Thomas F. Tedder

Multiple sclerosis (MS) is a debilitating autoimmune disease of the CNS, which is characterized by demyelination and axonal injury and frequently preceded by a demyelinating event called clinically isolated syndrome (CIS). Despite the importance of B cells and autoantibodies in MS pathology, their target specificities remain largely unknown. For an agnostic and comprehensive evaluation of autoantibodies in MS, we developed and employed what we believe to be a novel autoantigen discovery technology, the Antigenome Platform. This Platform is a high-throughput assay comprising large-fragment (approximately 100 amino acids) cDNA libraries, phage display, serum antibody screening technology, and robust bioinformatics analysis pipelines. For autoantibody discovery, we assayed serum samples from CIS patients who received either placebo or treatment who were enrolled in the REFLEX clinical trial, which assessed the effects of IFN-β-1a (Rebif) clinical and MRI activity in patients with CIS. Serum autoantibodies from patients with CIS were significantly and reproducibly enriched for known and previously unreported protein targets; 166 targets were selected by over 10% of patients’ sera. Further, 10 autoantibody biomarkers associated with disease activity and 17 associated with patient response to IFN-β-1a therapy. These findings indicate widespread autoantibody production in MS and provide biomarkers for continued study and prediction of disease progression.

D2C7-immunotoxin (IT), a dual-specific IT targeting wild-type epidermal growth factor receptor (EGFR) and mutant EGFR variant III (EGFRvIII) proteins, demonstrates encouraging survival outcomes in a subset of patients with glioblastoma. We hypothesized that immunosuppression in glioblastoma limits D2C7-IT efficacy. To improve the response rate and reverse immunosuppression, we combined D2C7-IT tumor cell killing with αCD40 costimulation of antigen-presenting cells. In murine glioma models, a single intratumoral injection of D2C7-IT+αCD40 treatment activated a proinflammatory phenotype in microglia and macrophages, promoted long-term tumor-specific CD8 ⁺ T cell immunity, and generated cures. D2C7-IT+αCD40 treatment increased intratumoral Slamf6 ⁺ CD8 ⁺ T cells with a progenitor phenotype and decreased terminally exhausted CD8 ⁺ T cells. D2C7-IT+αCD40 treatment stimulated intratumoral CD8 ⁺ T cell proliferation and generated cures in glioma-bearing mice despite FTY720-induced peripheral T cell sequestration. Tumor transcriptome profiling established CD40 up-regulation, pattern recognition receptor, cell senescence, and immune response pathway activation as the drivers of D2C7-IT+αCD40 antitumor responses. To determine potential translation, immunohistochemistry staining confirmed CD40 expression in human GBM tissue sections. These promising preclinical data allowed us to initiate a phase 1 study with D2C7-IT+αhCD40 in patients with malignant glioma (NCT04547777) to further evaluate this treatment in humans.

The protein tyrosine phosphatase CD45 plays a crucial role in B cell antigen receptor (BCR) signaling by activating Src family kinases. Cd45 ^−/− mice show altered B cell development and a phenotype likely due to reduced steady-state signaling; however, Cd45 ^−/− B cells show relatively normal BCR ligation–induced signaling. In our investigation of how BCR signaling was restored in Cd45 ^−/− cells, we found that the coreceptor CD22 switched from an inhibitory to a stimulatory function in these cells. We disrupted the ability of CD22 to interact with its ligands in Cd45 ^−/− B cells by generating Cd45 ^−/− St6galI ^−/− mice, which cannot synthesize the glycan ligand of CD22, or by treating Cd45 ^−/− B cells in vitro with the sialoside GSC718, which inhibits ligand binding to CD22. BCR ligation–induced signaling was reduced by ST6GalI deficiency, but not by GSC718 treatment, suggesting that CD22 restored BCR ligation–induced signaling in Cd45 ^−/− mature B cells by altering cellular phenotypes during development. CD22 was required for the increase in the surface amount of IgM-BCR on Cd45 ^−/− B cells, which augmented signaling. Because B cell survival depends on steady-state BCR signaling, IgM-BCR abundance was likely increased by the selective survival of IgM-BCR ^hi Cd45 ^−/− B cells because of CD22-mediated signaling under conditions of substantially reduced steady-state signaling. Because the amount of surface IgM-BCR is increased on B cells from patients with other BCR signaling deficiencies, including X-linked agammaglobulinemia, our findings suggest that CD22 may contribute to the partial restoration of B cell function in these patients.

