Design, synthesis, and optimization of a series of 2-azaspiro[3.3]heptane derivatives as orally bioavailable fetal hemoglobin inducers.

Pharmacological reactivation of the γ-globin gene for the production of fetal hemoglobin (HbF) is a promising approach for the management of β-thalassemia and sickle cell disease (SCD). We conducted a phenotypic screen in human erythroid progenitor cells to identify molecules that could induce HbF, which resulted in identification of the hit compound 1. Exploration of structure-activity relationships and optimization of ADME properties led to 2-azaspiro[3.

Development of an exon skipping therapy for X-linked Alport syndrome with truncating variants in COL4A5.

Currently, there are no treatments for Alport syndrome, which is the second most commonly inherited kidney disease. Here we report the development of an exon-skipping therapy using an antisense-oligonucleotide (ASO) for severe male X-linked Alport syndrome (XLAS). We targeted truncating variants in exon 21 of the COL4A5 gene and conducted a type IV collagen α3/α4/α5 chain triple helix formation assay, and in vitro and in vivo treatment efficacy evaluation.

Phenotypic-screening generates active novel fetal globin-inducers that downregulate Bcl11a in a monkey model.

Heritable disorders associated with hemoglobin production are the most common monogenic disorders. These are mainly represented by disorders such as β-thalassemia and sickle cell disease. Induction of fetal hemoglobin (HbF) has been known to ameliorate the clinical severity of these β hemoglobinopathies.

G protein-coupled receptor 39 plays an anti-inflammatory role by enhancing IL-10 production from macrophages under inflammatory conditions.

The possible role of G protein-coupled receptor 39 (GPR39) in inflammation was examined in macrophages. Gpr39 expression increased in thioglycollate-induced peritoneal macrophages. TC-G 1008, a G protein-coupled receptor 39 agonist, enhanced interleukin (IL)-10 production from thioglycollate-induced peritoneal macrophages stimulated with lipopolysaccharide (LPS) in vitro.

Heterogeneity of auto-antibodies against nAChR in myasthenic serum and their pathogenic roles in experimental autoimmune myasthenia gravis.

Many myasthenia gravis (MG) patients have auto-antibodies against the nicotinic acetylcholine receptor (nAChR), and monoclonal antibodies against the main immunogenic region (MIR) of nAChR can induce experimental autoimmune MG (EAMG). We investigated whether Fab fragment of MIR antibody (Fab35) could block the pathogenicity of polyclonal antibodies. Fab35 partially inhibited nAChR downmodulation, blocked EAMG serum-induced binding of polyclonal antibodies and complement deposition in vitro.

Analysis of peripheral B cells and autoantibodies against the anti-nicotinic acetylcholine receptor derived from patients with myasthenia gravis using single-cell manipulation tools.

The majority of patients with myasthenia gravis (MG), an organ-specific autoimmune disease, harbor autoantibodies that attack the nicotinic acetylcholine receptor (nAChR-Abs) at the neuromuscular junction of skeletal muscles, resulting in muscle weakness. Single cell manipulation technologies coupled with genetic engineering are very powerful tools to examine T cell and B cell repertoires and the dynamics of adaptive immunity. These tools have been utilized to develop mAbs in parallel with hybridomas, phage display technologies and B-cell immortalization.

Design, synthesis, and biological evaluation of 3-(1-Aryl-1H-indol-5-yl)propanoic acids as new indole-based cytosolic phospholipase A2α inhibitors.

This article describes the design, synthesis, and biological evaluation of new indole-based cytosolic phospholipase A2α (cPLA2α, a group IVA phospholipase A2) inhibitors. A screening-hit compound from our library, (E)-3-{4-[(4-chlorophenyl)thio]-3-nitrophenyl}acrylic acid (5), was used to design a class of 3-(1-aryl-1H-indol-5-yl)propanoic acids as new small molecule inhibitors. The resultant structure-activity relationships studied using the isolated enzyme and by cell-based assays revealed that the 1-(p-O-substituted)phenyl, 3-phenylethyl, and 5-propanoic acid groups on the indole core are essential for good inhibitory activity against cPLA2α.

Pharmacokinetic/pharmacodynamic analyses of chymase inhibitor SUN13834 in NC/Nga mice and prediction of effective dosage for atopic dermatitis patients.

A chymase inhibitor SUN13834 has been shown to improve skin condition in animal models for atopic dermatitis. In the present study, effective dosages of SUN13834 for atopic dermatitis patients were predicted by pharmacokinetic/pharmacodynamic (PK/PD) analyses of SUN13834 in NC/Nga mice, which spontaneously develop atopic dermatitis-like skin lesions. For the PK/PD analyses, we utilized the minimum effective plasma concentration of unbound SUN13834 in late-phase reaction of trinitrochlorobenzene (TNCB)-induced biphasic dermatitis in mice, based on the assumption that the minimum effective plasma concentrations are the same among the two animal models.

