S1PR3-G12-biased agonist ALESIA targets cancer metabolism and promotes glucose starvation.

Metabolic activities are altered in cancer cells compared with those in normal cells, and the cancer-specific pathway becomes a potential therapeutic target. Higher cellular glucose consumption, which leads to lower glucose levels, is a hallmark of cancer cells. In an objective screening for chemicals that induce cell death under low-glucose conditions, we discovered a compound, denoted as ALESIA (Anticancer Ligand Enhancing Starvation-induced Apoptosis). By our shedding assay of transforming growth factor α in HEK293A cells, ALESIA was determined to act as a sphingosine-1-phosphate receptor 3-G12-biased agonist that promotes nitric oxide production and oxidative stress. The oxidative stress triggered by ALESIA resulted in the exhaustion of glucose, cellular NADPH deficiency, and then cancer cell death. Intraperitoneal administration of ALESIA improved the survival of mice with peritoneally disseminated rhabdomyosarcoma, indicating its potential as a new type of anticancer drug for glucose starvation therapy.

Hydroxypropyl Cyclodextrin Improves Amiodarone-induced Aberrant Lipid Homeostasis of Alveolar Cells.

Alveolar epithelial type II (AT2) cells secrete pulmonary surfactant via lamellar bodies (LBs). Abnormalities in LBs are associated with pulmonary disorders, including fibrosis. However, high-content screening (HCS) for LB abnormalities is limited by the lack of understanding of AT2 cell functions. In the present study, we have developed LB cells harboring LB-like organelles that secrete surfactant proteins. These cells were more similar to AT2 cells than to parental A549 cells. LB cells recapitulated amiodarone (AMD)-induced LB enlargement, similar to AT2 cells of patients exposed to AMD. To reverse AMD-induced LB abnormalities, we performed HCS of approved drugs and identified 2-hydroxypropyl-ß-cyclodextrin (HPßCD), a cyclic oligosaccharide, as a potential therapeutic agent. A transcriptome analysis revealed that HPßCD modulates lipid homeostasis. In addition, HPßCD inhibited AMD-induced LB abnormalities in human induced pluripotent stem cell-derived AT2 cells. Our results demonstrate that LB cells are useful for HCS and suggest that HPßCD is a candidate therapeutic agent for AMD-induced interstitial pneumonia.

Necrostatin-7 suppresses RANK-NFATc1 signaling and attenuates macrophage to osteoclast differentiation.

Osteoclasts play a crucial role in osteolytic bone diseases, such as osteoporosis, rheumatoid arthritis, periodontitis, Paget's disease of bone and bone metastatic tumors. Therefore, controlling osteoclast differentiation and function has been considered a promising therapeutic strategy. Here, we show that necrostatin (Nec)-7, an inhibitor of programmed necrosis, strongly suppressed receptor activator of nuclear factor (NF)-κB ligand (RANKL)-induced osteoclastogenesis and bone resorption, without compromising macrophage colony-stimulating factor (M-CSF)-supported survival and growth of osteoclast precursor cells. Accordingly, Nec-7 significantly decreased the levels of RANKL-induced osteoclastogenic marker genes, such as cathepsin K. Mechanistically, Nec-7 neither affected MAPK nor NF-κB activation; however, it strongly inhibited the RANKL receptor (RANK) to nuclear factor of activated T cells c1 (NFATc1) signaling. Lentiviral expression of RANK in bone marrow-derived macrophages significantly restored osteoclastogenesis and NFATc1 amplification in Nec-7-treated cells. In this study, we revealed that Nec-7-sensitive pathways are crucially involved in osteoclast formation and function. Investigation of the molecular mechanism(s) through which Nec-7 inhibits RANK-NFATc1 signaling axis may lead to the development of new therapeutic strategies for bone disease.

Actin-binding protein coronin 1A controls osteoclastic bone resorption by regulating lysosomal secretion of cathepsin K.