Objective: Regulatory B cells that inhibit immune responses through interleukin‐10 (IL‐10) secretion (B10 cells) have been characterized in adult subjects with autoimmune disease. The aim of this study was to characterize B10 cells in individuals across the entire age range of normal human development and changes in their frequency and numbers in children with autoimmunity.Methods: The phenotype and numbers of B10 cells in blood were examined in healthy individuals and children with autoimmunity, using flow cytometry. B10 cell function was assessed by measuring the effect of B cell–derived IL‐10 on interferon‐γ (IFNγ) expression by CD4+ T cells. Serum cytokine levels were measured by enzyme‐linked immunosorbent assay.Results: The frequency of B10 cells transiently increased during childhood, when up to 30% of B cells were competent to produce IL‐10, compared with the low frequencies in healthy newborns (3–4%) and adults (7–9%). The surface phenotype of B10 cells in children revealed age‐dependent variability. B10 cells from children were distinct from proinflammatory cytokine–producing B cells and down‐regulated IFNγ production by CD4+ T cells in vitro. Compared with age‐matched healthy controls, children with autoimmunity had lower numbers and frequencies of B10 cells (decreased by 39% and 48%, respectively), higher IFNγ levels, and lower IL‐21 levels in serum. IFNγ inhibited, whereas IL‐21 promoted, B cell IL‐10 competence in vitro.Conclusion: B10 cells, a functionally defined cell subset with a variable surface phenotype reflective of overall B cell development, transiently expand during childhood. B10 cell frequencies and numbers were decreased in children with autoimmunity, which may be explained in part by alterations in serum IFNγ and IL‐21 that differentially regulate B10 cell development.

B cells are known to instigate and promulgate immune responses by producing antibodies and presenting antigens to T cells. However, a rare but potent B-cell subset in both humans and mice is capable of inhibiting immune responses through the production of the anti-inflammatory cytokine IL-10. Regulatory B cells do not express any unique combination of surface markers but instead represent a small population of B cells that have acquired the unique ability to produce IL-10. This numerically rare B-cell subset is therefore functionally referred to as ‘B10 cells’ to reflect both their molecular program and the fact that their anti-inflammatory effects in models of autoimmunity, infection and cancer are solely attributable to IL-10 production. As with most B cells, B10 cell development and function appear to be predominantly, if not exclusively, driven by antigen-receptor signals. Once generated, B10 cells respond to both innate and adaptive immune signals, with a requirement for antigen-specific local interactions with T cells to induce IL-10 production and to provide optimal immune suppression in mouse models of autoimmune disease. B10 cells therefore provide an antigen-specific mechanism for delivering IL-10 locally to sites of immune activation and inflammation. The ability of B10 cells to regulate innate and adaptive immune responses makes them an ideal therapeutic target for the treatment of many immune-related disorders.

B cells are commonly thought to enhance inflammatory immune responses. However, specific regulatory B cell subsets recently were identified that downregulate adaptive and innate immunity, inflammation, and autoimmunity through diverse molecular mechanisms. In both mice and humans, a rare, but specific, subset of regulatory B cells is functionally characterized by its capacity to produce IL-10, a potent inhibitory cytokine. For clarity, this regulatory B cell subset has been labeled as B10 cells, because their ability to downregulate immune responses and inflammatory disease is fully attributable to IL-10, and their absence or loss exacerbates disease symptoms in mouse models. This review preferentially focuses on what is known about mouse B10 cell development, phenotype, and effector function, as well as on mechanistic studies that demonstrated their functional importance during inflammation, autoimmune disease, and immune responses.

B cells provide humoral protection against pathogens and promote cellular immunity through diverse nonclassical effector functions. To assess B cell function in promoting T cell homeostasis, mature B cells were either acutely or chronically depleted in mice using CD20 mAb. Acute B cell depletion in either 2- or 4-mo-old mice significantly reduced spleen and lymph node CD4⁺ and CD8⁺ T cell numbers, including naive, activated, and Foxp3⁺CD25⁺CD4⁺ regulatory T cell subsets. The numbers of IFN-γ– and TNF-α–producing T cells were also significantly reduced. Chronic B cell depletion for 6 mo in aged naive mice resulted in a 40–70% reduction in activated CD4⁺ and CD8⁺ T cell numbers and 20–50% reductions in IFN-γ–producing T cells. Therefore, B cells were necessary for maintaining naive CD4⁺ and CD8⁺ T cell homeostasis for subsequent optimal T cell expansion in young and old mice. To determine the significance of this finding, a week of B cell depletion in 4-mo-old mice was followed by acute viral infection with lymphocytic choriomeningitis virus Armstrong. Despite their expansion, activated and cytokine-producing CD4⁺ and CD8⁺ T cell numbers were still significantly reduced 1 wk later. Moreover, viral peptide-specific CD4⁺ and CD8⁺ T cell numbers and effector cell development were significantly reduced in mice lacking B cells, whereas lymphocytic choriomeningitis virus titers were dramatically increased. Thus, T cell function is maintained in B cell–depleted mice, but B cells are required for optimal CD4⁺ and CD8⁺ T cell homeostasis, activation, and effector development in vivo, particularly during responses to acute viral infection.