Expert Review of Clinical Immunology

ISSN: 1744-666X (Print) 1744-8409 (Online) Journal homepage: http://www.tandfonline.com/loi/ierm20

Overcoming challenges in the diagnosis and treatment of myasthenia gravis Amelia Evoli, Raffaele Iorio & Emanuela Bartoccioni To cite this article: Amelia Evoli, Raffaele Iorio & Emanuela Bartoccioni (2016) Overcoming challenges in the diagnosis and treatment of myasthenia gravis, Expert Review of Clinical Immunology, 12:2, 157-168, DOI: 10.1586/1744666X.2016.1110487 To link to this article: http://dx.doi.org/10.1586/1744666X.2016.1110487

Published online: 16 Dec 2015.

Submit your article to this journal

Article views: 291

View related articles

View Crossmark data

Citing articles: 1 View citing articles

Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=ierm20 Download by: [University of New England]

Date: 11 November 2017, At: 09:05

EXPERT REVIEW OF CLINICAL IMMUNOLOGY, 2016 VOL. 12, NO. 2, 157–168 http://dx.doi.org/10.1586/1744666X.2016.1110487

REVIEW

Overcoming challenges in the diagnosis and treatment of myasthenia gravis Amelia Evolia, Raffaele Iorioa,b and Emanuela Bartoccionic

Downloaded by [University of New England] at 09:05 11 November 2017

a Institute of Neurology, Catholic University, Roma, Italy; bDon Gnocchi ONLUS Foundation, Milan, Italy; cInstitute of General Pathology, Catholic University, Roma, Italy

ABSTRACT

ARTICLE HISTORY

In recent years, the discovery of new autoantigens and the use of sensitive assays have expanded the clinical spectrum of myasthenia gravis (MG). In particular, antibodies binding to clustered acetylcholine receptors and to the low-density lipoprotein receptor-related protein 4 have not only bridged a significant gap in diagnosis but also have relevant clinical implications. MG management includes different therapeutic options, from symptomatic agents as the only therapy in mildly affected cases to combined long-term immunosuppression and thymectomy in patients with severe disabling disease. MG biological diversity can influence the response to therapies and should be taken into account when planning treatment. Biologic agents are promising, though their use is currently limited to patients with refractory disease.

Received 31 August 2015 Accepted 16 October 2015

Introduction Myasthenia gravis (MG) is the most common disorder of neuromuscular junction (NMJ), with a prevalence approaching 200/million. It affects all races and can occur at any age, from the first year to the nineties. In Western countries, MG shows a peak age of onset in the third decade in women and a larger peak in the sixth and seventh decades in men, while in Asian countries there is a high prevalence of limited forms of the disease with childhood onset.[1] MG is increasingly recognized as a heterogeneous disease including distinct clinical entities. MG is due to autoantibodies (Abs) that, with different mechanisms, impair neuromuscular transmission (NMT) and cause fatigable weakness of voluntary muscles. The great majority of patients have serum Abs to the acetylcholine receptor (AChR). These Abs are mainly of IgG1 and IgG3 isotypes, induce severe alterations of the postsynaptic membrane through complement activation and increased AChR degradation, and, to a lesser extent, interfere with the neurotransmitter binding site. [2] MG with AChR Abs (AChR-MG) is frequently associated with thymus alterations that are thought to play a critical role in the disease pathogenesis. Thymus follicular hyperplasia (TFH), typically found in female patients with early onset MG, is considered the site where the autoimmune response to the AChR is initiated and maintained. Conversely, defects of T cell selection appears to be responsible for MG association CONTACT Amelia Evoli © 2015 Taylor & Francis

[email protected]

