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Neuromyelitis optica (NMO) is an inflammatory autoimmune disorder of the CNS that predominantly affects the spinal cord and optic nerves. A majority (approximately 75%) of patients with NMO are seropositive for autoantibodies against the astrocyte water ch

Multiple sclerosis is an inflammatory demyelinating disease of the central nervous system (CNS) that is thought to be caused by a combination of genetic and environmental factors. To date, considerable evidence has associated Epstein-Barr virus (EBV) infection with disease development. However, it remains controversial whether EBV infects multiple sclerosis brain and contributes directly to CNS immunopathology. To assess whether EBV infection is a characteristic feature of multiple sclerosis brain, a large cohort of multiple sclerosis specimens containing white matter lesions (nine adult and three paediatric cases) with a heterogeneous B cell infiltrate and a second cohort of multiple sclerosis specimens (12 cases) that included B cell infiltration within the meninges and parenchymal B cell aggregates, were examined for EBV infection using multiple methodologies including in situ hybridization, immunohistochemistry and two independent real-time polymerase chain reaction (PCR) methodologies that detect genomic EBV or the abundant EBV encoded RNA (EBER) 1, respectively. We report that EBV could not be detected in any of the multiple sclerosis specimens containing white matter lesions by any of the methods employed, yet EBV was readily detectable in multiple Epstein-Barr virus-positive control tissues including several CNS lymphomas. Furthermore, EBV was not detected in our second cohort of multiple sclerosis specimens by in situ hybridization. However, our real-time PCR methodologies, which were capable of detecting very few EBV infected cells, detected EBV at low levels in only 2 of the 12 multiple sclerosis meningeal specimens examined. Our finding that CNS EBV infection was rare in multiple sclerosis brain indicates that EBV infection is unlikely to contribute directly to multiple sclerosis brain pathology in the vast majority of cases. [ABSTRACT FROM AUTHOR]

Autoimmunity affects 10% of the population. Within this umbrella, autoantibody-mediated diseases targeting one autoantigen provide a unique opportunity to comprehensively understand the developmental pathway of disease-causing B cells and autoantibodies. While such autoreactivities are believed to be generated during germinal centre reactions, the roles of earlier immune checkpoints in autoantigen-specific B cell tolerance are poorly understood. We address this concept in patients with CASPR2-autoantibody encephalitis and healthy controls. In both groups, comparable and high (∼0.5%) frequencies of unmutated CASPR2-reactive naïve B cells were identified. By contrast, CASPR2-reactive memory B cells were exclusive to patients, and their B cell receptors demonstrated affinity-enhancing somatic mutations with heterogenous binding kinetics. These effector molecules possessed epitope-dependent pathogenic effects in vitro neuronal cultures and in vivo. The unmutated common ancestors of these memory B cells showed a distinctive balance between strong CASPR2 reactivity and very limited binding across the remaining human proteome. Our results are the first to propose mechanisms underlying autoantigen-specific tolerance in humans. We identify permissive central tolerance, defective peripheral tolerance and heterogenous autoantibody binding properties as sequential pathogenic steps which licence CASPR2-directed pathology. By leveraging the basic immunobiology, we rationally direct tolerance-restoring approaches in CASPR2-antibody diseases. This paradigm is applicable across autoimmune conditions.

Autoimmune autonomic ganglionopathy (AAG) is a rare disease wherein autoantibodies target the ganglionic acetylcholine receptor (gAChR). Current diagnosis in the United States depends upon clinical symptoms and positive autoantibody detection using a radioimmunoprecipitation assay (RIA). Here we offer a proof-of-principle study on an alternative method, fluorescence-detection size-exclusion-chromatography (FSEC). We show FSEC can detect autoantibodies against gAChR from patient sera but not healthy controls or samples from other autoimmune diseases. We compare FSEC to RIA and find good correlation. We discuss potential advantages of using FSEC as an alternative or as a first-step diagnostic prior to pursuing existing methodologies., Competing Interests: Declaration of competing interest SV has received compensation as a consultant for argenx, Alterity, Amneal, Catalyst and LabCorp and has received research support from Quest Diagnostics (through a licensing contract). NM has received compensation from Genentech, Inc. as a member of the steering committee for CHIMES and is a co-founder of GenrAb, Inc., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Patients with autoimmune disease have exhausted antigen-specific T cells that remain capable of B cell support.

This post hoc analysis of the randomized, placebo-controlled N-MOmentum study (NCT02200770) of inebilizumab in neuromyelitis optica spectrum disorder (NMOSD) evaluated relationships between circulating B-cell subsets and aquaporin-4 immunoglobulin G (AQP4-lgG) titers and attacks. Among participants receiving placebo, CD20 + and CD27 + B-cell counts were modestly increased from the pre-attack visit to attack; plasmablast/plasma cell gene signature was increased from baseline to the pre-attack visit (p = 0.016) and from baseline to attack (p = 0.009). With inebilizumab treatment, B-cell subset counts decreased and did not increase with attacks. No difference in change of AQP4-IgG titers from baseline to time of attack was observed., (© 2024 The Author(s). Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association.)

Whole-proteome autoantibody profiling reveals an immunological signature that predates the clinical onset of multiple sclerosis.

Given the global surge in autoimmune diseases, it is critical to evaluate emerging therapeutic interventions. Despite numerous new targeted immunomodulatory therapies, comprehensive approaches to apply and evaluate the effects of these treatments longitudinally are lacking. Here, we leveraged advances in programmable-phage immunoprecipitation methodology to explore the modulation, or lack thereof, of autoantibody profiles, proteome-wide, in both health and disease. Using a custom set of over 730,000 human-derived peptides, we demonstrated that each individual, regardless of disease state, possesses a distinct and complex constellation of autoreactive antibodies. For each individual, the set of resulting autoreactivites constituted a unique immunological fingerprint, or "autoreactome," that was remarkably stable over years. Using the autoreactome as a primary output, we evaluated the relative effectiveness of various immunomodulatory therapies in altering autoantibody repertoires. We found that therapies targeting B cell maturation antigen (BCMA) profoundly altered an individual's autoreactome, while anti-CD19 and anti-CD20 therapies had minimal effects. These data both confirm that the autoreactome comprises autoantibodies secreted by plasma cells and strongly suggest that BCMA or other plasma cell-targeting therapies may be highly effective in treating currently refractory autoantibody-mediated diseases.

