Inhibitors of the Sialidase NEU3 as Potential Therapeutics for Fibrosis.

Fibrosing diseases are a major medical problem, and are associated with more deaths per year than cancer in the US. Sialidases are enzymes that remove the sugar sialic acid from glycoconjugates. In this review, we describe efforts to inhibit fibrosis by inhibiting sialidases, and describe the following rationale for considering sialidases to be a potential target to inhibit fibrosis. First, sialidases are upregulated in fibrotic lesions in humans and in a mouse model of pulmonary fibrosis. Second, the extracellular sialidase NEU3 appears to be both necessary and sufficient for pulmonary fibrosis in mice. Third, there exist at least three mechanistic ways in which NEU3 potentiates fibrosis, with two of them being positive feedback loops where a profibrotic cytokine upregulates NEU3, and the upregulated NEU3 then upregulates the profibrotic cytokine. Fourth, a variety of NEU3 inhibitors block pulmonary fibrosis in a mouse model. Finally, the high sialidase levels in a fibrotic lesion cause an easily observed desialylation of serum proteins, and in a mouse model, sialidase inhibitors that stop fibrosis reverse the serum protein desialylation. This then indicates that serum protein sialylation is a potential surrogate biomarker for the effect of sialidase inhibitors, which would facilitate clinical trials to test the exciting possibility that sialidase inhibitors could be used as therapeutics for fibrosis.

A CD209 ligand and a sialidase inhibitor differentially modulate adipose tissue and liver macrophage populations and steatosis in mice on the Methionine and Choline-Deficient (MCD) diet.

Non-alcoholic fatty liver disease (NAFLD) is associated with obesity and type 2 diabetes and is characterized by the accumulation of fat in the liver (steatosis). NAFLD can transition into non-alcoholic steatohepatitis (NASH), with liver cell injury, inflammation, and an increased risk of fibrosis. We previously found that injections of either 1866, a synthetic ligand for the lectin receptor CD209, or DANA, a sialidase inhibitor, can inhibit inflammation and fibrosis in multiple animal models. The methionine and choline-deficient (MCD) diet is a model of NASH which results in the rapid induction of liver steatosis and inflammation. In this report, we show that for C57BL/6 mice on a MCD diet, injections of both 1866 and DANA reversed MCD diet-induced decreases in white fat, decreases in adipocyte size, and white fat inflammation. However, these effects were not observed in type 2 diabetic db/db mice on a MCD diet. In db/db mice on a MCD diet, 1866 decreased liver steatosis, but these effects were not observed in C57BL/6 mice. There was no correlation between the ability of 1866 or DANA to affect steatosis and the effects of these compounds on the density of liver macrophage cells expressing CLEC4F, CD64, F4/80, or Mac2. Together these results indicate that 1866 and DANA modulate adipocyte size and adipose tissue macrophage populations, that 1866 could be useful for modulating steatosis, and that changes in the local density of 4 different liver macrophages cell types do not correlate with effects on liver steatosis.

Reduced Sialylation and Bioactivity of the Antifibrotic Protein Serum Amyloid P in the Sera of Patients with Idiopathic Pulmonary Fibrosis.

Pulmonary fibrosis is a chronic and generally fatal disorder characterized by progressive formation of scar-like tissue in the lungs. Sialic acids are often found as the terminal sugar on extracellular glycoconjugates such as protein glycosylations. Sialidases, also known as neuraminidases, desialylate glycoconjugates. Serum amyloid P (SAP), a pentameric serum glycoprotein that has two sialic acids on each polypeptide, inhibits the differentiation of monocytes into fibrocytes and promotes human PBMCs to accumulate high extracellular levels of IL-10. When SAP is desialylated with sialidase, the effects of SAP on fibrocyte differentiation and IL-10 accumulation are strongly inhibited. Intriguingly, in patients with pulmonary fibrosis, there are increased levels of sialidase activity in the bronchoalveolar lavage fluid, increased levels of sialidases in the lungs, and decreased levels of SAP in the sera. To elucidate the role of SAP desialylation in idiopathic pulmonary fibrosis (IPF) pathogenesis, we purified SAP from the serum of IPF patients and healthy controls and measured the extent of sialylation and bioactivity of the purified SAP. We find that some IPF patients have abnormally high levels of the sialidase NEU3 in their sera and that the SAP in the sera of IPF patients has an abnormally high extent of desialylation and an abnormally low ability to inhibit fibrocyte differentiation and induce extracellular IL-10 accumulation by PBMC. These results suggest that SAP desialylation may play a role in IPF pathogenesis and that inhibiting NEU3 could be a potential therapeutic target for IPF.

