Objective: To report on a novel neuronal target antigen in 3 patients with autoimmune cerebellar degeneration.Methods: Three patients with subacute to chronic cerebellar ataxia and controls underwent detailed clinical and neuropsychological assessment together with quantitative high-resolution structural MRI. Sera and CSF were subjected to comprehensive autoantibody screening by indirect immunofluorescence assay (IFA) and immunoblot. Immunoprecipitation with lysates of hippocampus and cerebellum combined with mass spectrometric analysis was used to identify the autoantigen, which was verified by recombinant expression in HEK293 cells and use in several immunoassays. Multiparameter flow cytometry was performed on peripheral blood and CSF, and peripheral blood was subjected to T-cell receptor spectratyping.Results: Patients presented with a subacute to chronic cerebellar and brainstem syndrome. MRI was consistent with cortical and cerebellar gray matter atrophy associated with subsequent neuroaxonal degeneration. IFA screening revealed strong immunoglobulin G1 reactivity in sera and CSF with hippocampal and cerebellar molecular and granular layers, but not with a panel of 30 recombinantly expressed established neural autoantigens. Neurochondrin was subsequently identified as the target antigen, verified by IFA and immunoblot with HEK293 cells expressing human neurochondrin as well as the ability of recombinant neurochondrin to neutralize the autoantibodies' tissue reaction. Immune phenotyping revealed intrathecal accumulation and activation of B and T cells during the acute but not chronic phase of the disease. T-cell receptor spectratyping suggested an antigen-specific T-cell response accompanying the formation of antineurochondrin autoantibodies. No such neurochondrin reactivity was found in control cohorts of various neural autoantibody-associated neurologic syndromes, relapsing-remitting multiple sclerosis, cerebellar type of multiple system atrophy, hereditary cerebellar ataxias, other neurologic disorders, or healthy donors. Conclusion:Neurochondrin is a neuronal target antigen in autoimmune cerebellar degeneration.Neurol Neuroimmunol Neuroinflamm 2017;4:e307; doi: 10.1212/NXI.0000000000000307 GLOSSARY AD 5 axial diffusivity; AI 5 antibody indices; DTI 5 diffusion tensor imaging; FA 5 fractional anisotropy; IFA 5 immunofluorescent assay; IgG 5 immunoglobulin G; IVIg 5 IV immunoglobulin; mGluR 5 metabotropic glutamate receptor; MSA-c 5 cerebellar type of multiple system atrophy; PB 5 peripheral blood; RD 5 radial diffusivity; RRMS 5 relapsing-remitting multiple sclerosis; SARA 5 Scale for the Assessment and Rating of Ataxia; TCR 5 T-cell receptor.Autoantibodies against neuronal constituents are associated with several severe immune-mediated CNS disorders. These disorders may predominantly affect gray matter structures of different brain regions such as the archicortex of the limbic system, neocortex, and basal ganglia, as well as cerebellar cortex and brainstem.

ObjectivesTo describe the neurologic spectrum and treatment outcomes for neurochondrin-IgG positive cases identified serologically in the Mayo Clinic Neuroimmunology Laboratory.MethodsArchived serum and CSF specimens previously scored positive for IgGs that stained mouse hippocampal tissue in a nonuniform synaptic pattern by immunofluorescence assay (89 among 616,025 screened, 1993–2019) were reevaluated. Antibody characterization experiments revealed specificity for neurochondrin, confirmed by recombinant protein assays.ResultsIgG in serum (9) or CSF (4) from 8 patients yielded identical neuron-restricted CNS patterns, most pronounced in hippocampus (stratum lucidum in particular), cerebellum (Purkinje cells and molecular layer), and amygdala. All were neurochondrin-IgG positive. Five were women; median symptom onset age was 43 years (range, 30–69). Of 7 with clinical data, 6 presented with rapidly progressive cerebellar ataxia, brainstem signs, or both; 1 had isolated unexplained psychosis 1 year prior. Five of 6 had cerebellar signs, 4 with additional brainstem symptoms or signs (eye movement abnormalities, 3; dysphagia, 2; nausea and vomiting, 1). One patient with brainstem signs (vocal cord paralysis and VII nerve palsy) had accompanying myelopathy (longitudinally extensive abnormality on MRI; aquaporin-4-IgG and myelin oligodendrocyte glycoprotein-IgG negative). The 7th patient had small fiber neuropathy only. Just 1 of 7 had contemporaneous cancer (uterine). Six patients with ataxia or brainstem signs received immunotherapy, but just 1 remained ambulatory. At last follow-up, 5 had MRI evidence of severe cerebellar atrophy.ConclusionIn our series, neurochondrin autoimmunity was usually accompanied by a nonparaneoplastic rapidly progressive rhombencephalitis with poor neurologic outcomes. Other phenotypes and occasional paraneoplastic causes may occur.