Neuromyelitis optica spectrum disorder (NMOSD), myelin oligodendrocyte glycoprotein Ab disease, and autoimmune myasthenia gravis (MG) are autoantibody-mediated neurologic conditions where autoantibodies can induce Ab-dependent cellular cytotoxicity (ADCC), a NK cell-mediated effector function. However, whether ADCC is a pathogenic mechanism in patients with these conditions has not been confirmed. We sought to characterize circulatory NK cells using functional assays, phenotyping, and transcriptomics to elucidate their role in pathology. NK cells from NMOSD patients and MG patients with elevated disease burden exhibited reduced ADCC and CD56dimCD16hi NK cells, along with an elevated frequency of CD56dimCD16dim/- NK cells. We determined that ADCC induces a similar phenotypic shift in vitro. Bulk RNA sequencing distinguished the CD56dimCD16dim/- population from the canonical CD56dimCD16hi cytotoxic and CD56hiCD16- immunomodulatory subsets, as well as CD56hiCD16+ NK cells. Multiparameter immunophenotyping of NK cell markers, functional proteins, and receptors similarly showed that the CD56dimCD16dim/- subset exhibits a unique profile while still maintaining expression of characteristic NK markers CD56, CD94, and NKp44. Notably, expression of perforin and granzyme is reduced in comparison with CD56dimCD16hi NK cells. Moreover, they exhibit elevated trogocytosis capability, HLA-DR expression, and many chemokine receptors, including CCR7. In contrast with NMOSD and MG, myelin oligodendrocyte glycoprotein Ab disease NK cells did not exhibit functional, phenotypic, or transcriptomic perturbations. In summary, CD56dimCD16dim/- NK cells are a distinct peripheral blood immune cell population in humans elevated upon prior cytotoxic activity by the CD56dimCD16hi NK cell subset. The elevation of this subset in NMOSD and MG patients suggests prior ADCC activity., (Copyright © 2024 by The American Association of Immunologists, Inc.)

Background:  Limb-threatening lower extremity injuries often require secondary bone grafting after soft tissue reconstruction. We hypothesized that there would be fewer wound complications when performing secondary bone grafting via a remote surgical approach rather than direct flap elevation., Methods:  A retrospective cohort study was performed at a single Level 1 trauma center comparing complications after secondary bone grafting in patients who had undergone previous soft tissue reconstruction after open tibia fractures between 2006 and 2020. Comparing bone grafting via a remote surgical incision versus direct flap elevation, we evaluated wound dehiscence requiring return to the operating room as the primary outcome. Secondary outcomes were deep infection and delayed amputation., Results:  We identified 129 patients (mean age: 40 years, 82% male) with 159 secondary bone grafting procedures. Secondary bone grafting was performed via a remote surgical approach in 54% ( n  = 86) and direct flap elevation in 46% ( n  = 73) of cases. Wound dehiscence requiring return to the operating room occurred in one patient in the flap elevation group (1%) and none of the patients in the remote surgical approach. The odds of deep wound infection (OR, 1.77; p  = 0.31) or amputation (OR, 1.43; p  = 0.73) did not significantly differ between surgical approaches. No significant differences were found in complications between the reconstructive surgeon elevating and re-insetting the flap and the orthopaedic trauma surgeon performing the flap elevation and re-inset., Conclusion:  Direct flap elevation for secondary bone grafting after soft tissue reconstruction for open tibia fractures did not result in more complications than bone grafting via a remote surgical approach. These findings should reassure surgeons to allow other clinical factors to influence the surgical approach for bone grafting., Competing Interests: N.N.O'H. reported receiving stock or stock options from Arbutus Medical, Inc. unrelated to this research. R.V.O'T. reported serving as a paid consultant for Smith & Nephew and Stryker, receiving stock options from Imagen, and receiving royalties from Lincotek, all unrelated to this study. The remaining authors report no conflict of interest., (Thieme. All rights reserved.)

The prevalence and burden of autoimmune and autoantibody mediated disease is increasing worldwide, yet most disease etiologies remain unclear. Despite numerous new targeted immunomodulatory therapies, comprehensive approaches to apply and evaluate the effects of these treatments longitudinally are lacking. Here, we leverage advances in programmable-phage immunoprecipitation (PhIP-Seq) methodology to explore the modulation, or lack thereof, of proteome-wide autoantibody profiles in both health and disease. We demonstrate that each individual, regardless of disease state, possesses a distinct set of autoreactivities constituting a unique immunological fingerprint, or "autoreactome", that is remarkably stable over years. In addition to uncovering important new biology, the autoreactome can be used to better evaluate the relative effectiveness of various therapies in altering autoantibody repertoires. We find that therapies targeting B-Cell Maturation Antigen (BCMA) profoundly alter an individual's autoreactome, while anti-CD19 and CD-20 therapies have minimal effects, strongly suggesting a rationale for BCMA or other plasma cell targeted therapies in autoantibody mediated diseases., Competing Interests: Competing Interest Declaration Potential Conflicts J.D.R. reports being a founder and paid consultant for Delve Bio, Inc., and a paid consultant for the Public Health Company and Allen & Co. M.A.S. receives unrelated research funding from the NIH, the CDC, Cepheid and Merck and unrelated Honoria from UpToDate, Inc. M.R.W. reports being a founder and paid consultant for Delve Bio, Inc. and receives unrelated research grant funding from Roche/Genentech and Novartis, and received speaking honoraria from Genentech, Takeda, WebMD, and Novartis. C.J.T. reports being on the Scientific Advisory Boards for Caribou Biosciences, T-CURX, Myeloid Therapeutics, ArsenalBio, Cargo Therapeutics, Celgene/BMS Cell Therapy; is a member a DSMB member of Kyverna; is on Ad hoc advisory boards/consulting (last 12 months) for Nektar Therapeutics, Legend Biotech, Prescient Therapeutics, Century Therapeutics, IGM Biosciences, Abbvie; has Stock options in Eureka Therapeutics, Caribou Biosciences, Myeloid Therapeutics, ArsenalBio, Cargo Therapeutics; and reports the right to receive payment from Fred Hutch as an inventor on patents related to CAR T-cell therapy. J.A.H. reports research funding from Merck and Takeda and consulting fees from Takeda, Gilead, SentiBio, and Century Therapeutics. J.G. reports research funding from Sobi, Juno Therapeutics (a BMS company), Celgene (a BMS company), Angiocrine Bioscience; is an Ad hoc consultant for Sobi, Legend Biotech, Janssen, Kite Pharma, MorphoSys. D.J.G. has received research funding, has served as an advisor and has received royalties from Juno Therapeutics, a Bristol-Myers Squibb company; has served as an advisor and received research funding from Seattle Genetics; has served as an advisor for GlaxoSmithKline, Celgene, Janssen Biotech, Ensoma and Legend Biotech; and has received research funding from SpringWorks Therapeutics, Sanofi, and Cellectar Biosciences. K.C.O. received unrelated research funding from, and is an equity shareholder of Cabaletta Bio, and receives unrelated research funding from Seismic, argenx, and Viela Bio/Horizon (now Amgen). R.J.N. reports unrelated research support from the National Institutes of Health, Genentech, Inc., Alexion Pharmaceuticals, Inc., argenx, Annexon Biosciences, Inc., Ra Pharmaceuticals, Inc. (now UCB S.A.), the Myasthenia Gravis Foundation of America, Inc., Momenta Pharmaceuticals, Inc. (now Janssen), Immunovant, Inc., Grifols, S.A., and Viela Bio, Inc. (now Horizon Therapeutics plc). R.J.N. has also served as a consultant/advisor unrelated to research in this manuscript for Alexion Pharmaceuticals, Inc., argenx, Cabaletta Bio, Inc., Cour Pharmaceuticals, CSL Behring, Grifols, S.A., Ra Pharmaceuticals, Inc. (now UCB S.A.), Immunovant, Inc., Momenta Pharmaceuticals, Inc. (now Janssen), and Viela Bio, Inc. (now Horizon Therapeutics plc). J.L.D., C.M.B. and M.R.W. receive licensing fees from CDI Labs.

