Kerri Prain

Use and monitoring of low dose rituximab in myasthenia gravis

Background Myasthenia gravis is an autoimmune disorder of the neuromuscular junction. Rituximab (RTX), a monoclonal antibody to CD20, leads to B lymphocyte depletion and has been used in some autoimmune disorders, including small case series of myasthenia gravis patients. Methods A retrospective analysis was performed of all patients with acetylcholine receptor (AChR) (11 subjects) or muscle specific kinase antibody (MuSK) positive myasthenia gravis (three subjects), who had been treated with RTX in Brisbane, Australia. In most patients 1 g of RTX, in two divided doses, was given. Patients were monitored by serial clinical assessments, flow cytometry of peripheral blood B lymphocytes and antibody testing. Results RTX led to a significant improvement in symptoms in 11 of 14 patients. Doses of immunosuppressive medications were able to be reduced in 12 of 14 patients but medications could be completely ceased in only one patient. A demonstrable reduction of autoantibody levels was found in only three AChR positive patients and one MuSK positive patient, independent of clinical improvement. Peripheral blood B lymphocyte depletion was achieved in 13 out of 14 patients. B lymphocyte recovery occurred between 9 and 30 months post RTX (median 12.3 months) and was consistently associated with worsening of clinical symptoms. Conclusion Rituximab at a dose of 1 g appears to be beneficial in the treatment of patients with severe myasthenia gravis. Serial monitoring of peripheral blood B lymphocytes appears to be useful in guiding the need for further RTX therapy.

Molecular Pathogenesis of Neuromyelitis Optica

Neuromyelitis optica (NMO) is a rare autoimmune disorder, distinct from multiple sclerosis, causing inflammatory lesions in the optic nerves and spinal cord. An autoantibody (NMO IgG) against aquaporin-4 (AQP4), a water channel expressed on astrocytes is thought to be causative. Peripheral production of the antibody is triggered by an unknown process in genetically susceptible individuals. Anti-AQP4 antibody enters the central nervous system (CNS) when the blood brain barrier is made permeable and has high affinity for orthogonal array particles of AQP4. Like other autoimmune diseases, Th17 cells and their effector cytokines (such as interleukin 6) have been implicated in pathogenesis. AQP4 expressing peripheral organs are not affected by NMO IgG, but the antibody causes extensive astrocytic loss in specific regions of the CNS through complement mediated cytotoxicity. Demyelination occurs during the inflammatory process and is probably secondary to oligodendrocyte apoptosis subsequent to loss of trophic support from astrocytes. Ultimately, extensive axonal injury leads to severe disability. Despite rapid advances in the understanding of NMO pathogenesis, unanswered questions remain, particularly with regards to disease mechanisms in NMO IgG seronegative cases. Increasing knowledge of the molecular pathology is leading to improved treatment strategies.

Incidence and prevalence of NMOSD in Australia and New Zealand

Objectives We have undertaken a clinic-based survey of neuromyelitis optica spectrum disorders (NMOSDs) in Australia and New Zealand to establish incidence and prevalence across the region and in populations of differing ancestry. Background NMOSD is a recently defined demyelinating disease of the central nervous system (CNS). The incidence and prevalence of NMOSD in Australia and New Zealand has not been established. Methods Centres managing patients with demyelinating disease of the CNS across Australia and New Zealand reported patients with clinical and laboratory features that were suspicious for NMOSD. Testing for aquaporin 4 antibodies was undertaken in all suspected cases. From this group, cases were identified who fulfilled the 2015 Wingerchuk diagnostic criteria for NMOSD. A capture–recapture methodology was used to estimate incidence and prevalence, based on additional laboratory identified cases. Results NMOSD was confirmed in 81/170 (48%) cases referred. Capture–recapture analysis gave an adjusted incidence estimate of 0.37 (95% CI 0.35 to 0.39) per million per year and a prevalence estimate for NMOSD of 0.70 (95% CI 0.61 to 0.78) per 100 000. NMOSD was three times more common in the Asian population (1.57 (95% CI 1.15 to 1.98) per 100 000) compared with the remainder of the population (0.57 (95% CI 0.50 to 0.65) per 100 000). The latitudinal gradient evident in multiple sclerosis was not seen in NMOSD. Conclusions NMOSD incidence and prevalence in Australia and New Zealand are comparable with figures from other populations of largely European ancestry. We found NMOSD to be more common in the population with Asian ancestry.

