Pathogenic myelin oligodendrocyte glycoprotein antibodies recognize glycosylated epitopes and perturb oligodendrocyte physiology

Antibodies to myelin components are routinely detected in multiple sclerosis patients. However, their presence in some control subjects has made it difficult to determine their contribution to disease pathogenesis. Immunization of C57BL/6 mice with either rat or human myelin oligodendrocyte glycoprotein (MOG) leads to experimental autoimmune encephalomyelitis (EAE) and comparable titers of anti-MOG antibodies as detected by ELISA. However, only immunization with human (but not rat) MOG results in a B cell-dependent EAE. In this study, we demonstrate that these pathogenic and nonpathogenic anti-MOG antibodies have a consistent array of differences in their recognition of antigenic determinants and biological effects. Specifically, substituting proline at position 42 with serine in human MOG (as in rat MOG) eliminates the B cell requirement for EAE. All MOG proteins analyzed induced high titers of anti-MOG (tested by ELISA), but only antisera from mice immunized with unmodified human MOG were encephalitogenic in primed B cell-deficient mice. Nonpathogenic IgGs bound recombinant mouse MOG and deglycosylated MOG in myelin (tested by Western blot), but only pathogenic IgGs bound glycosylated MOG. Only purified IgG to human MOG bound to live rodent oligodendrocytes in culture and, after cross-linking, induced repartitioning of MOG into lipid rafts, followed by dramatic changes in cell morphology. The data provide a strong link between in vivo and in vitro observations regarding demyelinating disease, further indicate a biochemical mechanism for anti-MOG-induced demyelination, and suggest in vitro tools for determining autoimmune antibody pathogenicity in multiple sclerosis patients.     

Antibodies to native myelin oligodendrocyte glycoprotein are serologic markers of early inflammation in multiple sclerosis

Myelin oligodendrocyte glycoprotein (MOG) is an integral membrane protein expressed in CNS oligodendrocytes and outermost myelin lamellae. Anti-MOG Abs cause myelin destruction (demyelination) in animal models of multiple sclerosis (MS); however, such pathogenic Abs have not yet been characterized in humans. Here, a method that specifically detects IgG binding to human MOG in its native, membrane-embedded conformation on MOG-transfected mammalian cells was used to evaluate the significance of these auto Abs. Compared with healthy controls, native MOG-specific IgGs were most frequently found in serum of clinically isolated syndromes (P < 0.001) and relapsing-remitting MS (P < 0.01), only marginally in secondary progressive MS (P < 0.05), and not at all in primary progressive MS. We demonstrate that epitopes exposed in this cell-based assay are different from those exposed on the refolded, extracellular domain of human recombinant MOG tested by solid-phase ELISA. In marmoset monkeys induced to develop MS-like CNS inflammatory demyelination, IgG reactivity against the native membrane-bound MOG is always detected before clinical onset of disease (P < 0.0001), unlike that against other myelin constituents. We conclude that (i) epitopes displayed on native, glycosylated MOG expressed in vivo are early targets for pathogenic Abs; (ii) these Abs, which are not detected in solid-phase assays, might be the ones to play a pathogenic role in early MS with predominant inflammatory activity; and (iii) the cell-based assay provides a practical serologic marker for early detection of CNS autoimmune demyelination including its preclinical stage at least in the primate MS model.     

Autoantibodies in inflammatory demyelinating diseases of the central nervous system

The determination of disease-specific autoantibodies (Abs) is a challenge in any autoimmune disease. The significance of Abs detected in inflammatory demyelinating diseases (IDD) of the central nervous system (CNS), such as multiple sclerosis (MS), is still unclear. Histopathological reports have demonstrated that a humoral (Abs)-mediated pattern of demyelination is detected in >50% of MS patients and is consistently associated with active demyelination. The observation that these patients specifically respond to plasmapheresis reinforces the hypothesis of a specific humoral MS subtype. One of the most intensively studied antigen targets in MS is a glycoprotein of the myelin sheath called the Myelin Oligodendrocyte Glycoprotein (MOG). Recent advances have shown that epitope specificity of MOG is crucial in terms of specificity of the Ab response. Several other auto-Abs, including anti-myelin, oligodendrocyte and neuronal Abs have been studied in MS. These auto-Abs may have pathogenic or protective properties, but could also have no functional role. Recently, the demonstration of a highly specific auto-Ab in an IDD of the CNS called neuromyelitis optica (NMO), directed against the aquaporin-4 (AQP-4) located at the blood brain barrier (BBB), has allowed a refinement of the diagnostic criteria of NMO and classification of this disease as an autoimmune channelopathy. These recent advances have reinforced the interest in tracking the role of the humoral response in the different IDD of the CNS.     