Objective: Myasthenia gravis (MG) is an autoantibody-mediated neuromuscular junction disorder involving the acetylcholine receptors on the motor endplate. The safety and response to high-dose chemotherapy (HDIT) and autologous hematopoietic cell transplantation (HCT) were assessed in a patient with severe refractory MG., Methods: As part of a pilot study of HDIT/HCT for patients with treatment-resistant autoimmune neurological disorders, a patient with severe refractory MG underwent treatment. After mobilization of hematopoietic stem cells with rituximab, prednisone, and G-CSF, the patient had HDIT consisting of carmustine, etoposide, cytarabine, melphalan, and rabbit antithymocyte globulin, followed by autologous HCT. The effect of treatment on the autoantibody to the acetylcholine receptor (AChR) was assessed., Results: The patient had been diagnosed with AChR antibody-positive MG 14 years before HDIT/HCT and had failed thymectomy, therapeutic plasma exchange, and multiple immunomodulatory agents. The Myasthenia Gravis Foundation of America (MGFA) clinical classification was IVb before HDIT/HCT. She tolerated HDIT/HCT well and started to improve clinically within days of treatment. At both 1 and 2 years after HDIT/HCT, patients remained symptom-free. After HDIT/HCT, AChR-binding autoantibodies persisted, and the relative frequency of immune cell subtypes shifted., Interpretation: HDIT/HCT induced a complete response of disease activity in a patient with severe refractory MG. This response may suggest that a cell-mediated etiology may be a significant contributing factor in refractory MG cases. A phase 2 clinical trial is warranted to establish if HDIT/HCT can be an effective therapy for severe refractory MG and to gain a further understanding of disease pathogenesis., (© 2023 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association.)

Curated expression of proteolipid protein 1 (PLP1) is essential for multiple sclerosis-derived autoantibody recognition.

Muscle-specific tyrosine kinase myasthenia gravis (MuSK MG) is an autoimmune disease that causes life-threatening muscle weakness due to anti-MuSK autoantibodies that disrupt neuromuscular junction signaling. To avoid chronic immunosuppression from current therapies, we engineered T cells to express a MuSK chimeric autoantibody receptor with CD137-CD3ζ signaling domains (MuSK-CAART) for precision targeting of B cells expressing anti-MuSK autoantibodies. MuSK-CAART demonstrated similar efficacy as anti-CD19 chimeric antigen receptor T cells for depletion of anti-MuSK B cells and retained cytolytic activity in the presence of soluble anti-MuSK antibodies. In an experimental autoimmune MG mouse model, MuSK-CAART reduced anti-MuSK IgG without decreasing B cells or total IgG levels, reflecting MuSK-specific B cell depletion. Specific off-target interactions of MuSK-CAART were not identified in vivo, in primary human cell screens or by high-throughput human membrane proteome array. These data contributed to an investigational new drug application and phase 1 clinical study design for MuSK-CAART for the treatment of MuSK autoantibody-positive MG., (© 2023. The Author(s).)

Tissue-resident memory T cells accumulate in mucosal sites during infancy and then mature through childhood.

Serum autoantibodies targeting the nicotinic acetylcholine receptor (AChR) in patients with autoimmune myasthenia gravis (MG) can mediate pathology via three distinct molecular mechanisms: complement activation, receptor blockade, and antigenic modulation. However, it is unclear whether multi-pathogenicity is mediated by individual or multiple autoantibody clones. Using an unbiased B cell culture screening approach, we generated a library of 11 human-derived AChR-specific recombinant monoclonal autoantibodies (mAb) and assessed their binding properties and pathogenic profiles using specialized cell-based assays. Five mAbs activated complement, three blocked α-bungarotoxin binding to the receptor, and seven induced antigenic modulation. Furthermore, two clonally related mAbs derived from one patient were each highly efficient at more than one of these mechanisms, demonstrating that pathogenic mechanisms are not mutually exclusive at the monoclonal level. Using novel Jurkat cell lines that individually express each monomeric AChR subunit (α 2 βδε), these two mAbs with multi-pathogenic capacity were determined to exclusively bind the α-subunit of AChR, demonstrating an association between mAb specificity and pathogenic capacity. These findings provide new insight into the immunopathology of MG, demonstrating that single autoreactive clones can efficiently mediate multiple modes of pathology. Current therapeutic approaches targeting only one autoantibody-mediated pathogenic mechanism may be evaded by autoantibodies with multifaceted capacity., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Myelin oligodendrocyte glycoprotein (MOG) antibody-associated disease (MOGAD) is an inflammatory demyelinating CNS condition characterized by the presence of MOG autoantibodies. We sought to investigate whether human MOG autoantibodies are capable of mediating damage to MOG-expressing cells through multiple mechanisms. We developed high-throughput assays to measure complement activity (CA), complement-dependent cytotoxicity (CDC), antibody-dependent cellular phagocytosis (ADCP), and antibody-dependent cellular cytotoxicity (ADCC) of live MOG-expressing cells. MOGAD patient sera effectively mediate all of these effector functions. Our collective analyses reveal that (a) cytotoxicity is not incumbent on MOG autoantibody quantity alone; (b) engagement of effector functions by MOGAD patient serum is bimodal, with some sera exhibiting cytotoxic capacity while others did not; (c) the magnitude of CDC and ADCP is elevated closer to relapse, while MOG-IgG binding is not; and (d) all IgG subclasses can damage MOG-expressing cells. Histopathology from a representative MOGAD case revealed congruence between lesion histology and serum CDC and ADCP, and we identified NK cells, mediators of ADCC, in the cerebrospinal fluid of relapsing patients with MOGAD. Thus, MOGAD-derived autoantibodies are cytotoxic to MOG-expressing cells through multiple mechanisms, and assays quantifying CDC and ADCP may prove to be effective tools for predicting risk of future relapses.

To compare the immunopathology of immune checkpoint inhibitor-induced myasthenia gravis (ICI-MG) and idiopathic MG, we profiled the respective AChR autoantibody pathogenic properties. Of three ICI-MG patients with AChR autoantibodies, only one showed complement activation and modulation/blocking potency, resembling idiopathic MG. In contrast, AChR autoantibody-mediated effector functions were not detected in the other two patients, questioning the role of their AChR autoantibodies as key mediators of pathology. The contrasting properties of AChR autoantibodies in these cases challenge the accuracy of serological testing in establishing definite ICI-MG diagnoses and underscore the importance of a thorough clinical assessment when evaluating ICI-related adverse events., (© 2023 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association.)

