Camilla Buckley

Antibodies to Kv1 potassium channel-complex proteins leucine-rich, glioma inactivated 1 protein and contactin-associated protein-2 in limbic encephalitis, Morvan’s syndrome and acquired neuromyotonia

Antibodies that immunoprecipitate 125I-α-dendrotoxin-labelled voltage-gated potassium channels extracted from mammalian brain tissue have been identified in patients with neuromyotonia, Morvan’s syndrome, limbic encephalitis and a few cases of adult-onset epilepsy. These conditions often improve following immunomodulatory therapies. However, the proportions of the different syndromes, the numbers with associated tumours and the relationships with potassium channel subunit antibody specificities have been unclear. We documented the clinical phenotype and tumour associations in 96 potassium channel antibody positive patients (titres >400 pM). Five had thymomas and one had an endometrial adenocarcinoma. To define the antibody specificities, we looked for binding of serum antibodies and their effects on potassium channel currents using human embryonic kidney cells expressing the potassium channel subunits. Surprisingly, only three of the patients had antibodies directed against the potassium channel subunits. By contrast, we found antibodies to three proteins that are complexed with 125I-α-dendrotoxin-labelled potassium channels in brain extracts: (i) contactin-associated protein-2 that is localized at the juxtaparanodes in myelinated axons; (ii) leucine-rich, glioma inactivated 1 protein that is most strongly expressed in the hippocampus; and (iii) Tag-1/contactin-2 that associates with contactin-associated protein-2. Antibodies to Kv1 subunits were found in three sera, to contactin-associated protein-2 in 19 sera, to leucine-rich, glioma inactivated 1 protein in 55 sera and to contactin-2 in five sera, four of which were also positive for the other antibodies. The remaining 18 sera were negative for potassium channel subunits and associated proteins by the methods employed. Of the 19 patients with contactin-associated protein-antibody-2, 10 had neuromyotonia or Morvan’s syndrome, compared with only 3 of the 55 leucine-rich, glioma inactivated 1 protein-antibody positive patients (P < 0.0001), who predominantly had limbic encephalitis. The responses to immunomodulatory therapies, defined by changes in modified Rankin scores, were good except in the patients with tumours, who all had contactin-associated-2 protein antibodies. This study confirms that the majority of patients with high potassium channel antibodies have limbic encephalitis without tumours. The identification of leucine-rich, glioma inactivated 1 protein and contactin-associated protein-2 as the major targets of potassium channel antibodies, and their associations with different clinical features, begins to explain the diversity of these syndromes; furthermore, detection of contactin-associated protein-2 antibodies should help identify the risk of an underlying tumour and a poor prognosis in future patients.

Potassium channel antibodies in two patients with reversible limbic encephalitis

Limbic encephalitis (LE) is often associated with lung, thymic, or testicular tumours and antibodies to Hu, CV2, or Ma2 (Ta) antigens. In these cases, it generally has a poor prognosis. Here we describe two patients with symptoms of LE, negative for typical paraneoplastic antibodies, in whom antibodies to voltage‐gated potassium channels (VGKC) were detected retrospectively in serial serum samples. Patient 1 had a thymoma recurrence, but in patient 2 no tumour has been detected in the years following presentation. Plasma exchange was effective in reducing VGKC antibody levels, with substantial improvement in mental symptoms in patient 1. In patient 2, the VGKC antibodies fell spontaneously over two years, with almost complete recovery of mental function. Although neither patient had obvious neuromyotonia at presentation, both showed excessive secretions. We suggest that patients with limbic symptoms and excessive secretions should be tested for VGKC antibodies, and, if they are present, prompt and effective immunosuppressive treatment should be considered.

Morvan syndrome: clinical and serological observations in 29 cases.

A study was undertaken to describe the clinical spectrum, voltage‐gated potassium channel (VGKC) complex antibody specificities, and central nervous system localization of antibody binding in 29 patients diagnosed with Morvan syndrome (MoS).