SARS-CoV-2-related Myelin Oligodendrocyte Glycoprotein Antibody-associated Disease: A Case Report and Literature Review

We herein report a case of myelin oligodendrocyte glycoprotein (MOG) antibody-associated disease after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. A 24-year-old woman developed unilateral optic neuritis 3 weeks after contracting coronavirus disease 2019 (COVID-19), followed by intracranial demyelinating lesions and myelitis. Since serum anti-MOG antibody was positive, we diagnosed MOG antibody-associated disease. Immunotherapy with steroids resulted in the rapid improvement of neurological symptoms. This is a suggestive case, as there are no reports of MOG antibody-associated disease with multiple neurological lesions occurring after COVID-19. The response to immunotherapy was favorable. This case suggests that it is important to measure anti-MOG antibodies in patients who develop inflammatory neurological disease after COVID-19.     

Identification of a pathogenic antibody response to native myelin oligodendrocyte glycoprotein in multiple sclerosis

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system. Although the cause of MS is still uncertain, many findings point toward an ongoing autoimmune response to myelin antigens. Because of its location on the outer surface of the myelin sheath and its pathogenicity in the experimental autoimmune encephalomyelitis model, myelin oligodendrocyte glycoprotein (MOG) is one of the potential disease-causing self antigens in MS. However, the role of MOG in the pathogenesis of MS has remained controversial. In this study we addressed the occurrence of autoantibodies to native MOG and its implication for demyelination and axonal loss in MS. We applied a high-sensitivity bioassay, which allowed detecting autoantibodies that bind to the extracellular part of native MOG. Antibodies, mostly IgG, were found in sera that bound with high affinity to strictly conformational epitopes of the extracellular domain of MOG. IgG but not IgM antibody titers to native MOG were significantly higher in MS patients compared with different control groups with the highest prevalence in primary progressive MS patients. Serum autoantibodies to native MOG induced death of MOG-expressing target cells in vitro. Serum from MS patients with high anti-MOG antibody titers stained white matter myelin in rat brain and enhanced demyelination and axonal damage when transferred to autoimmune encephalomyelitis animals. Overall these findings suggest a pathogenic antibody response to native MOG in a subgroup of MS patients.     

The crystal structure of myelin oligodendrocyte glycoprotein,a key autoantigen in multiple sclerosis

Myelin oligodendrocyte glycoprotein (MOG) is a key CNS-specific autoantigen for primary demyelination in multiple sclerosis. Although the disease-inducing role of MOG has been established, its precise function in the CNS remains obscure. To gain new insights into the physiological and immunopathological role of MOG, we determined the 1.8-A crystal structure of the MOG extracellular domain (MOGED). MOGED adopts a classical Ig (Ig variable domain) fold that was observed to form an antiparallel head-to-tail dimer. A dimeric form of native MOG was observed, and MOGED was also shown to dimerize in solution, consistent with the view of MOG acting as a homophilic adhesion receptor. The MOG35-55 peptide, a major encephalitogenic determinant recognized by both T cells and demyelinating autoantibodies, is partly occluded within the dimer interface. The structure of this key autoantigen suggests a relationship between the dimeric form of MOG within the myelin sheath and a breakdown of immunological tolerance to MOG that is observed in multiple sclerosis.     

Diagnostic algorithm for relapsing demyelinating syndromes of the CNS in children

Background

Paediatric relapsing demyelinating syndromes of the CNS define a group of diseases that have different phenotypes. Although for some of them, such as multiple sclerosis and neuromyelitis optica spectrum disorder (NMOSD), diagnostic criteria have been developed, diagnostic uncertainties are not uncommon. We aimed to identify the key features that unify phenotypes, and focused on patients with myelin oligodendrocyte glycoprotein (MOG) antibodies, to investigate whether they show distinctive clinical and radiological features, independently of their original diagnosis. We then generated a diagnostic algorithm for clinical use.