KEYWORDS

AChR; clustered-AChR; immunosuppressants; Lrp4; myasthenia gravis; MuSK; thymectomy

with thymoma, a tumor of thymic epithelial cells, that is present in 10–15% of patients, more commonly in the fifth and sixth decades of life.[3] Serum Abs to the muscle-specific tyrosine kinase receptor (MuSK) are detected in 5–8% of MG patients. MuSK is the core of a multiprotein complex, that is critical for AChR clustering and postsynaptic configuration.[4] As experimental models have shown, MuSK Abs interfere with the protein function[5]; although being mostly IgG4, they cannot activate complement and are rather inefficient in antigen cross-linking.[2] In addition, the low-density lipoprotein receptor-related protein 4 (Lrp4), a receptor for agrin that mediates MuSK activation,[6] has been identified as autoantigen in MG (see Figure 1). The frequency of Lrp4 Abs in patients with neither AChR- nor MuSK-Abs shows marked variability in different studies [7]; the Ab effects and clinical phenotypes are not fully characterized. While the search for novel antigens in MG is actively pursued, there are still some patients without detectable Abs. The frequency of these so-called ‘triple-negative’ cases is unknown. In MG, pathogenic heterogeneity is associated with marked variability in weakness extension and severity, from purely ocular symptoms to severe life-threatening disease. A correct diagnosis is crucial in order to offer patients appropriate treatment. MG management has steadily improved over the years. However, there are still uncertainties and controversies both in the diagnostic workup and therapeutic strategy. This review will discuss how recent advances

Downloaded by [University of New England] at 09:05 11 November 2017

158

A. EVOLI ET AL.

Figure 1. Schematic representation of the acetylcholine receptor, the muscle-specific tyrosine kinase receptor and low-density lipoprotein receptor-related protein 4 complex at the neuromuscular junction. AChE: acetylcholinesterase; AChR: acetylcholine receptor; Lrp4: Low-density lipoprotein receptor-related protein 4; MuSK: musclespecific tyrosine kinase receptor.Modified and reprinted with permission from Evoli and Iorio, Clinical Experimental Neuroimmunology, 2015; license number: 3696030334051.

in clinical and basic research may help clinicians in the care of these patients.

Establishing the diagnosis of MG MG is suspected in patients with history and signs of fluctuating weakness of voluntary muscles, worsening on exertion and improving at rest. Diagnosis confirmation is then achieved through: a) detection of serum Abs; b) electromyography (EMG) studies showing compound muscle action potential decrement on low-rate repetitive nerve stimulation (RNS) or increased jitter on single-fiber (SF) EMG; c) clinical response to acetylcholinesterase inhibitors (AChE-Is). Positive results on b) and c) confirm a postsynaptic defect of NMT, while detection of specific Abs establishes the diagnosis of MG. Routine serological testing includes AChR and MuSK Ab assays. Given to their high prevalence in MG, AChR Abs are the first to be tested when MG is suspected on clinical grounds; patients with negative results on this assay should be tested for MuSK Abs. The coexistence of these Abs is very rare.[8] AChR and MuSK Abs are considered very specific for MG and, in practice, their detection in patients with consistent symptoms confirms the diagnosis, with no real need for additional testing. On the other hand, when Ab assays are negative, EMG confirmation is crucial.

Electrophysiological examination should include conventional needle EMG and neurographic studies to rule out diseases that can mimic MG. RNS is the routine technique in the investigation of NMT disorders; it is considered fairly specific, though a decremental response can also be found in motor neuron disease and radiculopathy.[9] The diagnostic sensitivity of RNS is related to weakness pattern. The rate of positive results is around 70% in patients with generalized disease when both distal and proximal muscles are tested, while it is less than 50% in ocular myasthenia and in patients with focal cranial symptoms.[10] SF-EMG is regarded as the most sensitive diagnostic test in MG, as positive results can be recorded in >90% of patients, including those with limited weakness, when appropriate muscles are examined. However, increased jitter is far from specific and can be found in several neuropathic and myopathic conditions.[9] In addition, SF-EMG technique is time-consuming and requires patient’s cooperation. In clinical practice, the choice between RNS and SF-EMG depends on their relative sensitivity in relation to weakness extension: SF-EMG has a stronger indication in subjects with purely ocular or focal symptoms and in those with mild weakness; RNS is generally adequate in patients with more relevant generalized symptoms. A clear-cut clinical improvement on administration of short-acting AChE-Is, as edrophonium chloride iv. or