Evidence of underdiagnosis of myasthenia gravis in older people

Background: Myasthenia gravis is a potentially serious but treatable muscle disease caused by autoantibodies directed at the acetylcholine receptor (AChR) on the postsynaptic membrane of the neuromuscular junction. There is anecdotal evidence that the diagnosis is sometimes missed in older patients. Objective: To examine the incidence and age distribution of positive AChR antibodies in samples referred to diagnostic laboratories in the UK, and the prevalence of positive AChR antibodies in samples from a cohort of older individuals. Methods: Positive AChR antibody tests were identified from all UK centres registered for the assay with the European quality assurance scheme (EQAS) during 1997–99, and the age and sex specific incidence was calculated, based on the UK population. The prevalence of AChR antibodies in sera from a sample of 2000 individuals aged ≥ 60 years was determined. Results: 3183 individuals had positive AChR antibody tests on routine screening during the years 1997 to 1999 in the UK, giving an annual incidence of 1.8/100 000. In both sexes, the age specific incidence rose steeply between the ages of 45 and 74, reaching 9.9/100 000 in men, and then fell, with a sharp decline above the age of 80. In the prevalence study, whereas only one serum from individuals aged 60–74 years was positive for AChR antibodies (0.12%), sera from eight individuals aged ≥ 75 years were positive (0.7%). Only one had a previous clinical diagnosis of myasthenia gravis but four others had histories of stroke or transient ischaemic attacks. Conclusions: The sharp fall in the incidence of clinically recognised myasthenia gravis in people over 80 years of age in our national AChR antibody incidence study, and the high prevalence of previously unrecognised positive AChR antibodies in those ≥ 75 years old, suggest that myasthenia gravis may be substantially underdiagnosed in older people.

Randomized Trial of Thymectomy in Myasthenia Gravis

BACKGROUND Thymectomy has been a mainstay in the treatment of myasthenia gravis, but there is no conclusive evidence of its benefit. We conducted a multicenter, randomized trial comparing thymectomy plus prednisone with prednisone alone. METHODS We compared extended transsternal thymectomy plus alternate-day prednisone with alternate-day prednisone alone. Patients 18 to 65 years of age who had generalized nonthymomatous myasthenia gravis with a disease duration of less than 5 years were included if they had Myasthenia Gravis Foundation of America clinical class II to IV disease (on a scale from I to V, with higher classes indicating more severe disease) and elevated circulating concentrations of acetylcholine-receptor antibody. The primary outcomes were the time-weighted average Quantitative Myasthenia Gravis score (on a scale from 0 to 39, with higher scores indicating more severe disease) over a 3-year period, as assessed by means of blinded rating, and the time-weighted average required dose of prednisone over a 3-year period. RESULTS A total of 126 patients underwent randomization between 2006 and 2012 at 36 sites. Patients who underwent thymectomy had a lower time-weighted average Quantitative Myasthenia Gravis score over a 3-year period than those who received prednisone alone (6.15 vs. 8.99, P<0.001); patients in the thymectomy group also had a lower average requirement for alternate-day prednisone (44 mg vs. 60 mg, P<0.001). Fewer patients in the thymectomy group than in the prednisone-only group required immunosuppression with azathioprine (17% vs. 48%, P<0.001) or were hospitalized for exacerbations (9% vs. 37%, P<0.001). The number of patients with treatment-associated complications did not differ significantly between groups (P=0.73), but patients in the thymectomy group had fewer treatment-associated symptoms related to immunosuppressive medications (P<0.001) and lower distress levels related to symptoms (P=0.003). CONCLUSIONS Thymectomy improved clinical outcomes over a 3-year period in patients with nonthymomatous myasthenia gravis. (Funded by the National Institute of Neurological Disorders and Stroke and others; MGTX ClinicalTrials.gov number, NCT00294658.).

Mature, long-lived CD4+ and CD8+ T cells are generated by the thymoma in myasthenia gravis