Diagnostic Value of Distinguishing and Reporting Different Perinuclear ANCA (P-ANCA) Immunofluorescence Patterns

OBJECTIVES To investigate whether discriminating the classic perinuclear antineutrophil cytoplasmic antibody (P-ANCA) pattern from atypical P-ANCA and uninterpretable patterns improves the diagnostic utility of ANCA testing. METHODS All ANCA requests (n = 3,544) referred to Pathology Queensland were analyzed prospectively over 4 months for P-ANCA pattern subtypes and myeloperoxidase (MPO)-ANCA/PR3-ANCA results and correlated with clinical, laboratory, and radiologic evidence of necrotizing small vessel vasculitis. RESULTS Of the 436 perinuclear immunofluorescence-positive samples, 45 were classic P-ANCA, 163 were atypical P-ANCA, and 228 were antinuclear antibodies/uninterpretable. The classic P-ANCA pattern had a significantly stronger association with vasculitis (30/45) than atypical P-ANCA (2/163) (P <.0001) or ANA/uninterpretable patterns (8/228) (P <.0001). The combination of a classic P-ANCA pattern and positive MPO-ANCA/PR3-ANCA result was also more strongly associated with vasculitis than a positive MPO-ANCA/PR3-ANCA result in isolation (P = .003). CONCLUSIONS This study demonstrates that reporting different P-ANCA patterns (including ANA/uninterpretable patterns) provides additional diagnostic information to MPO-ANCA/PR3-ANCA results.

The prevalence and treatment outcomes of antineuronal antibody-positive patients admitted with first episode of psychosis

Background Antineuronal antibodies are associated with psychosis, although their clinical significance in first episode of psychosis (FEP) is undetermined. Aims To examine all patients admitted for treatment of FEP for antineuronal antibodies and describe clinical presentations and treatment outcomes in those who were antibody positive. Method Individuals admitted for FEP to six mental health units in Queensland, Australia, were prospectively tested for serum antineuronal antibodies. Antibody-positive patients were referred for neurological and immunological assessment and therapy. Results Of 113 consenting participants, six had antineuronal antibodies (anti-N-methyl-D-aspartate receptor antibodies [n = 4], voltage-gated potassium channel antibodies [n = 1] and antibodies against uncharacterised antigen [n = 1]). Five received immunotherapy, which prompted resolution of psychosis in four. Conclusions A small subgroup of patients admitted to hospital with FEP have antineuronal antibodies detectable in serum and are responsive to immunotherapy. Early diagnosis and treatment is critical to optimise recovery. Declaration of interest None.

059 Teratoma with glycine, NMDA & VGKC antibodies presenting with severe weakness, respiratory failure & corticospinal tract dysfunction

Introduction Glycine antibodies are associated with progressive encephalomyelitis with rigidity and myoclonus (PERM) but have been rarely reported to cause myasthenic-like symptoms. 1 We present a single patient seen at a tertiary neurology centre with fatigable muscle weakness and corticospinal tract dysfunction with positive glycine, VGKC and NMDA receptor antibodies, associated with an ovarian teratoma expressing neural tissue. Case A 32 year old female presented with fatigable muscle weakness. Physical examination was atypical for a disease of the neuromuscular junction with features of exaggerated reflexes and clonus. The weakness progressed over 1–2 weeks leading to respiratory failure. Investigations including MRI brain, repetitive stimulation and acetylcholine-receptor antibody studies were all within the normal limits. Studies for neuronal antibodies against intracellular targets were negative. Further investigation revealed positive autoantibodies against Glycine receptor, NMDA (N-Methyl-d-aspartate) receptor and VGKC (voltage-gated potassium channel) with an underlying ovarian teratoma. Complete clinical resolution was achieved with teratoma resection and 2 g/kg course of intravenous immunoglobulin. Histopathological examination of the tumour revealed a mature cystic teratoma with dystrophic calcification. The teratoma included skin, adipose tissue, intestinal type mucosa, and neuroglial tissue in which nerve fibres and ganglion cells are present. A lymphoid infiltrate was concentrated in areas of neural tissue within the teratoma. Conclusion Muscle weakness, corticospinal tract dysfunction and respiratory failure have previously been reported in the context of anti-glycine antibodies. This is the first case where anti-glycine, anti-NMDA and anti-VGKC antibodies have been found in the context of an ovarian teratoma. The autoimmune nature of this condition is emphasised by the lymphoid infiltrate around the neural tissue expressed within the teratoma. Reference 1. Carvajal-Gonzalez A, Leite MI, Waters P, Woodhall M, Coutinho E, Balint B, … Vincent A. Glycine receptor antibodies in PERM and related syndromes: characteristics, clinical features and outcomes. Brain2014;137(Pt 8):2178–2192.

9.: Characteristics of aquaporin 4 antibodies associated with lesions of the area postrema

Whilst testing for aquaporin 4 antibodies (AQP4 Ab) in a large neuroimmunology laboratory we encountered several patients with an atypical AQP4 Ab test result, which were associated with a distinct clinical picture and findings on neuroimaging. We postulated that in these patients there may be a different target antigen as compared to that of classical neuromyelitis optica. We studied a clinical case series of patients with positive AQP4 Ab testing, performed using both a cell based assay (CBA) and immunofluorescence on rodent brain sections. We describe the clinical presentation, MRI changes, serological markers and treatment outcome of several patients with an atypical AQP4 Ab test result, namely positive binding on the CBA, but an absence of staining on rodent brain sections. We propose that the clinical syndrome seen in our case series is distinct from that seen in classical neuromyelitis optica. We suggest that there are subclasses of AQP4 Ab which are associated with characteristic clinical and neuroimaging findings, and will discuss possible mechanisms.