Analysis of signal transduction pathways involved in anti-CD30 mAb-induced human eosinophil apoptosis.

Background: Activation of cell surface CD30 by immobilized anti-CD30 monoclonal antibodies (mAbs) induces extremely rapid and intense apoptosis in human eosinophils in vitro. This anti-CD30 mAb-induced eosinophil apoptosis was inhibited by the addition of inhibitors of p38 and ERK1/2 mitogen-activated protein kinases (MAPKs). However, the signal transduction pathways other than for MAPKs remain unclear.

Oral chymase inhibitor SUN13834 ameliorates skin inflammation as well as pruritus in mouse model for atopic dermatitis.

Chymase is a chymotrypsin-like serine protease exclusively stored in secretory granules of mast cells and has been thought to participate in allergic diseases. It has already been shown that chymase inhibitor SUN13834 improves dermatitis in NC/Nga mice that spontaneously develop dermatitis resembling atopic dermatitis. In the present study, effect of chymase inhibitor SUN13834 on itch, the major feature of atopic dermatitis, was examined using a mouse dermatitis model induced by repeated topical application of 2,4-dinitrofluorobenzene (DNFB).

Rapid and strong induction of apoptosis in human eosinophils by anti-CD30 mAb-coated microspheres and phagocytosis by macrophages.

Background: Eosinophils represent a potential therapeutic target in allergic diseases. We previously reported that two clones of anti-CD30 mAbs (HRS-4 and Ber-H8) induced extremely rapid and intense apoptosis in human eosinophils in vitro, but only when the mAbs were immobilized on plates [Matsumoto K, J Immunol 2004;172:2186]. As the initial step towards clinical application of these anti-CD30 mAbs in the treatment of allergic diseases, we made an attempt to clarify two issues; first, whether or not anti-CD30 mAb-coated microspheres can efficiently induce apoptosis in human eosinophils, and second, whether or not these apoptotic eosinophils can be phagocytosed by macrophages without the release of granular proteins.

CpG oligodeoxynucleotide prolongs eosinophil survival through activation of contaminating B cells and plasmacytoid dendritic cells in vitro.

Background: In our previous study, oligodeoxynucleotides containing unmethylated CpG motifs (CpG ODNs) significantly prolonged eosinophil survival without inducing active release of eosinophil-derived neurotoxin or interleukin 8. In addition, this survival-promoting activity was nuclear factor-kappaB dependent. However, some eosinophil preparations from different donors hardly responded to CpG ODNs at all.

Mast cell chymase induces expression of chemokines for neutrophils in eosinophilic EoL-1 cells and mouse peritonitis eosinophils.

Human chymase induced release of interleukin-8 (IL-8) in human EoL-1 cells that had been differentiated into eosinophil-like cells with butyric acid. The chymase-induced IL-8 production was specific in that other cytokines/chemokines examined were not induced. Human chymase also increased mRNA for IL-8 in the differentiated EoL-1 cells, showing involvement of mRNA synthesis.

NR4A orphan nuclear receptor family in peripheral blood eosinophils from patients with atopic dermatitis and apoptotic eosinophils in vitro.

To identify novel genes related to the clinical signs of atopic dermatitis (AD), differentially expressed genes were sought in peripheral blood eosinophils from both AD patients and healthy volunteers. RNA was prepared from eosinophils, expression of various genes was monitored using the Affymetrix GeneChip, and expression was quantified by real-time RT-PCR. Two genes, Nur77 and NOR1, members of NR4A orphan nuclear receptor family, were expressed at a significantly higher level in AD patients than in healthy volunteers.

Eosinophil migration induced by mast cell chymase is mediated by extracellular signal-regulated kinase pathway.

Mast cell chymase is known to induce eosinophil migration in vivo and in vitro. In the present study, we investigated possible involvement of mitogen-activated protein (MAP) kinases; extracellular signal-regulated kinase (ERK), c-Jun amino-terminal kinase (JNK), and p38, in the chymase-induced eosinophil migration. Human chymase induced a rapid phosphorylation of ERK1/2 and p38 in human eosinophilic leukemia EoL-1 cells, while no phosphorylation was detected in JNK.

Extremely rapid and intense induction of apoptosis in human eosinophils by anti-CD30 antibody treatment in vitro.

Apoptosis is an important cellular mechanism for controlling cell viability and proliferation. With respect to eosinophils, cytokines prolong their survival, whereas corticosteroids reduce their survival in vitro. CD30, a member of the TNFR family, is expressed on the surface of many cell types, including Hodgkin's lymphoma cells.