Structural insights into the antigenicity of myelin oligodendrocyte glycoprotein

Multiple sclerosis is a chronic disease of the central nervous system (CNS) characterized by inflammation, demyelination, and axonal loss. The immunopathogenesis of demyelination in multiple sclerosis involves an autoantibody response to myelin oligodendrocyte glycoprotein (MOG), a type I transmembrane protein located at the surface of CNS myelin. Here we present the crystal structures of the extracellular domain of MOG (MOGIgd) at 1.45-A resolution and the complex of MOGIgd with the antigen-binding fragment (Fab) of the MOG-specific demyelinating monoclonal antibody 8-18C5 at 3.0-A resolution. MOGIgd adopts an IgV like fold with the A'GFCC'C" sheet harboring a cavity similar to the one used by the costimulatory molecule B7-2 to bind its ligand CTLA4. The antibody 8-18C5 binds to three loops located at the membrane-distal side of MOG with a surprisingly dominant contribution made by MOG residues 101-108 containing a strained loop that forms the upper edge of the putative ligand binding site. The sequence R101DHSYQEE108 is unique for MOG, whereas large parts of the remaining sequence are conserved in potentially tolerogenic MOG homologues expressed outside the immuno-privileged environment of the CNS. Strikingly, the only sequence identical to DHSYQEE was found in a Chlamydia trachomatis protein of unknown function, raising the possibility that Chlamydia infections may play a role in the MOG-specific autoimmune response in man. Our data provide the structural basis for the development of diagnostic and therapeutic strategies targeting the pathogenic autoantibody response to MOG.     

Infectious Mononucleosis Triggers Generation of IgG Auto-Antibodies against Native Myelin Oligodendrocyte Glycoprotein

A history of infectious mononucleosis (IM), symptomatic primary infection with the Epstein Barr virus, is associated with the development of autoimmune diseases and increases the risk to develop multiple sclerosis. Here, we hypothesized that immune activation during IM triggers autoreactive immune responses. Antibody responses towards cellular antigens using a HEp-2 based indirect immunofluorescence assay and native myelin oligodendrocyte glycoprotein (MOG) using a flow cytometry-based assay were determined in 35 patients with IM and in 23 control subjects. We detected frequent immunoglobulin M (IgM) reactivity to vimentin, a major constituent of the intermediate filament family of proteins, in IM patients (27/35; 77%) but rarely in control subjects (2/23; 9%). IgG autoantibodies binding to HEp-2 cells were absent in both groups. In contrast, IgG responses to native MOG, present in up to 40% of children with inflammatory demyelinating diseases of the central nervous system (CNS), were detectable in 7/35 (20%) patients with IM but not in control subjects. Normalization of anti-vimentin IgM levels to increased total IgM concentrations during IM resulted in loss of significant differences for anti-vimentin IgM titers. Anti-MOG specific IgG responses were still detectable in a subset of three out of 35 patients with IM (9%), even after normalization to increased total IgG levels. Vimentin-specific IgM and MOG-specific IgG responses decreased following clinical resolution of acute IM symptoms. We conclude from our data that MOG-specific memory B cells are activated in subset of patients with IM.     

Identification of human neutralizing antibodies that bind to complex epitopes on dengue virions

Dengue is a mosquito-borne flavivirus that is spreading at an unprecedented rate and has developed into a major health and economic burden in over 50 countries. Even though infected individuals develop potent and long-lasting serotype-specific neutralizing antibodies (Abs), the epitopes engaged by human neutralizing Abs have not been identified. Here, we demonstrate that the dengue virus (DENV)-specific serum Ab response in humans consists of a large fraction of cross-reactive, poorly neutralizing Abs and a small fraction of serotype-specific, potently inhibitory Abs. Although many mouse-generated, strongly neutralizing monoclonal antibodies (mAbs) recognize epitopes that are present on recombinant DENV envelope (E) proteins, unexpectedly, the majority of neutralizing Abs in human immune sera bound to intact virions but not to the ectodomain of purified soluble E proteins. These conclusions with polyclonal Abs were confirmed with newly generated human mAbs derived from DENV-immune individuals. Two of three strongly neutralizing human mAbs bound to E protein epitopes that were preserved on the virion but not on recombinant E (rE) protein. We propose that humans produce Abs that neutralize DENV infection by binding a complex, quaternary structure epitope that is expressed only when E proteins are assembled on a virus particle. Mapping studies indicate that this epitope has a footprint that spans adjacent E protein dimers and includes residues at the hinge between domains I and II of E protein. These results have significant implications for the DENV Ab and vaccine field.     