Inclusion body myositis (IBM) is an autoimmune and degenerative disorder of skeletal muscle. The B cell infiltrates in IBM muscle tissue are predominantly fully differentiated Ab-secreting plasma cells, with scarce naive or memory B cells. The role of this infiltrate in the disease pathology is not well understood. To better define the humoral response in IBM, we used adaptive immune receptor repertoire sequencing, of human-derived specimens, to generate large BCR repertoire libraries from IBM muscle biopsies and compared them to those generated from dermatomyositis, polymyositis, and circulating CD27+ memory B cells, derived from healthy controls and Ab-secreting cells collected following vaccination. The repertoire properties of the IBM infiltrate included the following: clones that equaled or exceeded the highly clonal vaccine-associated Ab-secreting cell repertoire in size; reduced somatic mutation selection pressure in the CDRs and framework regions; and usage of class-switched IgG and IgA isotypes, with a minor population of IgM-expressing cells. The IBM IgM-expressing population revealed unique features, including an elevated somatic mutation frequency and distinct CDR3 physicochemical properties. These findings demonstrate that some of IBM muscle BCR repertoire characteristics are distinct from dermatomyositis and polymyositis and circulating Ag-experienced subsets, suggesting that it may form through selection by disease-specific Ags., (Copyright © 2023 The Authors.)

An approach for identifying antibodies derived from distinct B cell populations demonstrates how secondary immunization responses are dominated by mature B cells generated during primary responses.

Background: Neuromyelitis optica spectrum disorder (NMOSD) is a rare, autoimmune disease of the central nervous system that produces acute, unpredictable relapses causing cumulative neurological disability. Satralizumab, a humanized, monoclonal recycling antibody that targets the interleukin-6 receptor, reduced NMOSD relapse risk vs. placebo in two Phase 3 trials: SAkuraSky (satralizumab ± immunosuppressive therapy; NCT02028884) and SAkuraStar (satralizumab monotherapy; NCT02073279). Satralizumab is approved to treat aquaporin-4 IgG-seropositive (AQP4-IgG+) NMOSD. SAkuraBONSAI (NCT05269667) will explore fluid and imaging biomarkers to better understand the mechanism of action of satralizumab and the neuronal and immunological changes following treatment in AQP4-IgG+ NMOSD., Objectives: SAkuraBONSAI will evaluate clinical disease activity measures, patient-reported outcomes (PROs), pharmacokinetics, and safety of satralizumab in AQP4-IgG+ NMOSD. Correlations between imaging markers (magnetic resonance imaging [MRI] and optical coherence tomography [OCT]) and blood and cerebrospinal fluid (CSF) biomarkers will be investigated., Study Design: SAkuraBONSAI is a prospective, open-label, multicenter, international, Phase 4 study that will enroll approximately 100 adults (18-74 years) with AQP4-IgG+ NMOSD. This study includes two patient cohorts: newly diagnosed, treatment-naïve patients (Cohort 1; n = 60); and inadequate responders to recent (<6 months) rituximab infusion (Cohort 2; n = 40). Satralizumab monotherapy (120 mg) will be administered subcutaneously at Weeks 0, 2, 4, and Q4W thereafter for a total of 92 weeks., Endpoints: Disease activity related to relapses (proportion relapse-free, annualized relapse rate, time to relapse, and relapse severity), disability progression (Expanded Disability Status Scale), cognition (Symbol Digit Modalities Test), and ophthalmological changes (visual acuity; National Eye Institute Visual Function Questionnaire-25) will all be assessed. Peri-papillary retinal nerve fiber layer and ganglion cell complex thickness will be monitored using advanced OCT (retinal nerve fiber layer and ganglion cell plus inner plexiform layer thickness). Lesion activity and atrophy will be monitored by MRI. Pharmacokinetics, PROs, and blood and CSF mechanistic biomarkers will be assessed regularly. Safety outcomes include the incidence and severity of adverse events., Conclusions: SAkuraBONSAI will incorporate comprehensive imaging, fluid biomarker, and clinical assessments in patients with AQP4-IgG+ NMOSD. SAkuraBONSAI will provide new insights into the mechanism of action of satralizumab in NMOSD, while offering the opportunity to identify clinically relevant neurological, immunological, and imaging markers., Competing Interests: JLB reports payment for study design/consultation from MedImmune/Viela Bio; personal fees from AbbVie, Antigenomycs, Alexion, Chugai, Clene Nanomedicine, Genentech, Genzyme, Mitsubishi-Tanabe, Reistone Bio, Beigene, TG Therapeutics, Novartis, and Roche; grants from Novartis, Mallinckrodt, and the National Institutes of Health; and has a patent for Aquaporumab issued. KF serves as an advisor or on scientific advisory boards for Biogen, Mitsubishi Tanabe, Novartis, Chugai/Roche, Alexion, Viela Bio/Horizon Therapeutics, UCB, Merck Biopharma, Japan Tobacco, and AbbVie; has received funding for travel and speaker honoraria from Biogen, Eisai, Mitsubishi Tanabe, Novartis, Chugai, Roche, Alexion, Viela Bio, Teijin, Asahi Kasei Medical, Merck, and Takeda; and has received the Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan and the Grants-in-Aid for Scientific Research from the Ministry of Health, Welfare and Labor of Japan. HJK has received a grant from the National Research Foundation of Korea and research support from Aprilbio and Eisai; received consultancy/speaker fees from Alexion, Aprilbio, Altos Biologics, Biogen, Celltrion, Daewoong, Eisai, GC Pharma, HanAll BioPharma, Handok, Horizon Therapeutics (formerly Viela Bio), MDimune, Mitsubishi Tanabe Pharma, Merck Serono, Novartis, Roche, Sanofi Genzyme, Teva-Handok, and UCB; is a co-editor for the Multiple Sclerosis Journal and an associate editor for the Journal of Clinical Neurology. RM serves on scientific advisory boards for Viela Bio/Horizon Therapeutics, Roche, Alexion, UCB; and has received funding for travel and fees from Alexion, Biogen, Merck, Novartis, Roche, and Viela Bio/Horizon Therapeutics. KCO'C has received research support from Alexion, now part of AstraZeneca, and Viela Bio, now part of Horizon Therapeutics, Cabaletta Bio, and Argenx. He is a consultant and equity shareholder of Cabaletta Bio. During the last 2 years, he has served as consultant/advisor for Alexion Pharmaceuticals, now part of AstraZeneca, and for Roche. RCS has been a consultant to Roche Pharmaceuticals and directs the William H. Annesley EyeBrain Center, including the OCT reading center at Thomas Jefferson University partnered with Wills Eye Hospital. AT is the MS Society of Canada Research Chair at UBC, with support from the MSMRI Research Group. He has research funding from Hilton Foundation, Roche, and Biogen; and receives honoraria from Roche, Sanofi Genzyme, and Biogen. HW receives honoraria for acting as a member of scientific advisory boards for AbbVie, Alexion, Argenx, Bristol Myers Squibb/Celgene, Janssen, Merck, Novartis, and Sandoz, as well as speaker honoraria and travel support from Alexion, Biogen, Bristol Myers Squibb, Genzyme, Merck, Neurodiem, Novartis, Roche, TEVA, and WebMD Global. He is acting as a paid consultant for AbbVie, Actelion, Argenx, Beckton Dickinson, Biogen, Bristol Myers Squibb, EMD Serono, Fondazione Cariplo, Gossamer Bio, Idorsia, Immunic, Immunovant, Janssen, Lundbeck, Merck, NexGen, Novartis, PSI CRO, Roche, Sanofi, Swiss Multiple Sclerosis Society, UCB, and Worldwide Clinical Trials. His research was funded by the German Federal Ministry for Education and Research (BMBF), Deutsche Forschungsgesellschaft (DFG), Deutsche Myasthenie Gesellschaft e.V., Alexion, Amicus, Therapeutics Inc., Argenx, Biogen, CSL Behring, F. Hoffmann-La Roche, Genzyme, Merck KgaA, Novartis, Roche Pharma, and UCB Biopharma. JW, RB, and TK are employees of F. Hoffmann-La Roche Ltd. SSZ has served, or serves, as a consultant and received honoraria from Alexion, Biogen, EMD-Serono, Horizon, Novartis, Roche/Genentech, Sanofi-Genzyme, and Teva Pharmaceuticals, Inc., and has served on Data Safety Monitoring Boards for Lilly, BioMS, Teva, and Opexa Therapeutics. VGA is an employee of Genentech, Inc. ANL is an employee of Genentech, Inc., and stockholder of Genentech, Inc., and F. Hoffmann-La Roche Ltd. SL-C is an employee of Roche Products Ltd. SJP has received personal compensation for serving as a consultant for Genentech/Roche, Sage Therapeutics, and Astellas. He has received personal compensation for serving on scientific advisory boards or data safety monitoring boards for F. Hoffmann-La Roche AG, Genentech, and UCB. His institution has received compensation for serving as a consultant for Astellas, Alexion, and Viela Bio/MedImmune. All compensation is paid to Mayo Clinic. He has received research support from Alexion, Viela Bio/MedImmune, and Roche/Genentech. He has two patents issued: Patent# 8,889,102 (Application#12-678350, Neuromyelitis Optica Autoantibodies as a Marker for Neoplasia); Patent# 9,891,219B2 (Application#12-573942, Methods for Treating Neuromyelitis Optica [NMO] by Administration of Eculizumab to an individual that is Aquaporin-4 [AQP4]-IgG Autoantibody positive). The authors declare that this study is funded by F. Hoffmann-La Roche Ltd. F. Hoffmann-La Roche Ltd. contributed to the study design and writing of this article. All authors, including those employed by Roche, had final responsibility for the decision to submit for publication and will be involved in future collection, analysis, and interpretation of data., (Copyright © 2023 Bennett, Fujihara, Kim, Marignier, O'Connor, Sergott, Traboulsee, Wiendl, Wuerfel, Zamvil, Anania, Buffels, Künzel, Lekkerkerker, Lennon-Chrimes and Pittock.)

