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B cells rewire complement for optimal germinal center responses. When B cell C3 activity is limited, germinal centers collapse and outputs are impaired.

Binding of the complement component C1q to the CH2 domain of antigen-bound immunoglobulin gamma (IgG) activates the classical complement pathway and depends on its close proximity to Fc fragments of neighboring antibodies. IgG subclasses contain a highly conserved asparagine 297 (N)-linked biantennary glycan within their CH2 domains, the core structure of which can be extended with terminal galactose and sialic acid residues. To investigate whether Fc-glycosylation regulates effector functions of human IgG subclasses, we cloned the antigen-binding region of the CD20-specific monoclonal antibody rituximab into IgG isotype expression vectors. We found that Fc-galactosylation enhances the efficacy of CD20-targeting complement-fixing antibodies for C1q binding and complement-mediated tumor cell lysis. Increased efficacies were restricted to IgG1 and IgG3 subclasses indicating that Fc-galactosylation alone is not sufficient for IgG2 and IgG4 to acquire complement-fixing properties. Addition of terminal galactose to the N-glycan specifically improved binding of C1q without changing antigen- and FcγRIIIa-binding affinities of IgG isotypes. These data indicate that Fc galactosylation can be harnessed to enhance the complement-activating properties of IgG1 and IgG3 antibodies.

Immunoglobulin gamma (IgG) antibodies are key effector proteins of the immune system. They recognize antigens with high specificity and are indispensable for immunological memory following pathogen exposure or vaccination. The constant, crystallizable fragment (Fc) of IgG molecules mediates antibody effector functions such as complement-dependent cytotoxicity, antibody-mediated cellular cytotoxicity, and antibody-dependent cell-mediated phagocytosis. These functions are regulated by a single N-linked, biantennary glycan of the heavy chain, which resides just below the hinge region, and the presence of specific sugar moieties on the glycan has profound implications on IgG effector functions. Emerging knowledge of how Fc glycans contribute to IgG structure and functions has opened new avenues for the therapeutic exploitation of defined antibody glycoforms in the treatment of cancer and autoimmune diseases. Here, we review recent advances in understanding proinflammatory IgG effector functions and their regulation by Fc glycans.

Humoral immune responses require germinal centres (GC) for antibody affinity maturation. Within GC, B cell proliferation and mutation are segregated from affinity-based positive selection in the dark zone (DZ) and light zone (LZ) substructures, respectively. While IL-21 is known to be important in affinity maturation and GC maintenance, here we show it is required for both establishing normal zone representation and preventing the accumulation of cells in the G1 cell cycle stage in the GC LZ. Cell cycle progression of DZ B cells is unaffected by IL-21 availability, as is the zone phenotype of the most highly proliferative GC B cells. Collectively, this study characterises the development of GC zones as a function of time and B cell proliferation and identifies IL-21 as an important regulator of these processes. These data help explain the requirement for IL-21 in normal antibody affinity maturation. How IL-21 functions during development of high affinity antibody in germinal centres (GC) is not fully known. Here using a cell cycle reporter mouse, the authors show that IL-21 promotes cell cycle progression within the GC light zone and enables release from the G1 cell cycle stage.

IgG molecules exert both pro- and antiinflammatory effector functions based on the composition of the fragment crystallizable (Fc) domain glycan. Sialylated IgG Fc domains have antiinflammatory properties that are attributed to their ability to increase the activation threshold of innate effector cells to immune complexes by stimulating the upregulation of the inhibitory Fcγ receptor IIB (FcγRIIB). Here, we report that IgG Fc sialylation of human monoclonal IgG1 molecules impairs their efficacy to induce complement-mediated cytotoxicity (CDC). Fc sialylation of a CD20-targeting antibody had no impact on antibody-dependent cellular cytotoxicity and did not change the affinity of the antibody for activating Fcγ receptors. In contrast, the presence of sialic acid abrogated the increased binding of C1q to Fc-galactosylated IgG1 and resulted in decreased levels of C3b deposition on the cell surface. Similar to monoclonal antibodies, sialic acid inhibited the increased C1q binding to galactosylated Fc fragments in human polyclonal IgG. In sera derived from patients with chronic inflammatory demyelinating polyneuropathy, an autoimmune disease of the peripheral nervous system in which humoral immune responses mediate tissue damage, induction of IgG Fc sialylation was associated with clinical disease remission. Thus, impairment of CDC represents an FcγR-independent mechanism by which Fc-sialylated glycovariants might limit proinflammatory IgG effector functions.