Elicitation of structure-specific antibodies by epitope scaffolds

Elicitation of antibodies against targets that are immunorecessive, cryptic, or transient in their native context has been a challenge for vaccine design. Here we demonstrate the elicitation of structure-specific antibodies against the HIV-1 gp41 epitope of the broadly neutralizing antibody 2F5. This conformationally flexible region of gp41 assumes mostly helical conformations but adopts a kinked, extended structure when bound by antibody 2F5. Computational techniques were employed to transplant the 2F5 epitope into select acceptor scaffolds. The resultant “2F5-epitope scaffolds” possessed nanomolar affinity for antibody 2F5 and a range of epitope flexibilities and antigenic specificities. Crystallographic characterization of the epitope scaffold with highest affinity and antigenic discrimination confirmed good to near perfect attainment of the target conformation for the gp41 molecular graft in free and 2F5-bound states, respectively. Animals immunized with 2F5-epitope scaffolds showed levels of graft-specific immune responses that correlated with graft flexibility (p < 0.04), while antibody responses against the graft—as dissected residue-by-residue with alanine substitutions—resembled more closely those of 2F5 than sera elicited with flexible or cyclized peptides, a resemblance heightened by heterologous prime-boost. Lastly, crystal structures of a gp41 peptide in complex with monoclonal antibodies elicited by the 2F5-epitope scaffolds revealed that the elicited antibodies induce gp41 to assume its 2F5-recognized shape. Epitope scaffolds thus provide a means to elicit antibodies that recognize a predetermined target shape and sequence, even if that shape is transient in nature, and a means by which to dissect factors influencing such elicitation.     

Anti-phosphatidylserine-prothrombin complex antibodies in patients with localized scleroderma

OBJECTIVE: Although some antiphospholipid antibodies (Abs) are found in patients with localized scleroderma (LSc), Ab against phosphatidylserine-prothrombin complex (PS/PT) has not been examined. We investigated anti-PS/PT Ab levels in patients with LSc. METHODS: IgG anti-PS/PT Ab levels in serum samples taken from patients with LSc (n = 42) were measured using ELISA. RESULTS: IgG anti-PS/PT Ab was detected in 17% of the LSc patients, while it was not detected in any normal controls (n = 32) or psoriasis vulgaris (n = 25), and this frequency was similar to that of systemic sclerosis (17%, n = 41). Among 3 LSc subgroups, generalized morphea, the severest form of LSc, had a frequency (27%) comparable with that of systemic lupus erythematosus (32%, n = 25). Among 7 LSc patients with anti-PS/PT Ab, 2 developed symptomatic thromboembolism (A 70-year-old man developed deep vein thrombosis and pulmonary infarction, although he was negative for other antiphospholipid Abs. A 6-year-old boy positive for lupus anticoagulant had cerebral infarction). By contrast, symptomatic thromboembolism was not detected in 35 LSc patients without anti-PS/PT Ab. CONCLUSION: Patients with LSc, especially generalized morphea, exhibit anti-PS/PT Ab at a frequency comparable with collagen diseases such as systemic sclerosis and systemic lupus erythematosis. Examination of this Ab may be useful to recognize the risk of thromboembolism in patients with LSc.     

HIV-1 CTL-based vaccine immunogen selection: antigen diversity and cellular response features

Much of the current effort in HIV-1 vaccine design is directed at achieving T-cell immunity that will result in enough immunological memory to contain HIV-1 infection after acquisition. However, antigenic diversity, plus a lack of understanding of HIV-1 vaccine immunology, have hindered the development of a globally effective cytotoxic T-lymphocyte (CTL)-based vaccine. Cellular response, in using a finite immune system to recognize an infinite number of potential pathogens, exhibits a series of parsimonious features. These features are considered critical in modulating HIV-1 vaccine multiple specificities. We took the features into consideration when the potential epitope coverage (E(c)) to circulating strains by current vaccine strategies was analyzed. Based on these analyses, several approaches are proposed to enhance the breadth of vaccine responses and, hence, the potential protective efficacy.     