Myasthenia gravis (MG) is an autoantibody-mediated autoimmune disorder of the neuromuscular junction. A small subset of patients (<10%) with MG, have autoantibodies targeting muscle-specific tyrosine kinase (MuSK). MuSK MG patients respond well to CD20-mediated B cell depletion therapy (BCDT); most achieve complete stable remission. However, relapse often occurs. To further understand the immunomechanisms underlying relapse, we studied autoantibody-producing B cells over the course of BCDT. We developed a fluorescently labeled antigen to enrich for MuSK-specific B cells, which was validated with a novel Nalm6 cell line engineered to express a human MuSK-specific B cell receptor. B cells (≅ 2.6 million) from 12 different samples collected from nine MuSK MG patients were screened for MuSK specificity. We successfully isolated two MuSK-specific IgG4 subclass-expressing plasmablasts from two of these patients, who were experiencing a relapse after a BCDT-induced remission. Human recombinant MuSK mAbs were then generated to validate binding specificity and characterize their molecular properties. Both mAbs were strong MuSK binders, they recognized the Ig1-like domain of MuSK, and showed pathogenic capacity when tested in an acetylcholine receptor (AChR) clustering assay. The presence of persistent clonal relatives of these MuSK-specific B cell clones was investigated through B cell receptor repertoire tracing of 63,977 unique clones derived from longitudinal samples collected from these two patients. Clonal variants were detected at multiple timepoints spanning more than five years and reemerged after BCDT-mediated remission, predating disease relapse by several months. These findings demonstrate that a reservoir of rare pathogenic MuSK autoantibody-expressing B cell clones survive BCDT and reemerge into circulation prior to manifestation of clinical relapse. Overall, this study provides both a mechanistic understanding of MuSK MG relapse and a valuable candidate biomarker for relapse prediction., (© 2022. The Author(s).)

Patients with multiple sclerosis display broad EBV-specific TCR repertoires driven by an enduring anti-EBV immune response.

Purpose of Review: This review summarizes recent insights into the immunopathogenesis of autoimmune myasthenia gravis (MG). Mechanistic understanding is presented according to MG disease subtypes and by leveraging the knowledge gained through the use of immunomodulating biological therapeutics., Recent Findings: The past two years of research on MG have led to a more accurate definition of the mechanisms through which muscle-specific tyrosine kinase (MuSK) autoantibodies induce pathology. Novel insights have also emerged from the collection of stronger evidence on the pathogenic capacity of low-density lipoprotein receptor-related protein 4 autoantibodies. Clinical observations have revealed a new MG phenotype triggered by cancer immunotherapy, but the underlying immunobiology remains undetermined. From a therapeutic perspective, MG patients can now benefit from a wider spectrum of treatment options. Such therapies have uncovered profound differences in clinical responses between and within the acetylcholine receptor and MuSK MG subtypes. Diverse mechanisms of immunopathology between the two subtypes, as well as qualitative nuances in the autoantibody repertoire of each patient, likely underpin the variability in therapeutic outcomes. Although predictive biomarkers of clinical response are lacking, these observations have ignited the development of assays that might assist clinicians in the choice of specific therapeutic strategies., Summary: Recent advances in the understanding of autoantibody functionalities are bringing neuroimmunologists closer to a more detailed appreciation of the mechanisms that govern MG pathology. Future investigations on the immunological heterogeneity among MG patients will be key to developing effective, individually tailored therapies., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