New autoantibodies in neurological diseases

Introduction The spectrum of antibodies against neural tissues detected by indirect immunofluorescence (IIF) has expanded rapidly. There are now approximately 20 anti-neuronal antibodies (PNAs) detected in association with neurological diseases, especially with underlying malignancies. Many PNAs have been correlated with disease and neurological defects and appear to fall into two groups: those that are cytotoxic (PCA-1 and ANNA-1); and others that are inhibitory (anti-NMDAR, anti-VGCC and anti-VGKC). We present a series of novel antibodies that stain a variety of CNS structures in patients with diverse neurological presentations. Patients and Methods All patients in this study were referred for either anti-neuronal, anti-NMO-IgG (neuromyelitis optica) or anti-NMDAR (N-methyl-D-aspartate receptor) antibody testing. Discussion With the introduction of NMO-IgG IIF testing in 2007 and anti-NMDAR testing in 2010, 4377 patients with increasingly diverse neurological and psychiatric presentations were tested. We have now identified several cohorts of patients with a variety of anti-CNS antibodies targeting (1) cerebellar white matter fibres, (2) glia, (3) axons, and (4) astrocytes. Although the targets for these antibodies appear to be hetero-genous, several patients have staining patterns in common and the pattern of tissue staining often correlates with the clinical presentation. We are undertaking further studies to identify the precise nature of the target antigens.

The importance of distinguishing and reporting different perinuclear ANCA (P-ANCA) immunofluorescence patterns: a prospective study

Objectives: To investigate whether discriminating the classic perinuclear antineutrophil cytoplasmic antibody (P-ANCA) pattern from atypical P-ANCA and uninterpretable patterns improves the diagnostic utility of ANCA testing. Methods: All ANCA requests (n = 3,544) referred to Pathology Queensland were analyzed prospectively over 4 months for P-ANCA pattern subtypes and myeloperoxidase (MPO)-ANCA/PR3-ANCA results and correlated with clinical, laboratory, and radiologic evidence of necrotizing small vessel vasculitis. Results: Of the 436 perinuclear immunofluorescencepositive samples, 45 were classic P-ANCA, 163 were atypical P-ANCA, and 228 were antinuclear antibodies/ uninterpretable. The classic P-ANCA pattern had a significantly stronger association with vasculitis (30/45) than atypical P-ANCA (2/163) ( P <.0001) or ANA/ uninterpretable patterns (8/228) ( P <.0001). The combination of a classic P-ANCA pattern and positive MPO-ANCA/PR3-ANCA result was also more strongly associated with vasculitis than a positive MPO-ANCA/ PR3-ANCA result in isolation ( P = .003). Conclusions: This study demonstrates that reporting different P-ANCA patterns (including ANA/uninterpretable patterns) provides additional diagnostic information to MPO-ANCA/PR3-ANCA results. The antineutrophil cytoplasmic antibody (ANCA)–associated vasculitides are characterized by the presence of autoantibodies directed against constituents of neutrophil cytoplasmic granules. These conditions, which include granulomatosis with polyangiitis (GPA), microscopic polyangiitis (MPA), and eosinophilic granulomatosis with polyangiitis (EGPA), are characterized by the presence of systemic necrotizing small vessel vasculitis and are frequently associated with serious complications, including acute renal failure and potentially life-threatening diffuse alveolar hemorrhage. 1

Update on anti-NMO IgG antibodies

Neuromyelitis optica (NMO) or Devic’s syndrome is a severe idiopathic and relapsing, demyelinating disease characterised by optic neuritis (ON) and longitudinally extensive transverse myelitis (LETM). 1 Neuropathologically, NMO is characterised by extensive demyelination with partial necrosis of the spinal cord white and grey matter, acute axonal injury, neutrophil and eosinophil infiltrates, IgG and IgM deposition and perivascular complement activation. NMO is distinguished from classical multiple sclerosis (MS) by clinical, neuroimaging, CSF and serological criteria. 1 The identification of NMO-IgG by indirect immunofluorescence (IIF) was initially described as a highly specific marker with the titre paralleling the disease activity. However, the sensitivity of IIF is variable (58–75%) for clinical NMO. Consequently, numerous methods using aquaporin 4 (AQP4) as the target antigen have been developed in an attempt to improve sensitivity. Apart from cell-based assays, these have failed to improve on the sensitivity of IIF. Recently, it has been demonstrated that NMO-IgG targets mainly the M-23 isoform of AQP4 that structurally organises itself into orthogonol arrays of particles in cell membranes. As AQP4-Ab recognises conformational epitopes, the lower sensitivity of all but the cell-based assays may be due to changes resulting during purification, fixation or other causes of NMO, i.e., parainfectious NMO. As indirect immunofluorescence is the most widely available method for detecting NMO-IgG, the Pathology Queensland experience has raised issues of the ‘true’ sensitivity of NMO-IgG testing for NMO as well as identifying other autoantibodies (including anti-reticulin and anti-nuclear antibodies) that cause interference and make IIF interpretation difficult. These issues may contribute to both false positive and false negative IIF results. Some authors have suggested 2 that the published sensitivities of NMO-IgG may be artificially high and this has prompted the proposed Australian and New Zealand NMO study designed to evaluate the sensitivity and specificity of all currently available methods for NMO-IgG.