Downloaded by [University of New England] at 09:05 11 November 2017

EXPERT REVIEW OF CLINICAL IMMUNOLOGY

neostigmine im., strongly supports the diagnosis of MG and a positive response to this ‘pharmacological’ test has been reported in approximately 90% of MG patients.[10] However, the response rate is much lower (50–70%) in MuSK-MG [11]; in addition, a positive reaction to AChE-I is not specific for MG, as it is usually present in congenital myasthenic syndromes and can be found in Lambert–Eaton myasthenic syndrome, amyotrophic lateral sclerosis (ALS), Guillain–Barrè syndrome, and, even, in patients with intracranial tumors.[1] In patients with negative AChR and MuSK Ab assays, MG must first be differentiated from other primary disorders of NMT, while, in selected cases, ALS, Miller Fisher syndrome, cranial neuropathy and mitochondrial myopathy must be considered in the differential diagnosis. Among NMT disorders, Lambert–Eaton myasthenic syndrome and botulism show characteristic clinical and EMG findings that can assist in establishing the correct diagnosis, while MG differentiation from a congenital myasthenic syndrome may be far more challenging. Patients with purely ocular symptoms, and negative or equivocal results of EMG (including SFEMG) tests should undergo brain MRI, and MG diagnosis should be reconsidered after the exclusion of other conditions that may cause ptosis and/or diplopia. New Abs have recently added to laboratory testing in MG. Herein, we discuss the contribution of standard and newly described Ab assays to the clinical diagnosis.

Diagnostic yield of AChR and MuSK Abs AChR- and MuSK Abs are routinely measured by a radioimmunoprecipitation assay (RIPA) that is sensitive, quantitative and widely available. AChR Ab testing by enzyme-linked immunoassay (ELISA), that offers the obvious advantage of not requiring radioactive isotopes, proved less sensitive, especially in patients with border-line Ab titers.[12] By RIPA, Abs to AChR are detected as IgG binding to 125 I-α-bungarotoxin-labeled solubilized AChR (binding Abs). Modulating and blocking Abs, which can be measured with distinct assays,[13] increase to a limited extent the sensitivity of the standard RIPA and have not entered routine practice. Binding AChR Abs are detected in 80–90% of patients with generalized MG, in roughly 50% of those with symptoms restricted to ocular muscles, and in virtually all cases with thymomaassociated MG.[8,14] Diagnostic sensitivity is increased by testing nonimmunosuppressed patients, while AChR-negative patients with recent-onset disease may turn positive when retested after 6–12 months (seroconversion).[8] On the other hand, AChR Ab positivity

159

rate is lower in childhood-onset MG,[15] and in milder forms of the disease.[13] MuSK Abs are detected by the standard RIPA in 30–40% of AChR-negative patients, with variable frequency across populations.[8] These Abs are very rare in ocular MG and are most commonly associated with a clinical pattern of predominant weakness in cranial, neck and respiratory muscles. MuSK-MG is uncommon in childhood, with a mean age of onset in the fourth decade.[11] The diagnostic accuracy of the standard RIPA for AChR Ab ranges 97–99%,[10] as positive results may be rarely found in other autoimmune conditions and in thymoma without MG, while MuSK Abs have never been reported in non-MG patients.[8] On the whole, standard RIPA for AChR- and MuSK Abs can confirm the diagnosis of MG in around 90% patients, with a higher rate of positive results in adults with generalized disease. MG with neither AChR nor MuSK Abs on the standard assay (so-called double seronegative myasthenia gravis (dSN-MG) has variable frequency in different patient cohorts (in part, depending on the relative rate of MuSK-MG), wide clinical spectrum and broad range of age-of-onset. In recent years, development of sensitive cell-based assays (CBAs) has led to detection of new Abs in this heterogeneous population. In CBA, antigens are expressed in cell lines, usually human embryonic kidney cells, transfected with the appropriate cDNAs and the Ab binding is detected by indirect immunofluorescence. This sensitive nonradioactive method could ideally be used in patients’ first screening. However, in comparison with RIPA, CBA cannot provide an exact quantification of Ab titer, and its use is currently limited to selected laboratories with specific facilities and expertise.[16,17]