Antibodies to muscle acetylcholine receptors, to other muscle antigens, and to some cytokines are found in the majority of patients with thymic tumors (thymomas) and myasthenia gravis (MG). The role of the tumor in initiating autoimmunity, however, is unclear; in particular, it is not known whether the thymoma exports mature and long‐lived T cells, which could provide help for antibody production in the periphery. Here, we quantified recently exported thymic T cells using the approach of measuring episomal DNA fragments [T‐cell receptor excision circles (TRECs)], generated by T‐cell receptor gene rearrangement. Compared to values in healthy individuals (n = 10) or in patients with late‐onset MG (n = 8), TREC levels were significantly raised in both the CD4+ and CD8+ peripheral blood compartments of patients with thymoma and MG (n = 14, p = 0.002 and p = 0.0004 compared to healthy controls) but only in the CD8+ compartment of the 3 patients with thymoma without MG (p = 0.4 and p = 0.01 for CD4+ and CD8+). TREC levels decreased following thymectomy to values similar to controls but were substantially raised in patients who had developed tumor recurrence (n = 6, p = 0.04 and p = 0.02 for CD4+ and CD8+); this was associated with increased antibodies to interferon‐α and interleukin‐12 in the one case studied serially. Collectively, these results support the hypothesis that the neoplastic thymoma tissue itself can generate and export mature, long‐lived T cells and that these T cells reflect the thymic pathology and are likely to be related to the associated autoimmune diseases. The results also provide a new approach for early diagnosis of thymoma recurrence.

Do titin and cytokine antibodies in MG patients predict thymoma or thymoma recurrence

Background: Patients with MG often have other autoantibodies in addition to those against the acetylcholine receptor (AChR). It has been suggested that antibodies to the muscle protein titin may be diagnostic of a thymoma, but they have also been found in patients with late-onset MG. Antibodies to certain cytokines have also been detected in patients with MG and thymoma, and it is not clear whether these antibodies could be more useful clinically. The authors measured antibodies against titin and the cytokines interferon α (IFNα) and interleukin 12 (IL12) in patients with MG and thymoma or thymoma recurrence, and in patients with MG but without thymoma presenting before (early-onset MG) or after (late-onset MG) 40 years of age. Method: Levels of titin, IFNα, and IL12 antibodies were determined by radioimmunoassay in 191 patients with MG and 82 controls. Results: As previously reported, titin antibodies were uncommon in patients with early-onset MG. However, in patients with late-onset MG, titin antibodies had similar prevalence and levels to those in patients with MG and thymoma, although the antibodies were uncommon in patients between 40 and 60 years of age presenting without a tumor. By contrast, cytokine antibodies were more common in patients with thymoma than in patients without thymoma, and cytokine antibodies typically increased substantially if the thymoma recurred. Conclusions: Measurement of titin antibodies has limited use in predicting the presence of a tumor, unless the patient is less than 60 years of age, but measurement of IFNα and IL12 antibodies may be helpful in identifying patients with a thymoma recurrence, particularly when mediastinal imaging is equivocal.

Antibodies to GABAA receptor α1 and γ2 subunits: clinical and serologic characterization.

Objective: To search for antibodies against neuronal cell surface proteins. Methods: Using immunoprecipitation from neuronal cultures and tandem mass spectrometry, we identified antibodies against the α1 subunit of the γ-aminobutyric acid A receptor (GABAAR) in a patient whose immunoglobulin G (IgG) antibodies bound to hippocampal neurons. We searched 2,548 sera for antibodies binding to GABAAR α, β, and γ subunits on live HEK293 cells and identified the class, subclass, and GABAAR subunit specificities of the positive samples. Results: GABAAR-Abs were identified in 40 of 2,046 (2%) referred sera previously found negative for neuronal antibodies, in 5/502 (1%) previously positive for other neuronal surface antibodies, but not in 92 healthy individuals. The antibodies in 40% bound to either the α1 (9/45, 20%) or the γ2 subunits (9/45, 20%) and were of IgG1 (94%) or IgG3 (6%) subclass. The remaining 60% had lower antibody titers (p = 0.0005), which were mainly immunoglobulin M (IgM) (p = 0.0025), and showed no defined subunit specificity. Incubation of primary hippocampal neurons with GABAAR IgG1 sera reduced surface GABAAR membrane expression. The clinical features of 15 patients (GABAAR α1 n = 6, γ2 n = 5, undefined n = 4) included seizures (47%), memory impairment (47%), hallucinations (33%), or anxiety (20%). Most patients had not been given immunotherapies, but one with new-onset treatment-resistant catatonia made substantial improvement after plasma exchange. Conclusions: The GABAAR α1 and γ2 are new targets for antibodies in autoimmune neurologic disease. The full spectrum of clinical features, treatment responses, correlation with antibody specificity, and in particular the role of the IgM antibodies will need to be assessed in future studies.