Ischemic stroke generates an immune response that contributes to neuronal loss as well as tissue repair. This is a complex process involving a range of cell types and effector molecules and impacts tissues outside of the CNS. Recent reviews address specific aspects of this response, but several years have passed and important advances have been made since a high-level review has summarized the overall state of the field. The present review examines the initiation of the inflammatory response after ischemic stroke, the complex impacts of leukocytes on patient outcome, and the potential of basic science discoveries to impact the development of therapeutics. The information summarized here is derived from broad PubMed searches and aims to reflect recent research advances in an unbiased manner. We highlight valuable recent discoveries and identify gaps in knowledge that have the potential to advance our understanding of this disease and therapies to improve patient outcomes., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Objective: To quantify patient preferences towards time to return to driving relative to compromised reaction time and potential complication risks., Design: Cross-sectional discrete choice experiment., Setting: Academic trauma center., Patients: Ninety-six adult patients with an operative lower extremity fracture from December 2019 through December 2020., Intervention: None., Main Outcome Measurement: Patient completed a discrete choice experiment survey consisting of 12 hypothetical return to driving scenarios with varied attributes: time to return to driving (range: 1 to 6 months), risk of implant failure (range: 1% to 12%), pain upon driving return (range: none to severe), and driving safety measured by braking distance (range: 0 to 40 feet at 60 mph). The relative importance of each attribute is reported on a scale of 0% to 100%., Results: Patients most valued a reduced pain level when resuming driving (62%), followed by the risk of implant failure (17%), time to return to driving (13%), and braking safety (8%). Patients were indifferent to returning to driving at 1 month (median utility: 28, interquartile range [IQR] -31 to 80) or 2 months (median utility: 59, IQR: 41 to 91) postinjury., Conclusion: Patients with lower extremity injuries demonstrated a willingness to forego earlier return to driving if it might mean a decrease in their pain level. Patients are least concerned about their driving safety, instead placing higher value on their own pain level and chance of implant failure. The findings of this study are the first to rigorously quantify patient preferences toward a return to driving and heterogeneity in patient preferences., Level of Evidence: V., Competing Interests: The remaining authors have no conflicts of interest to disclose., (Copyright © 2022 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of the Orthopaedic Trauma Association.)

Trial eligibility in myasthenia gravis (MG) remains largely dependent on a positive autoantibody serostatus. This significantly hinders seronegative MG (SNMG) patients from receiving potentially beneficial new treatments. In a subset of SNMG patients, acetylcholine receptor (AChR) autoantibodies are detectable by a clustered AChR cell-based assay (CBA). Of 99 SNMG patients from two academic U.S. centers, 18 (18.2%) tested positive by this assay. Autoantibody positivity was further validated in 17/18 patients. In a complementary experiment, circulating AChR-specific B cells were identified in a CBA-positive SNMG patient. These findings corroborate the clinical need for clustered AChR CBA testing when evaluating SNMG patients., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Background: Sporadic inclusion body myositis (IBM) is a debilitating disease which leads to impaired ambulation and loss of hand function. Yale IBM Registry (IBMR) was launched in November 2016 to address the knowledge gap in IBM natural history data. The registry interface provides an IBM personalized index calculator (IBM-PIC) based on the IBM-functional rating scale (IBM-FRS). While the calculator is based on the IBM-FRS, it has not been directly compared to the IBM-FRS score. Therefore, in this study, we compared the patient-reported IBM-PIC score from this calculator with the physician-obtained IBM-FRS score., Method: IBM-FRS was administered over the phone within two weeks of their most recent IBM-PIC entry in the IBMR to 35 participants. To compare the agreement between IBM-FRS and IBM-PIC scores, Interrater Correlation Coefficient (ICC) analysis was performed. For individual questions, Fleiss Kappa statistics was used., Results: Thirty-five active IBM-PIC users participated. Eighty percent of the participants were men, and 91% were White Caucasians. The reported IBM-FRS score of this group was 23.5 ± 7.4 (range 1-38). The Interrater Correlation Coefficient (ICC) between the physician-administered IBM-FRS score and the IBM-PIC was 0.98 (0.96-0.99). There was moderate to substantial agreement on all the questions on IBM-FRS except for handwriting and fine motor skills., Discussion: IBM-PIC is a reliable indicator of the IBM-FRS score obtained by the physician. It is anticipated that this online platform will be a valuable tool for assessing IBM severity and monitoring disease progression remotely both in clinical practice and research studies., (Copyright © 2022 Elsevier B.V. All rights reserved.)

T cells can acquire CD20 from B cells via trogocytosis, then contribute to autoimmune neurological disease while becoming targets for therapeutically effective B cell depletion.

Background and Objectives: Autoantibodies targeting the acetylcholine receptor (AChR), found in patients with myasthenia gravis (MG), mediate pathology through 3 mechanisms: complement-directed tissue damage, blocking of the acetylcholine binding site, and internalization of the AChR. Clinical assays, used to diagnose and monitor patients, measure only autoantibody binding. Consequently, they are limited in providing association with disease burden, understanding of mechanistic heterogeneity, and monitoring therapeutic response. The objective of this study was to develop a cell-based assay that measures AChR autoantibody-mediated complement membrane attack complex (MAC) formation., Methods: An HEK293T cell line-modified using CRISPR/Cas9 genome editing to disrupt expression of the complement regulator genes (CD46, CD55, and CD59)-was used to measure AChR autoantibody-mediated MAC formation through flow cytometry., Results: Serum samples (n = 155) from 96 clinically confirmed AChR MG patients, representing a wide range of disease burden and autoantibody titer, were tested along with 32 healthy donor (HD) samples. AChR autoantibodies were detected in 139 of the 155 (89.7%) MG samples through a cell-based assay. Of the 139 AChR-positive samples, autoantibody-mediated MAC formation was detected in 83 (59.7%), whereas MAC formation was undetectable in the HD group or AChR-positive samples with low autoantibody levels. MAC formation was positively associated with autoantibody binding in most patient samples; ratios (mean fluorescence intensity) of MAC formation to AChR autoantibody binding ranged between 0.27 and 48, with a median of 0.79 and an interquartile range of 0.43 (0.58-1.1). However, the distribution of ratios was asymmetric and included extreme values; 16 samples were beyond the 10-90 percentile, with high MAC to low AChR autoantibody binding ratio or the reverse. Correlation between MAC formation and clinical disease scores suggested a modest positive association (rho = 0.34, p = 0.0023), which included a subset of outliers that did not follow this pattern. MAC formation did not associate with exposure to immunotherapy, thymectomy, or MG subtypes defined by age-of-onset., Discussion: A novel assay for evaluating AChR autoantibody-mediated complement activity was developed. A subset of patients that lacks association between MAC formation and autoantibody binding or disease burden was identified. The assay may provide a better understanding of the heterogeneous autoantibody molecular pathology and identify patients expected to benefit from complement inhibitor therapy., (Copyright © 2022 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.)

Acetylcholine receptor (AChR) autoantibodies, found in patients with autoimmune myasthenia gravis (MG), can directly contribute to disease pathology through activation of the classical complement pathway. Activation of the complement pathway in autoimmune diseases can lead to a secondary complement deficiency resulting in reduced complement activity, due to consumption, during episodes of disease activity. It is not clear whether complement activity in MG patients associates with measurements of disease activity or the titer of circulating pathogenic AChR autoantibodies. To explore such associations, as a means to identify a candidate biomarker, we measured complement activity in AChR MG samples (N = 51) using a CH50 hemolysis assay, then tested associations between these values and both clinical status and AChR autoantibody titer. The majority of the study subjects (88.2%) had complement activity within the range defined by healthy controls, while six patients (11.8%) showed reduced activity. No significant association between complement activity and disease status or AChR autoantibody titer was observed., Competing Interests: The authors have read the journal’s policy and have the following competing interests: KCO has received research support from Ra Pharma, now a part of UCB Pharma. DDV and MDH are paid employees of UCB Pharma. RJN has received research support from UCB Pharma and served as consultant/advisor for UCB Pharma. The study did not receive funding via any of the authors’ salaries. These competing interests do not alter our adherence to PLOS ONE policies on sharing data and materials. There are no patents, products in development or marketed products associated with this research to declare.