Clustered-AChR Abs A significant improvement in the serological diagnosis of MG was the demonstration of serum Abs to clustered AChR. Earlier reports had shown that a proportion of dSN patients had a clinical picture undistinguishable from AChR-MG [18,19] and often harbored thymus hyperplastic changes, that are definitely uncommon in MuSK-MG.[20–22] These observations suggested the possible presence of Abs unable to attach to AChR in solution, but able to bind to densely packed AChRs, as they are in vivo.[23] To test this hypothesis, the neuroscience group in Oxford developed a CBA, using human embryonic kidney 293 cells transfected with cDNA from AChR subunits and from rapsyn, in order to have, on cell surface,

Downloaded by [University of New England] at 09:05 11 November 2017

160

A. EVOLI ET AL.

AChR clusters similar to those at the NMJ. With this assay, they were able to detect AChR Abs in a high proportion of dSN patients, including some cases with purely ocular disease.[23–25] These Abs were found to be mostly complement-activating IgG1,[23] and proved pathogenic in a passive transfer model.[25] Recently, the same authors reported clustered-AChR Abs in 38% dSN-MG patients most frequently in patients with prepubertal age of onset and purely ocular symptoms.[26] These results, if confirmed, would strengthen the diagnostic value of clusteredAChR Abs, given their high positivity rate in patient subgroups, as ocular myasthenia and pediatric cases, in whom standard Ab assays have a lower diagnostic yield and diagnostic confirmation may be trying. Two recent reports [27,28] came to somewhat different conclusions. In a French study, clustered-AChR Abs were detected in 16% dSN-MG, mostly affected with mild disease.[27] In a Chinese study, patients with Abs only to clustered AChR accounted for 45.8% dSN cases, with a 27% rate of ocular myasthenia and a strong association with thymoma.[28] These discrepancies may be due to technical differences, possible biases in patients’ selection or referral, and ethnic factors, as already noted for MuSK Abs. CBA may improve MuSK Ab detection in RIPA-negative patients.[24,29] In a multicenter study, MuSK Abs were detected in 13% of ‘triple-negative’ patients, in 1.9% of controls and in 5.1% of patients with other neurologic autoimmune diseases; interestingly, subjects with MuSK Abs only on CBA had a milder phenotype that those positive on standard RIPA.[29]

Low-density lipoprotein receptor-related protein 4Abs Of late, clinical research has made significant progress in the search for new pathogenic Abs in MG. Lrp4 represented an ideal candidate antigen since its identification with the long-sought coreceptor for agrin, in 2008.[30] In the subsequent years, while the crucial role of Lrp4 at the NMJ was further clarified in experimental studies,[6,31] several groups reported serum IgG to Lrp4 in MG patients. Lrp4 Abs mainly belong to IgG1 subclass and were shown to interfere with Lrp4-agrin binding [32] and with agrin-induced AChR clusters.[33] Mice immunized with Lrp4 ectodomain developed MG-like weakness associated with electrophysiological signs of NMT impairment.[34] Similar alterations were induced by injection of Abs from Lrp4-immunized rabbits.[34] Passive-transfer studies by patients’ IgG injection have not been reported so far.

The proportion of Lrp4-positive patients among dSN cases varied from 0 to 50% in different reports. [7,24,32,33,35,36] This variability could be due to the use of different assays, such as luciferase-reporter immunoprecipitation,[32] ELISA [33] and CBA.[7,24,35,36] Furthermore, a multicenter study, in which Lrp4 Abs were detected in 18.7% of 635 dSN patients, showed differences in Ab frequency (from 7 to 37%) in samples from different countries,[7] suggesting a possible role of ethnic factors in disease susceptibility. In the same study, more than 20% of Lrp4-positive patients had ocular myasthenia, and most of those with generalized disease had mild to moderate symptoms. In some patients, Lrp4 Abs were found to be associated, with AChR or MuSK Abs, more commonly with the latter [7,32,35,36]; these double positive cases tended to have more severe symptoms.[7] Recently, Lrp4 Abs were detected in serum and cerebrospinal fluid samples from patients with ALS. In this study, Lrp4 positivity rate was much higher in ALS (23%) than in MG (1–2%).[37] These findings are worrying, as they cut down Lrp4-Ab specificity for MG, and may even complicate clinical diagnosis. ALS with bulbar onset is sometimes mistaken for MG, and, in these cases, a slight reaction to AChE-Is may be seen and a decremental response on RNS and increased jitter on SF-EMG can be present.[9] Ab-associated MG clinical patterns are summarized in Table 1. In the immunological diagnosis of MG, current knowledge confirms RIPA as first-line assay in the detection of AChR and MuSK Abs and in Ab status monitoring. The diagnostic yield of these Abs can be increased by the use of sensitive CBAs. Clustered-AChR Ab assay proved an effective and specific tool in MG diagnosis; larger size studies including samples from different countries could be helpful in clarifying its epidemiological and clinical association. Lrp4 confirms the third most frequent Ab in MG patients; standardization of assay techniques, further evaluation of Ab specificity and Ab-phenotype correlation will better define its clinical relevance.