Background and Objectives: We sought to determine clinical significance of neuronal septin autoimmunity and evaluate for potential IgG effects. Methods: Septin-IgGs were detected by indirect immunofluorescence assays (IFAs; mouse tissue and cell based) or Western blot. IgG binding to (and internalization of) extracellular septin epitopes were evaluated for by live rat hippocampal neuron assay. The impact of purified patient IgGs on murine cortical neuron function was determined by recording extracellular field potentials in a multielectrode array platform. Results: Septin-IgGs were identified in 23 patients. All 8 patients with septin-5-IgG detected had cerebellar ataxia, and 7 had prominent eye movement disorders. One of 2 patients with co-existing septin-7-IgG had additional psychiatric phenotype (apathy, emotional blunting, and poor insight). Fifteen patients had septin-7 autoimmunity, without septin-5-IgG detected. Disorders included encephalopathy (11; 2 patients with accompanying myelopathy, and 2 were relapsing), myelopathy (3), and episodic ataxia (1). Psychiatric symptoms (≥1 of agitation, apathy, catatonia, disorganized thinking, and paranoia) were prominent in 6 of 11 patients with encephalopathic symptoms. Eight of 10 patients with data available (from 23 total) improved after immunotherapy, and a further 2 patients improved spontaneously. Staining of plasma membranes of live hippocampal neurons produced by patient IgGs (subclasses 1 and 2) colocalized with pre-and post-synaptic markers. Decreased spiking and bursting behavior in mixed cultures of murine glutamatergic and GABAergic cortical neurons produced by patient IgGs were attributable to neither antigenic crosslinking and internalization nor complement activation. Interpretation: Septin-IgGs are predictive of distinct treatment-responsive autoimmune central nervous system (CNS) disorders. Live neuron binding and induced electrophysiologic effects by patient IgGs may support septin-specific pathophysiology.

BackgroundA plurality of neurological syndromes is associated with autoantibodies against neural antigens relevant for diagnosis and therapy. Identification of these antigens is crucial to understand the pathogenesis and to develop specific immunoassays. Using an indirect immunofluorescence assay (IFA)-based approach and applying different immunoprecipitation (IP), chromatographic and mass spectrometric protocols was possible to isolate and identify a spectrum of autoantigens from brain tissue.MethodsSera and CSF of 320 patients suspected of suffering from an autoimmune neurological syndrome were comprehensively investigated for the presence of anti-neural IgG autoantibodies by IFA using mosaics of biochips with brain tissue cryosections and established cell-based recombinant antigen substrates as well as immunoblots. Samples containing unknown brain tissue-specific autoantibodies were subjected to IP with cryosections of cerebellum and hippocampus (rat, pig, and monkey) immobilized to glass slides or with lysates produced from homogenized tissue, followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, tryptic digestion, and matrix-assisted laser desorption/ionization–time of flight mass spectrometry analysis. Identifications were confirmed by IFA with recombinant HEK293 cells and by neutralizing the patients’ autoantibodies with the respective recombinantly expressed antigens in the tissue-based immunofluorescence test.ResultsMost samples used in this study produced speckled, granular, or homogenous stainings of the hippocampal and cerebellar molecular and/or granular layers. Others exclusively stained the Purkinje cells. Up to now, more than 20 different autoantigens could be identified by this approach, among them ATP1A3, CPT1C, Flotillin1/2, ITPR1, NBCe1, NCDN, RGS8, ROCK2, and Syntaxin-1B as novel autoantigens.DiscussionThe presented antigen identification strategy offers an opportunity for identifying up to now unknown neural autoantigens. Recombinant cell substrates containing the newly identified antigens can be used in serology and the clinical relevance of the autoantibodies can be rapidly evaluated in cohort studies.