Serum Amyloid P and a Dendritic Cell-Specific Intercellular Adhesion Molecule-3-Grabbing Nonintegrin Ligand Inhibit High-Fat Diet-Induced Adipose Tissue and Liver Inflammation and Steatosis in Mice.

High-fat diet (HFD)-induced inflammation is associated with a variety of health risks. The systemic pentraxin serum amyloid P (SAP) inhibits inflammation. SAP activates the high-affinity IgG receptor Fcγ receptor I (FcγRI; CD64) and the lectin receptor dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN; CD209). Herein, we show that for mice on an HFD, injections of SAP and a synthetic CD209 ligand (1866) reduced HFD-increased adipose and liver tissue inflammation, adipocyte differentiation, and lipid accumulation in adipose tissue. HFD worsened glucose tolerance test results and caused increased adipocyte size; for mice on an HFD, SAP improved glucose tolerance test results and reduced adipocyte size. Mice on an HFD had elevated serum levels of IL-1ß, IL-23, interferon (IFN)-ß, IFN-γ, monocyte chemoattractant protein 1 [MCP-1; chemokine (C-C motif) ligand 2 (CCL2)], and tumor necrosis factor-α. SAP reduced serum levels of IL-23, IFN-ß, MCP-1, and tumor necrosis factor-α, whereas 1866 reduced IFN-γ. In vitro, SAP, but not 1866, treated cells isolated from white fat tissue (stromal vesicular fraction) produced the anti-inflammatory cytokine IL-10. HFD causes steatosis, and both SAP and 1866 reduced it. Conversely, compared with control mice, SAP knockout mice fed on a normal diet had increased white adipocyte cell sizes, increased numbers of inflammatory cells in adipose and liver tissue, and steatosis; and these effects were exacerbated on an HFD. SAP and 1866 may inhibit some, but not all, of the effects of a high-fat diet.

Monocyte differentiation and macrophage priming are regulated differentially by pentraxins and their ligands.

BACKGROUND: Circulating bone marrow-derived monocytes can leave the blood, enter a tissue, and differentiate into M1 inflammatory, M2a remodeling/fibrotic, or M2c/Mreg resolving/immune-regulatory macrophages. Macrophages can also convert from one of the above types to another. Pentraxins are secreted proteins that bind to, and promote efficient clearance of, microbial pathogens and cellular debris during infection, inflammation, and tissue damage. The pentraxins C-reactive protein (CRP), serum amyloid P (SAP), and pentraxin-3 (PTX3) can also bind a variety of endogenous ligands. As monocytes and macrophages are exposed to differing concentrations of pentraxins and their ligands during infection, inflammation, and tissue damage, we assessed what effect pentraxins and their ligands have on these cells. RESULTS: We found that many polarization markers do not discriminate between the effects of pentraxins and their ligands on macrophages. However, pentraxins, their ligands, and cytokines differentially regulate the expression of the hemoglobin-haptoglobin complex receptor CD163, the sialic acid-binding lectin CD169, and the macrophage mannose receptor CD206. CRP, a pentraxin generally thought of as being pro-inflammatory, increases the extracellular accumulation of the anti-inflammatory cytokine IL-10, and this effect is attenuated by GM-CSF, mannose-binding lectin, and factor H. CONCLUSIONS: These results suggest that the presence of pentraxins and their ligands regulate macrophage differentiation in the blood and tissues, and that CRP may be a potent inducer of the anti-inflammatory cytokine IL-10.