Other Abs Abs to agrin, ColQ and cortactin have recently been reported in MG patients. Their role in the disease pathogenesis has not been proved and they are not currently used in diagnostic assays. Neuronal agrin is a large proteoglycan secreted by motor neurons and enriched at the basal lamina. AgrinLrp4 binding promotes the formation of a tetrameric complex that binds to and activates MuSK, triggering intracellular signaling leading to AChR clustering.[4] Abs to agrin were detected by ELISA and CBA, in a total of

EXPERT REVIEW OF CLINICAL IMMUNOLOGY

161

Table 1. Clinical characteristics of MG subtypes. AChR-MG (85% patients)† Age of onset Males:Females Ocular myasthenia Weakness pattern Thymus changes (prevalent)

Early-onset 20% Any Hyperplastic changes in some cases

†

An additional 3–4% of MG patients should have clustered-AChR Abs. According to WHO classification. AChR: acetylcholine receptor; Lrp4: low-density lipoprotein receptor-related protein 4; MG: myasthenia gravis; MuSK: muscle-specific tyrosine kinase receptor.

Downloaded by [University of New England] at 09:05 11 November 2017

‡

34 MG patients in different studies, often in association with other disease-specific Abs, mostly AChR Abs.[38– 41] Sera from agrin-positive patients inhibited agrininduced MuSK phosphorylation and AChR clustering in myotubes.[38] At the NMJ, AChE is anchored at the basal lamina by its triple collagen tail (ColQ). Abs to ColQ were detected in 12/415 (2.9%) MG sera and in 1/43 (2.3%) control samples. All ColQ-positive patients were women and were prevalently affected with generalized disease; 7/ 12 of these cases were also positive for AChR-Abs or MuSK-Abs.[42] Cortactin is a tyrosine kinase substrate and a regulator of F-actin assembly that acts downstream of agrin/ MuSK in promoting AChR clustering.[43] Abs to cortactin were reported in 19.7% of dSN patients, but also in 4.8% of AChR- or MuSK-positive patients, 12.5% of subjects with other autoimmune diseases and 5.2% of healthy controls.[44]

Additional testing Patients with AChR-MG may have serum Abs to titin and the ryanodine receptor (striatonal Abs). These Abs are not diagnostic of MG, but are strongly associated with thymoma (titin Abs are positive in 95% and ryanodine receptor Abs in 70% of thymoma patients) and to a lesser extent with late-onset nonthymoma MG.[8,44] Striatonal Abs are markers of thymoma, at least in young MG patients,[45] though binding intracellular antigens and their pathogenicity remain uncertain. Kv1.4 Abs that target the muscle voltage-gated potassium channel were reported in 12–15% of Japanese MG patients, often in association with severe MG and myocarditis with arrhythmias.[46] These findings were not confirmed in Caucasian patients.[47] Upon MG diagnosis, all patients should undergo a radiological study of the mediastinum by computed tomography or MRI. As the presence of a thymoma is

uncommon in ocular myasthenia, and very rare in children and in patients with MuSK or Lrp4 Abs, its exclusion should be especially careful in patients at higher risk for such an association, that is, adult subjects with generalized AChR-MG.