NMDA receptor antibodies associated with distinct white matter syndromes

Objective: To report the clinical and radiologic findings of children with NMDA receptor (NMDAR) antibodies and white matter disorders. Method: Ten children with significant white matter involvement, with or without anti-NMDAR encephalitis, were identified from 46 consecutive NMDAR antibody–positive pediatric patients. Clinical and neuroimaging features were reviewed and the treatment and outcomes of the neurologic syndromes evaluated. Results: Three distinct clinicoradiologic phenotypes were recognized: brainstem encephalitis (n = 3), leukoencephalopathy following herpes simplex virus encephalitis (HSVE) (n = 2), and acquired demyelination syndromes (ADS) (n = 5); 3 of the 5 with ADS had myelin oligodendrocyte glycoprotein as well as NMDAR antibodies. Typical NMDAR antibody encephalitis was seen in 3 patients remote from the first neurologic syndrome (2 brainstem, 1 post-HSVE). Six of the 7 patients (85%) who were treated acutely, during the original presentation with white matter involvement, improved following immunotherapy with steroids, IV immunoglobulin, and plasma exchange, either individually or in combination. Two patients had escalation of immunotherapy at relapse resulting in clinical improvement. The time course of clinical features, treatments, and recoveries correlated broadly with available serum antibody titers. Conclusion: Clinicoradiologic evidence of white matter involvement, often distinct, was identified in 22% of children with NMDAR antibodies and appears immunotherapy responsive, particularly when treated in the acute phase of neurologic presentation. When observed, this clinical improvement is often mirrored by reduction in NMDAR antibody levels, suggesting that these antibodies may mediate the white matter disease.

Clinical Features and Diagnostic Usefulness of Antibodies to Clustered Acetylcholine Receptors in the Diagnosis of Seronegative Myasthenia Gravis.

IMPORTANCE Cell-based assays (CBAs) were shown to improve detection of acetylcholine receptor (AChR) antibodies in patients with myasthenia gravis (MG). Herein, we asked whether these assays were able to help determine the diagnosis in patients studied in routine clinical practice. OBJECTIVES To determine the diagnostic usefulness of CBAs in the diagnosis of MG and to compare the clinical features of patients with antibodies only to clustered AChRs with those of patients with seronegative MG (SNMG). DESIGN, SETTING, AND PARTICIPANTS All patients with clinical suspicion of MG who were seen within the Division of Clinical Neurology at the John Radcliffe Hospital in Oxford, England, between November 1, 2009, and November 30, 2013. Their serum antibodies and clinical features were studied. EXPOSURES Radioimmunoprecipitation assay (RIPA) and CBA were used to test for standard AChR antibodies and antibodies to clustered AChRs in 138 patients. All available samples from patients with SNMG were retrospectively tested for lipoprotein receptor-related protein 4 (LRP4) antibodies. MAIN OUTCOMES AND MEASURES Demographic, clinical, neurophysiological, and laboratory data. RESULTS In total, 138 patients were tested for antibodies to clustered AChRs, and 42 had a final diagnosis of MG. The clustered AChR CBA detected antibodies in 38.1% (16 of 42) of RIPA-negative patients with MG with 100% specificity. All patients with SNMG who were tested for LRP4 antibodies (21 of 26) were negative by CBA. Compared with patients with SNMG, patients with antibodies only to clustered AChRs had frequent prepubertal onset (62.5% [median age, 6 years; age range, 1-52 years] vs 11.5% [median age, 38 years; age range, 2-72 years], P ≤ .05), high prevalence of ocular MG (62.5% vs 42.3%), milder disease severity with less bulbar involvement (25.0% vs 46.2%), and absence of respiratory symptoms (0% vs 23.1%). Response to treatment and prognosis was good, with a reduced need for thymectomy (6.3% vs 19.2%) and a high proportion of patients going into remission (50.0% vs 8.3%, P ≤ .05). These observations also apply to the classic AChR MG phenotype seen in large series. CONCLUSIONS AND RELEVANCE Cell-based assay is a useful procedure in the routine diagnosis of RIPA-negative MG, particularly in children. Patients with antibodies only to clustered AChRs appear to be younger and have milder disease than other patients with MG. These observations will have implications in planning treatment.