Osteoclasts degrade bone matrix proteins via the secretion of lysosomal enzymes. However, the precise mechanisms by which lysosomal components are transported and fused to the bone-apposed plasma membrane, termed ruffled border membrane, remain elusive. Here, we identified coronin 1A as a negative regulator of exocytotic release of cathepsin K, one of the most important bone-degrading enzymes in osteoclasts. The modulation of coronin 1A expression did not alter osteoclast differentiation and extracellular acidification, but strongly affected the secretion of cathepsin K and osteoclast bone-resorption activity, suggesting the coronin 1A-mediated regulation of lysosomal trafficking and protease exocytosis. Further analyses suggested that coronin 1A prevented the lipidation-mediated sorting of the autophagy-related protein LC3 to the ruffled border and attenuated lysosome-plasma membrane fusion. In this process, the interactions between coronin 1A and actin were crucial. Collectively, our findings indicate that coronin 1A is a pivotal component that regulates lysosomal fusion and the secretion pathway in osteoclast-lineage cells and may provide a novel therapeutic target for bone diseases.

Anti-arthritic effect of E3 ubiquitin ligase, c-MIR, expression in the joints.

Cellular modulator of immune recognition (c-MIR) is an E3 ubiquitin ligase that ubiquitinates MHC class II and CD86 for their endocytosis and subsequent lysosomal degradation. In accordance with their importance in antigen presentation, systemic c-MIR over-expression downmodulates adaptive immune responses. Rheumatoid arthritis (RA) is a chronic synovitis driven by autoimmunity in the joints. Since antigen-presenting cells, such as macrophages, dendritic cells (DCs) and rheumatoid factor-positive B cells are abundant in the rheumatoid synovial tissues, autoantigens released by tissue damage should be presented locally, leading to amplification of systemic arthritogenic immune responses. Assuming that inhibition of the antigen presentation in the synovial tissues should suppress systemic arthritis, we transferred the c-MIR gene to the hind leg synovial tissues from mice with type II collagen (CII)-induced arthritis, an animal model of RA. The gene was transferred adenovirally because adenoviruses can infect DC and macrophages in vivo. Unexpectedly, therapeutic effect was observed only in the treated joints. Splenocyte responses and serum antibodies against CII were not suppressed. Moreover, in vitro studies disclosed that c-MIR gene transfer suppressed IL-6 production from synovial fibroblasts stimulated with tumor necrosis factor (TNF)-α or IL-1ß. Bone marrow-derived macrophages and DC from c-MIR transgenic mice were impaired in IL-6 and TNF-α production when stimulated with LPS. This suppression was controlled at the post-transcriptional level since their mRNA was not affected. These results have disclosed a new function of c-MIR, inhibition of inflammatory cytokine production. Induction of c-MIR in the joints could be a new therapeutic approach to the treatment of RA.

Intervention of an inflammation amplifier, triggering receptor expressed on myeloid cells 1, for treatment of autoimmune arthritis.

OBJECTIVE: Triggering receptor expressed on myeloid cells 1 (TREM-1) is inducible on monocyte/macrophages and neutrophils and accelerates tissue destruction by propagating inflammatory responses in disease related to bacterial infections. Its blockade rescues the hosts in murine models of sepsis, to clear the bacteria without impairing the host defense. The aim of this study was to investigate the involvement of TREM-1 in an autoimmune, noninfectious disease. METHODS: Synovial tissue specimens from the joints of patients with rheumatoid arthritis (RA) and the joints of mice with collagen-induced arthritis (CIA) were examined for TREM-1 expression, using flow cytometric analysis. Expression of TREM-1 on macrophages was induced by lipopolysaccharide, with or without a cyclooxygenase inhibitor. Rheumatoid synovial cells were stimulated with agonistic anti-TREM-1 antibodies. Recombinant adenovirus encoding the extracellular domain of TREM-1 fused with IgG-Fc (AxCATREM-1 Ig) or synthetic TREM-1 antagonistic peptides were injected to treat CIA, and the clinical manifestations of the antigen-specific T cell and B cell responses were evaluated. RESULTS: TREM-1 was expressed on CD14+ cells in rheumatoid synovial tissue and synovial macrophages from mice with CIA. Unlike murine macrophages, human monocyte/macrophages did not depend on prostaglandin E2 for up-regulation of TREM-1. Agonistic anti-TREM-1 antibodies promoted tumor necrosis factor alpha production from rheumatoid synovial cells. Blockade of TREM-1 using AxCATREM-1 Ig and antagonistic peptides ameliorated CIA without affecting the serum levels of anti-type II collagen antibodies or the proliferative responses of splenocytes to type II collagen. CONCLUSION: TREM-1 ligation contributes to the pathology of autoimmune arthritis. The results of this study implied that blockade of TREM-1 could be a new approach to rheumatic diseases that is safer than the presently available immunosuppressive treatments.