Treating MG There is general agreement that MG is one of the more treatable neuromuscular diseases. Current treatment, although largely unspecific, has dramatically reduced mortality and can restore satisfactory life conditions in the great majority of patients. However, there are still relevant drawbacks and controversies that can complicate management even in specialized centers: MG is a chronic condition and many patients need long-term treatment with exposure to serious side effects; few therapeutic modalities have been evaluated in randomized controlled trials (RCTs) and current guidelines mainly rely on expert opinions and retrospective studies; the response to treatment can differ among patients depending on MG biological diversity. Most patients present with isolated eyelid ptosis and diplopia, but only in 15–20% of these cases,[48] symptoms remain confined to extrinsic ocular muscles; in the other patients, generally within 2 years from the onset, weakness spreads to other muscle groups. The outcome of generalized MG is largely unpredictable, as some patients may suffer from rapidly progressive symptoms with early respiratory crises, while others have mild weakness for the whole course of their disease. Validated biomarkers that may predict disease severity and response to therapy have not yet been identified. Clinical management is based on the use, usually in combination, of different therapeutic options. Intensity of care is determined by weakness extension and severity, while treatment strategy should also take into account the disease pathophysiology (associated Abs

162

A. EVOLI ET AL.

and thymus pathology), and should be tailored, as far as possible, on individual patients.

Downloaded by [University of New England] at 09:05 11 November 2017

Symptomatic treatment Symptomatic agents relieve MG symptoms by enhancing NMT. They do not affect Ab production and pathogenic effects, and do not influence the natural history of the disease. Oral AChE-Is (pyridostigmine is the agent most commonly used) are the first-line treatment in MG, although normal muscle strength can be restored only in subjects with mild symptoms. Most AChR-MG patients respond more or less to pyridostigmine that is well tolerated at standard dosage ranging 180–360 mg/day in adults. Side effects are generally mild and subside with dose reduction. AChEI overmedication can lead to depolarization blockade at NMJ with increased muscle weakness.[48] On the other hand, MuSK-MG patients, for the most part, do not improve on AChE-Is, and standard pyridostigmine doses frequently induce nicotinic side effects and may even worsen weakness up to a cholinergic crisis.[11] A relative deficiency of AChE at motor endplate due to Abs interference with MuSK-ColQ binding could account for the cholinergic hypersensitivity in MuSK-MG.[49] In recent passive transfer studies, pyridostigmine injection exacerbated morphological and functional end-plate alterations, while 3,4-diaminopyridine, which increases quantal release without prolonging ACh half-life at the synaptic cleft, enhanced NMT without causing postsynaptic alterations.[50] In addition, treatment with the β2-adrenergic agent, albuterol reduced weakness in mice injected with IgG from MuSK-MG patients.[51] There are very few data on the effects of these agents in the human disease. To date, a partial effect of 3,4-dyaminopiridine in two children,[52] and response to both ephedrine and albuterol in a patient with severe MuSK-MG,[53] have been reported.

Long-term immunomodulation: thymectomy Thymectomy is indicated in all MG patients with evidence of thymoma. In these cases, surgery is the mainstay of oncologic treatment (that includes radio- and chemotherapy for invasive tumors), while it does not appear to improve significantly MG course. In patients without thymoma, thymectomy is aimed at the treatment of MG (‘therapeutic thymectomy’), its rationale resting on the pathogenic role of the thymus that has been convincingly shown in AChR-MG.[3,54,55] The efficacy of therapeutic thymectomy has never been evaluated in RCTs, and confounding factors, as