Immunoglobulin G (IgG) molecules are glycoproteins and residues in the sugar moiety attached to the IgG constant fragment (Fc) are essential for IgG functionality such as binding to cellular Fc receptors and complement activation. The core of this sugar moiety consists of a bi-antennary heptameric structure of mannose and N-acetylglucosamine (GlcNAc), further decorated with terminal and branching residues including galactose, sialic acid, fucose, and GlcNAc. Presence or absence of distinct residues such as fucose and sialic acid can dramatically alter pro- and anti-inflammatory IgG activities which could be harnessed for immunotherapeutic purposes. Here we review recent advances in understanding the role of the IgG-Fc glycan during immune responses and for immunotherapy with a focus on sialic acid and intravenous immunoglobulin (IVIG) treatment.

Effective vaccines elicit neutralizing antibodies and long-lasting memory, but this can be challenging with some pathogens, such as HIV. A new study shows how a slow-delivery protein immunization strategy administered in dose-escalation format over 12 days increased the durability of germinal centers and improved immunological outcomes.

Intravenous immunoglobulin (IVIG) are purified IgG preparations made from the pooled plasma from thousands of healthy donors and are being tested in preclinical mouse models. Inherent challenges, however, are the pluripotency of IVIG and its xenogeneicity in animals. IVIG can alter the viability of human neutrophils via agonistic antibodies to Fas and Siglec-9. In this study, we compared the effects of IVIG on human and mouse neutrophils using different death assays. Different commercial IVIG preparations similarly induced cytokine-dependent death in human neutrophils, whereas they had no effects on the survival of either peripheral blood or bone marrow neutrophils from C57BL/6 or BALB/c mice. F(ab’) 2 but not Fc fragments of IVIG induced death of human neutrophils, whereas neither of these IVIG fragments, nor agonistic monoclonal antibodies to human Fas or Siglec-9 affected the viability of mouse neutrophils. Pooled mouse IgG, which exhibited a different immunoprofile compared to IVIG, also had no effect on mouse cells. Together, these observations demonstrate that effects of IVIG on neutrophil survival are not adequately reflected in current mouse models, despite the key role of these cells in human inflammatory and autoimmune diseases.

Although reactivation and accumulation of autoreactive CD4⁺ T cells within the CNS are considered to play a key role in the pathogenesis of multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE), the mechanisms of how these cells recognize their target organ and induce sustained inflammation are incompletely understood. Here, we report that mice with conditional deletion of the essential autophagy protein ATG5 in classical dendritic cells (DCs), which are present at low frequencies in the nondiseased CNS, are completely resistant to EAE development following adoptive transfer of myelin-specific T cells and show substantially reduced in situ CD4⁺ T cell accumulation during the effector phase of the disease. Endogenous myelin peptide presentation to CD4⁺ T cells following phagocytosis of injured, phosphatidylserine-exposing oligodendroglial cells is abrogated in the absence of ATG5. Pharmacological inhibition of ATG-dependent phagocytosis by the cardiac glycoside neriifolin, an inhibitor of the Na⁺, K⁺-ATPase, delays the onset and reduces the clinical severity of EAE induced by myelin-specific CD4⁺ T cells. These findings link phagocytosis of injured oligodendrocytes, a pathological hallmark of MS lesions and during EAE, with myelin antigen processing and T cell pathogenicity, and identify ATG-dependent phagocytosis in DCs as a key regulator in driving autoimmune CD4⁺ T cell-mediated CNS damage.

The B cell-depleting IgG1 monoclonal antibody rituximab can persistently suppress disease progression in some patients with autoimmune diseases. However, the mechanism underlying these long-term beneficial effects has remained unclear. Here, we evaluated Ig gene usage in patients with anti-myelin-associated glycoprotein (anti-MAG) neuropathy, an autoimmune disease of the peripheral nervous system that is mediated by IgM autoantibodies binding to MAG antigen. Patients with anti-MAG neuropathy showed substantial clonal expansions of blood IgM memory B cells that recognized MAG antigen. The group of patients showing no clinical improvement after rituximab therapy were distinguished from clinical responders by a higher load of clonal IgM memory B cell expansions before and after therapy, by persistence of clonal expansions despite efficient peripheral B cell depletion, and by a lack of substantial changes in somatic hypermutation frequencies of IgM memory B cells. We infer from these data that the effectiveness of rituximab therapy depends on efficient depletion of noncirculating B cells and is associated with qualitative immunological changes that indicate reconfiguration of B cell memory through sustained reduction of autoreactive clonal expansions. These findings support the continued development of B cell-depleting therapies for autoimmune diseases.