green), prehypertrophic (yellow), and hypertrophic (red) zones are indicated by bars. Scale bar = 200 μm. (B) Verapamil suppresses the number of chondrocytes in the hypertrophic zone in (A). (C) Immunofluorescence with antibody against β-catenin in proximal tibiae of mouse embryo (E17.5). Color bars indicate layers as indicated in (A). Scale bar = 200 μm. (D) Verapamil suppresses the number of β-catenin-positive cells in the proliferative zone. The mean and SEM (<i>n</i> = 3) are indicated. *<i>p</i><0.05 versus control by one-way ANOVA with Tukey's test.</p