The IgG-specific endoglycosidase EndoS inhibits both cellular and complement-mediated autoimmune hemolysis

Abstract

EndoS from IntroductionThe presence and pathogenicity of autoantibodies directed against red blood cells (RBCs) have been extensively investigated and implicated in a number of autoimmune diseases, such as autoimmune hemolytic anemia (AIHA) 1 and systemic lupus erythematosus. 2 The RBC surface contains many epitopes recognizable by alloantibodies and autoantibodies, including the more than 300 blood group antigens located on glycoproteins and glycolipids but also nonpolymorphic/nonblood group protein structures. 3 The most common targets of pathogenic RBC autoantibodies are epitopes on the Rh protein, encoded by 2 homologous genes on chromosome 1, RHD and RHCE. 4 More than half of all autoantibody specificities in patients with AIHA of the common so-called warm type include immunoglobulin G (IgG) antibodies against these Rh epitopes, which are consequently often used to model AIHA experimentally.Human antibodies against RhD (anti-D) or rabbit antibodies against human RBC (anti-RBC) can result in accelerated phagocytosis and removal of RBC from the circulation by monocytes/ macrophages in the spleen and liver. 5,6 Fc␥ receptor (Fc␥R)-mediated erythrophagocytosis, in addition to other IgG effector functions, such as activated complement, oxidative burst, and cytokine production by Fc␥R expressing effectors cells, play an essential role in shortened RBC survival. [7][8][9][10][11] The ability of normal human monocytes to bind and phagocyte IgG-sensitized RBCs via Fc␥R and mediate immune RBC destruction is highly related to the glycosylation status of the Fc domain on the anti-RBC antibodies. [12][13][14][15][16][17] Endoglycosidase S (EndoS) is secreted by Streptococcus pyogenes and has a specific endoglycosidase activity on native IgG by hydrolyzing the conserved asparagine-linked glycans on the heavy chains of IgG. 18,19 EndoS is the first known bacterial enzyme with a unique specificity for native IgG. EndoS affects the functionality of opsonizing IgG by decreased binding to Fc␥Rs on a monocyte-like cell line and impaired classic complement activation in vitro. 19 EndoS has recently been shown to modulate human IgG/Fc␥R interactions by influencing the binding/dissociation of IgG to soluble and cell-bound Fc␥R. 20 Furthermore, the pretreatment of pathogenic antibodies by EndoS has a protective function in a mouse model of collagen-induced arthritis, and EndoS cures mice from lethal IgG-driven immune thrombocytopenic purpura (ITP). 21,22 The present study was undertaken to establish the effects of EndoS on antibody-mediated destruction of RBCs with a future therapeutic application in mind. Here we show, using monocytes from blood and the THP-1 monocytic cell line, that pretreatment of anti-RBC with EndoS abrogates erythrophagocytosis and subsequent activation of monocytes. The complement-dependent hemolytic effects of anti-RBC added to whole human blood were greatly decreased by pretreatment of this antibody with EndoS. Finally and most importantly, EndoS showed inhibitory effects on a model of AIHA in mice. These...

A Novel Assay for Antibody-Dependent Cell-Mediated Cytotoxicity against HIV-1- or SIV-Infected Cells Reveals Incomplete Overlap with Antibodies Measured by Neutralization and Binding Assays

“…Streptococcus pyogenes encodes an enzyme, endoglycosidase S (EndoS), which specifically hydrolyzes this N-linked glycan (3,27). Treatment of IgG with EndoS inhibits ADCC and complement-mediated lysis (2). To confirm that the cytotoxicity measured in our assay is dependent upon an Fc receptor-mediated interaction, we compared EndoS-treated IgG with untreated IgG (Fig.…”

IgG glycan hydrolysis by endoglycosidase S diminishes the proinflammatory properties of immune complexes from patients with systemic lupus erythematosus: A possible new treatment?

“…Since then, several studies have shown beneficial effects of EndoS in the treatment of disease in animal models (21,22,49,50) as well as in in vitro experiments (20,27,36). However, the exact mechanism of how EndoS would operate in the human in vitro system was not known.…”

“…The serum proteins were separated using NuPAGE 4-12% gradient gels (Invitrogen). Western blot analysis using antibodies against human IgG and LCA were performed as described previously (27). Human IgG (Immuno) was heat aggregated for 30 minutes at 63°C to create IC-like IgG complexes.…”

Antibody-mediated rejection across solid organ transplants: manifestations, mechanisms, and therapies

“…Alternatively, an anti-C1s antibody is in clinical trials for the treatment of complement-mediated diseases (NCT02502903). Since the capacity of IgG to activate complement depends on interaction with C1q via the glycosylated CH2 domain in the Fc region, other possible therapies to attenuate complementmediated injury include the Staphylococcus aureus enzymes IdeS and EndoS (111)(112)(113). These enzymes cause cleavage of the Fc tail (IdeS) or of the N-linked glycan (EndoS) and abolish complement activation in animal models of antibody-mediated inflammation (111)(112)(113)(114).…”

“…Since the capacity of IgG to activate complement depends on interaction with C1q via the glycosylated CH2 domain in the Fc region, other possible therapies to attenuate complementmediated injury include the Staphylococcus aureus enzymes IdeS and EndoS (111)(112)(113). These enzymes cause cleavage of the Fc tail (IdeS) or of the N-linked glycan (EndoS) and abolish complement activation in animal models of antibody-mediated inflammation (111)(112)(113)(114). Clinical trials for desensitization with IdeS are currently under way in Europe and the United States (NCT02426684); however, such approaches are likely to be limited because of (a) risk of allergic reaction due to antibody responses against these bacterial enzymes, and (b) reconstitution of circulating IgG, which occurs in 2-3 weeks.…”