Discordant memory B cell and circulating anti-Env antibody responses in HIV-1 infection

Long-lived memory B cells (B_Mem) provide an archive of historic Ab responses. By contrast, circulating Abs typically decline once the immunogen is cleared. Consequently, circulating Abs can underestimate the nature of cognate humoral immunity. On the other hand, the B_Mem pool should provide a comprehensive picture of Ab specificities that arise over the entire course of infection. To test this hypothesis, we compared circulating Ab and B_Mem from natural virus suppressors who control HIV-1 without therapy and maintain a relatively intact immune system. We found high frequencies of B_Mem specific for the conserved neutralizing CD4 induced or CD4 binding site epitopes of gp120, whereas low Ab titers to these determinants were detected in contemporaneous plasma. These data suggest that plasma Ab repertoires can underestimate the breadth of humoral immunity, and analyses of B_Mem should be included in studies correlating Ab specificity with protective immunity to HIV-1.     

Unilateral cerebral cortical encephalitis (CCE) with positive anti-MOG antibodies: A case report

Rationale:Nowadays, myelin oligodendrocyte glycoprotein (MOG)-antibody-associated disease (MOGAD) is regarded as an independent inflammatory demyelinating disease. Here, we report a rare case of unilateral cerebral cortical encephalitis (CCE) with positive anti-MOG antibodies.Patient concerns:A 19-year-old woman was admitted to our hospital owing to acute onset fever and headache. Four days later, she experienced a focal seizure that progressed to generalized tonic-clonic seizures.Diagnosis:Brain magnetic resonance imaging (MRI) demonstrated cortical lesions in the left cerebral hemisphere on T2-weighted fluid-attenuated inversion recovery imaging. The patient was positive for anti-MOG antibodies in serum and diagnosed with anti-MOG antibody-associated unilateral CCE.Interventions:She was administrated with intravenous methylprednisolone followed by oral corticosteroids.Outcomes:On day 14 after admission, a repeat MRI revealed partial resolution of the initial abnormalities. The patient received a quick recovery without residual symptoms.Conclusions:Unilateral CCE with positive anti-MOG antibodies has emerged as a special clinical phenotype of MOGAD. It should be emphasized that the characteristic neuroradiological features of CCE would be an important clue to the correct diagnosis of MOGAD.     

Mapping of distinct von Willebrand factor domains interacting with platelet GPIb and GPIIb/IIIa and with collagen using monoclonal antibodies

We have used monoclonal antibodies (M Abs) and proteolytic fragmentation to localize structurally the functional sites of human von Willebrand factor (vWF) responsible for interaction with membrane glycoproteins GPIb, GPIIb/IIIa, and with collagen. SpII (215 kd) and SpIII (320 kd), the S aureus V-8 protease homodimeric fragments representing the carboxy-terminal and amino-terminal segments of the vWF subunit, competitively inhibited the binding of multimeric vWF to thrombin-stimulated or ristocetin-stimulated platelets, respectively. Specific saturable binding of each fragment was observed to stimulate platelets appropriately and was inhibited only by selected M Abs that both bound to the specific fragment and inhibited the corresponding function. M Ab 9, which blocks thrombin-induced binding of vWF to platelets, inhibited binding of SpII to platelets and bound to SpII as well as to a dimeric, 86-kd thermolysin fragment composed of 42-kd and 23-kd subunits, each possessing the epitope. Binding of SpII was also inhibited by a M Ab to GPIIb/IIIa. Thus, it appears that a portion of the carboxy-terminal end of vWF contains the ligand site for the GPIIb/IIIa receptor. In contrast, M Ab H9, which blocks ristocetin- induced binding of vWF to platelets, inhibited binding of SpIII to platelets and bound to SpIII as well as to monomeric 33-kd and 28-kd subtilisin fragments. Binding of SpIII to platelets was also inhibited by a M Ab to GPIb. Thus, it appears that a small segment of the amino- terminal part of vWF contains the ligand for the platelet GPIb receptor. The collagen binding site of vWF was localized with M Ab B203, which inhibits vWF interaction with collagen. This M Ab also bound to SpIII as well as to monomeric 26-kd and 23-kd subtilisin fragments. Thus, the third functional site responsible for collagen binding appears to be localized on the amino-terminal portion of vWF, in a linear sequence different from those responsible for interaction with either of the platelet receptors. These assignments of functional sites should facilitate the localization of structural defects of vWF in the various forms of vWD and support the role of vWF as an adhesive protein with multiple interactive sites.     