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Objective:To report on a Caucasian patient who developed steroid-responsive transverse myelitis, graft vs host disease of the gut, and anti-GluRδ2 after allogenic stem cell transplantation.Methods:Histoimmunoprecipitation (HIP) with the patient's serum and cryosections of rat and porcine cerebellum followed by mass spectrometry was used to identify the autoantigen. Correct identification was verified by indirect immunofluorescence using recombinant GluRδ2 expressed in HEK293 cells.Results:The patient's serum produced a granular staining of the cerebellar molecular layer (immunoglobulin G1 and immunoglobulin G3; endpoint titer: 1:1,000) but did not react with other CNS tissues or 28 established recombinant neural autoantigens. HIP revealed a unique protein band at ∼110 kDa that was identified as GluRδ2. The patient's serum also stained GluRδ2 transfected but not mock-transfected HEK293 cells. Control sera from 38 patients with multiple sclerosis, 85 patients with other neural autoantibodies, and 205 healthy blood donors were negative for anti-GluRδ2. Preadsorption with lysate from HEK293-GluRδ2 neutralized the patient's tissue reaction whereas control lysate had no effect. In addition to anti-GluRδ2, the patient's serum contained immunoglobulin G autoantibodies against the pancreatic glycoprotein CUZD1, which are known to be markers of Crohn disease.Conclusions:In the present case, the development of anti-GluRδ2 was associated with transverse myelitis, which was supposedly triggered by the stem cell transplantation. Similar to encephalitis in conjunction with anti-GluRδ2 reported in a few Japanese patients, the patient's neurologic symptoms ameliorated after steroid therapy.

We describe a case of an anti-ATP1A3-associated neurologic disorder. Mutations in the gene encoding this neuronal surface protein have already been recognized as the cause of infantile alternating hemiplegia, rapid-onset dystonia parkinsonism, and CAPOS syndrome. Although the autoantibodies are unlikely to be pathogenic, they are likely to be rare biomarkers for the apparently paraneoplastic neurologic syndrome or for the tumor itself.

ObjectiveTo describe the identification of regulator of G-protein signaling 8 (RGS8) as an autoantibody target in patients with cerebellar syndrome associated with lymphoma.MethodsSera of 4 patients with a very similar unclassified reactivity against cerebellar Purkinje cells were used in antigen identification experiments. Immunoprecipitations with cerebellar lysates followed by mass spectrometry identified the autoantigen, which was verified by recombinant immunofluorescence assay, immunoblot, and ELISA with the recombinant protein.ResultsThe sera and CSF of 4 patients stained the Purkinje cells and molecular layer of the cerebellum. RGS8 was identified as the target antigen in all 4 sera. In a neutralization experiment, recombinant human RGS8 was able to neutralize the autoantibodies' tissue reaction. Patient sera and CSF showed a specific reactivity against recombinant RGS8 in ELISA and immunoblot, whereas no such reactivity was detectable in the controls. Clinical data were available for 2 of the 4 patients, remarkably both presented with cerebellar syndrome accompanied by B-cell lymphoma of the stomach (patient 1, 53 years) or Hodgkin lymphoma (patient 2, 74 years).ConclusionOur results indicate that autoantibodies against the intracellular Purkinje cell protein RGS8 represent new markers for paraneoplastic cerebellar syndrome associated with lymphoma.Classification of EvidenceThis study provided Class IV evidence that autoantibodies against the intracellular Purkinje cell protein RGS8 are associated with paraneoplastic cerebellar syndrome in lymphoma.