Objective: To determine whether rituximab is safe and potentially beneficial, warranting further investigation in an efficacy trial for acetylcholine receptor antibody-positive generalized MG (AChR-Ab+ gMG)., Methods: The B-Cell Targeted Treatment in MG (BeatMG) study was a randomized, double-blind, placebo-controlled, multicenter phase-2 trial that utilized a futility design. Individuals 21-90 years of age, with AChR-Ab+ gMG (MG Foundation of America Class II-IV) and receiving prednisone ≥15 mg/day were eligible. The primary outcome was a measure of steroid-sparing effect, defined as the proportion achieving ≥75% reduction in mean daily prednisone dose in the 4-weeks prior to week 52 and with clinical improvement or no significant worsening as compared to the 4-week period prior to randomization. The co-primary outcome was safety. Secondary outcomes included MG-specific clinical assessments. Fifty-two individuals were randomized (1:1) to either a two-cycle rituximab/placebo regimen, with follow-up through 52-weeks., Results: Of the 52 participants included, mean (±SD) age at enrollment was 55.1 (±17.1) years; 23 (44.2%) were female, and 31 (59.6%) were MGFA Class II. The mean (±SD) baseline prednisone dose was 22.1 (±9.7) mg/day. The primary steroid-sparing outcome was achieved in 60% of those on rituximab vs. 56% on placebo. The study reached its futility endpoint (p=0.03) suggesting that the pre-defined clinically meaningful improvement of 30% due to rituximab over placebo was unlikely to be achieved in a subsequent, larger trial. No safety issues identified., Conclusions: While rituximab was safe and well-tolerated, these results suggest that there is a low probability of observing the defined clinically meaningful steroid-sparing effect over a 12-month period in a phase-3 trial of mild-moderately symptomatic AChR-Ab+ gMG., Classification of Evidence: This study provides Class I evidence that for mild-to-moderate AChR-Ab+ gMG, compared with placebo, rituximab is safe but unlikely to reduce steroid use by an absolute difference of at least 30% at 1 year., Trial Registration: ClinicalTrials.gov Identifier: NCT02110706., (© 2021 American Academy of Neurology.)

Epileptic seizures can be mediated by patient autoantibodies targeting ion channels expressed in the brain.

Background and Objectives: There are limited data on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine reactogenicity in persons with multiple sclerosis (PwMS) and how reactogenicity is affected by disease-modifying therapies (DMTs). The objective of this retrospective cross-sectional study was to generate real-world multiple sclerosis-specific vaccine safety information, particularly in the context of specific DMTs, and provide information to mitigate specific concerns in vaccine hesitant PwMS., Methods: Between 3/2021 and 6/2021, participants in iConquerMS, an online people-powered research network, reported SARS-CoV-2 vaccines, experiences of local (itch, pain, redness, swelling, or warmth at injection site) and systemic (fever, chills, fatigue, headache, joint pain, malaise, muscle ache, nausea, allergic, and other) reactions within 24 hours (none, mild, moderate, and severe), DMT use, and other attributes. Multivariable models characterized associations between clinical factors and reactogenicity., Results: In 719 PwMS, 64% reported experiencing a reaction after their first vaccination shot, and 17% reported a severe reaction. The most common reactions were pain at injection site (54%), fatigue (34%), headache (28%), and malaise (21%). Younger age, being female, prior SARS-CoV-2 infection, and receiving the ChAdOx1 nCoV-19 (Oxford-AstraZeneca) vs BNT162b2 (Pfizer-BioNTech) vaccine were associated with experiencing a reaction after the first vaccine dose. Similar relationships were observed for a severe reaction, including higher odds of reactions among PwMS with more physical impairment and lower odds of reactions for PwMS on an alpha4-integrin blocker or sphingosine-1-phosphate receptor modulator. In 442 PwMS who received their second vaccination shot, 74% reported experiencing a reaction, whereas 22% reported a severe reaction. Reaction profiles after the second shot were similar to those reported after the first shot. Younger PwMS and those who received the mRNA-1273 (Moderna) vs BNT162b2 vaccine reported higher reactogenicity after the second shot, whereas those on a sphingosine-1-phosphate receptor modulator or fumarate were significantly less likely to report a reaction., Discussion: SARS-CoV-2 vaccine reactogenicity profiles and the associated factors in this convenience sample of PwMS appear similar to those reported in the general population. PwMS on specific DMTs were less likely to report vaccine reactions. Overall, the short-term vaccine reactions experienced in the study population were mostly self-limiting, including pain at the injection site, fatigue, headache, and fever., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.)

Elevated N-linked glycosylation of IgG V regions (IgG-V N-Glyc ) is an emerging molecular phenotype associated with autoimmune disorders. To test the broader specificity of elevated IgG-V N-Glyc , we studied patients with distinct subtypes of myasthenia gravis (MG), a B cell-mediated autoimmune disease. Our experimental design focused on examining the B cell repertoire and total IgG. It specifically included adaptive immune receptor repertoire sequencing to quantify and characterize N-linked glycosylation sites in the circulating BCR repertoire, proteomics to examine glycosylation patterns of the total circulating IgG, and an exploration of human-derived recombinant autoantibodies, which were studied with mass spectrometry and Ag binding assays to respectively confirm occupation of glycosylation sites and determine whether they alter binding. We found that the frequency of IgG-V N-Glyc motifs was increased in the total BCR repertoire of patients with MG when compared with healthy donors. The elevated frequency was attributed to both biased V gene segment usage and somatic hypermutation. IgG-V N-Glyc could be observed in the total circulating IgG in a subset of patients with MG. Autoantigen binding, by four patient-derived MG autoantigen-specific mAbs with experimentally confirmed presence of IgG-V N-Glyc , was not altered by the glycosylation. Our findings extend prior work on patterns of Ig V region N-linked glycosylation in autoimmunity to MG subtypes., (Copyright © 2021 by The American Association of Immunologists, Inc.)

Elevated frequency of afucosylated IgG1 antibodies during dengue virus infection is associated with prior infection and predicts severe disease., (Copyright © 2021, American Association for the Advancement of Science.)

Background: The current global pandemic has created unprecedented challenges in the blood supply network. Given the recent shortages, there must be a civilian plan for massively bleeding patients when there are no blood products on the shelf. Recognizing that the time to death in bleeding patients is less than 2 h, timely resupply from unaffected locations is not possible. One solution is to transfuse emergency untested whole blood (EUWB), similar to the extensive military experience fine-tuned over the last 19 years. While this concept is anathema in current civilian transfusion practice, it seems prudent to have a vetted plan in place., Methods and Materials: During the early stages of the 2020 global pandemic, a multidisciplinary and international group of clinicians with broad experience in transfusion medicine communicated routinely. The result is a planning document that provides both background information and a high-level guide on how to emergently deliver EUWB for patients who would otherwise die of hemorrhage., Results and Conclusions: Similar plans have been utilized in remote locations, both on the battlefield and in civilian practice. The proposed recommendations are designed to provide high-level guidance for experienced blood bankers, transfusion experts, clinicians, and health authorities. Like with all emergency preparedness, it is always better to have a well-thought-out and trained plan in place, rather than trying to develop a hasty plan in the midst of a disaster. We need to prevent the potential for empty shelves and bleeding patients dying for lack of blood., (© 2021 AABB.)