Tumor-specific Ab-mediated targeting of MHC-peptide complexes induces regression of human tumor xenografts in vivo

A cancer immunotherapy strategy is described herein that combines the advantage of the well established tumor targeting capabilities of high-affinity recombinant fragments of Abs with the known efficient, specific, and potent killing ability of CD8 T lymphocytes directed against highly antigenic MHC-peptide complexes. Structurally, it consists of a previously uncharacterized class of recombinant chimerical molecules created by the genetic fusion of single-chain (sc) Fv Ab fragments, specific for tumor cell surface antigens, to monomeric scHLA-A2 complexes containing immunodominant tumor- or viral-specific peptides. The fusion protein can induce very efficiently tumor cell lysis, regardless of the expression of self peptide-MHC complexes. Moreover, these molecules exhibited very potent antitumor activity in vivo in nude mice bearing preestablished human tumor xenografts. These in vitro and in vivo results suggest that recombinant scFv-MHC-peptide fusion molecules could represent an approach to immunotherapy, bridging Ab and T lymphocyte attack on cancer cells.     

Crystal structure of an anti-carbohydrate antibody directed against Vibrio cholerae O1 in complex with antigen: molecular basis for serotype specificity

The crystal structure of the murine Fab S-20-4 from a protective anti-cholera Ab specific for the lipopolysaccharide Ag of the Ogawa serotype has been determined in its unliganded form and in complex with synthetic fragments of the Ogawa O-specific polysaccharide (O-SP). The upstream terminal O-SP monosaccharide is shown to be the primary antigenic determinant. Additional perosamine residues protrude outwards from the Ab surface and contribute only marginally to the binding affinity and specificity. A complementary water-excluding hydrophobic interface and five Ab-Ag hydrogen bonds are crucial for carbohydrate recognition. The structure reported here explains the serotype specificity of anti-Ogawa Abs and provides a rational basis toward the development of a synthetic carbohydrate-based anti-cholera vaccine.     

Antibodies against denatured and reduced thyroid microsomal antigen in autoimmune thyroid disease

Different antigenic determinants on thyroid microsomal antigen (MAg) could induce different antibodies, which, in turn, could have differing importance in the pathogenesis of autoimmune thyroid disease. In this study we demonstrated three types of microsomal antibodies (MAbs) present in serum of patients with autoimmune thyroid disease by Western blot analysis. These MAbs reacted with native, denatured, and denatured and reduced MAg. Methods for detecting the MAbs were developed, and the clinical significance of these Abs was analyzed in 197 patients with thyroid disorders. For measurement of Ab against native and denatured MAg, an enzyme-linked immunosorbent assay, in which wells were coated with untreated or sodium dodecyl sulfate-denatured MAg, was used. For measurement of Ab against denatured and reduced MAg, 6% SDS-polyacrylamide gel electrophoresis followed by Western blot analysis was used. Native MAb enzyme-linked immunosorbent assay values correlated highly with microsomal hemagglutination titers, but patients with high titers of Ab against native MAg did not necessarily have Ab against denatured MAg or reduced MAg. Abs against denatured MAg or denatured and reduced MAg were found in patients with Hashimoto's disease (28.8% and 13.6%) and Graves' disease (22.4% and 11.2%). The percentage of patients with positive denatured and reduced MAb was higher in Graves' disease patients who had a longer sick interval or who developed hypothyroidism after radioiodine treatment. This study provides the first clear demonstration that MAbs are heterogeneous. The data suggest that antibodies against denatured or denatured and reduced MAg may be related to destruction of the thyroid gland.