Charting the gene-expression landscape of glioma-infiltrating T cells demonstrates that these T cells can express NK cell receptors, which inhibit the killing of glioma cells., (Copyright © , American Association for the Advancement of Science.)

Preclinical animal studies are essential to the development of safe and effective stem cell therapies. Bioluminescence imaging (BLI) is a powerful tool in animal studies that enables the real-time longitudinal monitoring of stem cells in vivo to elucidate their regenerative properties. This review describes the application of BLI in preclinical stem cell research to address critical challenges in producing successful stem cell therapeutics. These challenges include stem cell survival, proliferation, homing, stress response, and differentiation. The applications presented here utilize bioluminescence to investigate a variety of stem and progenitor cells in several different in vivo models of disease and implantation. An overview of luciferase reporters is provided, along with the advantages and disadvantages of BLI. Additionally, BLI is compared to other preclinical imaging modalities and potential future applications of this technology are discussed in emerging areas of stem cell research., (© 2020 The Authors. Biotechnology Journal published by Wiley-VCH GmbH.)

CD4+ follicular regulatory T (Tfr) cells control B cell responses through the modulation of follicular helper T (Tfh) cells and germinal center development while suppressing autoreactivity; however, their role in the regulation of productive germinal center B cell responses and humoral memory is incompletely defined. We show that Tfr cells promote antigen-specific germinal center B cell responses upon influenza virus infection. Following viral challenge, we found that Tfr cells are necessary for robust generation of virus-specific, long-lived plasma cells, antibody production against both hemagglutinin (HA) and neuraminidase (NA), the two major influenza virus glycoproteins, and appropriate regulation of the BCR repertoire. To further investigate the functional relevance of Tfr cells during viral challenge, we used a sequential immunization model with repeated exposure of antigenically partially conserved strains of influenza viruses, revealing that Tfr cells promote recall antibody responses against the conserved HA stalk region. Thus, Tfr cells promote antigen-specific B cell responses and are essential for the development of long-term humoral memory., Competing Interests: Disclosures: K.C. O’Connor reported personal fees from Alexion and grants from Ra Pharma outside the submitted work; is the recipient of a sponsored research subaward from the University of Pennsylvania, the primary financial sponsor of which is Cabaletta Bio; and is a consultant and equity shareholder of Cabaletta Bio. No other disclosures were reported., (© 2020 Lu et al.)

Pathogenic muscle-specific tyrosine kinase (MuSK)-specific IgG4 autoantibodies in autoimmune myasthenia gravis (MG) are functionally monovalent as a result of Fab-arm exchange. The development of these unique autoantibodies is not well understood. We examined MG patient-derived monoclonal autoantibodies (mAbs), their corresponding germline-encoded unmutated common ancestors (UCAs), and monovalent antigen-binding fragments (Fabs) to investigate how affinity maturation contributes to binding and immunopathology. Mature mAbs, UCA mAbs, and mature monovalent Fabs bound to MuSK and demonstrated pathogenic capacity. However, monovalent UCA Fabs bound to MuSK but did not have measurable pathogenic capacity. Affinity of the UCA Fabs for MuSK was 100-fold lower than the subnanomolar affinity of the mature Fabs. Crystal structures of two Fabs revealed how mutations acquired during affinity maturation may contribute to increased MuSK-binding affinity. These findings indicate that the autoantigen drives autoimmunity in MuSK MG through the accumulation of somatic mutations such that monovalent IgG4 Fab-arm-exchanged autoantibodies reach a high-affinity threshold required for pathogenic capacity., Competing Interests: Disclosures: M.L. Fichtner reported grants from Grifols outside the submitted work. P. Stathopoulos reported grants from Onassis Foundation outside the submitted work. R.J. Nowak reported no conflicts directly related to this work. R.J. Nowak had received research support from Alexion, argenx, Ra Pharma, Momenta, Genentech, Immunovant, and Viela Bio related to the conduct of clinical trials for myasthenia gravis. K.C. O'Connor reported grants from Ra Pharma and personal fees from Alexion outside the submitted work; and received research support from Ra Pharma and is a consultant and equity shareholder of Cabaletta Bio. K.C. O'Connor is the recipient of a sponsored research subaward from the University of Pennsylvania, the primary financial sponsor of which is Cabaletta Bio. No other disclosures were reported., (© 2020 Fichtner et al.)

Myasthenia gravis (MG) is a neuromuscular, autoimmune disease caused by autoantibodies that target postsynaptic proteins, primarily the acetylcholine receptor (AChR) and inhibit signaling at the neuromuscular junction. The majority of patients under 50 y with AChR autoantibody MG have thymic lymphofollicular hyperplasia. The MG thymus is a reservoir of plasma cells that secrete disease-causing AChR autoantibodies and although thymectomy improves clinical scores, many patients fail to achieve complete stable remission without additional immunosuppressive treatments. We speculate that thymus-associated B cells and plasma cells persist in the circulation after thymectomy and that their persistence could explain incomplete responses to resection. We studied patients enrolled in a randomized clinical trial and used complementary modalities of B cell repertoire sequencing to characterize the thymus B cell repertoire and identify B cell clones that resided in the thymus and circulation before and 12 mo after thymectomy. Thymus-associated B cell clones were detected in the circulation by both mRNA-based and genomic DNA-based sequencing. These antigen-experienced B cells persisted in the circulation after thymectomy. Many circulating thymus-associated B cell clones were inferred to have originated and initially matured in the thymus before emigration from the thymus to the circulation. The persistence of thymus-associated B cells correlated with less favorable changes in clinical symptom measures, steroid dose required to manage symptoms, and marginal changes in AChR autoantibody titer. This investigation indicates that the diminished clinical response to thymectomy is related to persistent circulating thymus-associated B cell clones., Competing Interests: Competing interest statement: K.C.O. has received research support from Ra Pharma and is a consultant and equity shareholder of Cabaletta Bio. K.C.O. is the recipient of a sponsored research subaward from the University of Pennsylvania, the primary financial sponsor of which is Cabaletta Bio. S.H.K. receives consulting fees from Northrop Grumman. R.J.N. has received research support from Alexion Pharmaceuticals, Genentech, Grifols, and Ra Pharma. H.J.K. has served as an advisor to Alnylam Pharmaceuticals, Ra Pharmaceuticals, and UCB Pharmaceuticals, and is Chief Exectutive Officer and Chief Marketing Officer of ARC Biotechnology, LLC, based on US Patent 8,961,98.