interstudy variability in surgical approach, patients’ selection, associated therapies and results’ evaluation, make the comparison between retrospective reports difficult.[56–58] Currently, thymectomy is recommended, in nonthymomatous patients with generalized MG, as an option to increase the chance of improvement and remission.[56] Recent studies support this view, reporting a better outcome in thymectomized than in unthymectomized patients, in particular in subjects with early onset MG associated with TFH [58,59] and AChR Abs.[22,60] The results of a multicenter, single-blinded randomized trial in nonthymomatous AChR-MG patients are awaited soon.[61] The indication to thymectomy should be extended to generalized MG with clustered-AChR Abs, as in these patients, the disease pathogenesis is very likely to be the same as in ‘typical’ AChR-MG and both TFH and thymoma have been reported.[23,27,28] On the other hand, the role of thymectomy in patients with other Abs is highly controversial. In MuSK-MG, thymus hyperplastic changes are uncommon [3,20,21] and clinical studies failed to show a definite benefit from thymectomy, as most thymectomized patients remained dependent on immunosuppressive therapy and few achieved remission.[62,63] In Lrp4-MG, TFH appears to be more rare than in AChR-MG [8]; the response to thymectomy has not been specifically addressed. The presence of a thymoma has been reported so far in three MuSK-MG patients [11,64] and in a single case with Lrp4 Abs.[36] This association, rare as it may be, justifies a radiological study of the mediastinum in these patients.

Short-term immunomodulation: plasma-exchange and intravenous immunoglobulin Plasma-exchange (PLEX), that removes Abs from circulation, and intravenous immunoglobulin (IVIg) that interferes with Ab activity, have a rapid albeit shortlived effect in MG, and additional immunosuppression is usually needed. PLEX standard protocol consists of five exchanges of one plasma volume on alternate days; IVIg are usually administered at 2 g/kg over 2–5 days. Their main indication is the treatment of disease exacerbations in patients with severe symptoms or respiratory crises. Additional indications include prevention of MG deterioration at the start of steroid therapy, preparation for thymectomy and as periodic treatment in selected cases unresponsive to conventional immunosuppression.[54,55] Some years ago, PLEX and IVIg were shown to have comparable efficacy in the treatment of severe MG

Downloaded by [University of New England] at 09:05 11 November 2017

EXPERT REVIEW OF CLINICAL IMMUNOLOGY

exacerbations.[65] A recent RCT has reached the same conclusion in patients with moderate to severe disease, showing that the two treatments were similar for extent of clinical improvement, response rate, duration of effect and tolerance.[66] The choice between PLEX and IVIg is based on patient’s comorbidities, procedure’s potential risks and medical facilities. However, in patients with life-threatening symptoms, PLEX is preferred because of its more rapid effect, while for outpatients IVIg is more suitable.[67] Both AChR-MG and MuSK-MG respond very well to PLEX, while IVIg effectiveness appears to be less consistent in patients with MuSK Abs.[11] Adverse effects can be reduced by screening patient’s comorbidities before treatment. IVIg may induce anaphylaxis in IgA-deficient patients and, at the dosage used in MG, is contraindicated in subjects with congestive heart disease and renal function impairment. Otherwise, side effects observed with IVIg are generally mild (nausea, headache, chills and fever) though, more rarely hemolytic anemia and thromboembolic complications can occur. PLEX is contraindicated in patients with sepsis, epilepsy, severe arterial hypotension and heart failure. Most adverse events are associated with vascular access such as infection, thrombosis; hemodynamic instability related to volume shifts requires careful monitoring during the procedure.[55] Different techniques of immunoadsorption (IA) have been successfully used in MG (mostly in AChR-MG), with the same indications as PLEX. This semiselective IA, that removes circulating IgG leaving other plasma components unaltered, can represent a valuable alternative in patients requiring intensive PLEX protocols.[68] An even more attractive option would be the selective depletion of pathogenic Abs. The efforts to develop antigen-specific IA have achieved encouraging results in experimental studies.[69] Subcutaneous IgG has recently been evaluated for maintenance therapy in inflammatory neuromuscular diseases.[70] Because of ease of administration and tolerability, it appears to be a sensible alternative to IVIg. An open label study of subcutaneous IgG in MG patients is under way.

Immunosuppressive therapy Immunosuppression is the mainstay treatment for MG. It is performed in patients with disabling symptoms, not adequately controlled with symptomatic agents, in association or as an alternative to thymectomy. Treatment approach is determined by weakness severity and rate of progression, and by patient’s characteristics (age, lifestyle and associated conditions). Once

163

sustained improvement is achieved, medications are gradually reduced to the minimum effective dose in order to minimize adverse effects.[48] Corticosteroids have been in use for many years, and treatment drawbacks and complications (including the risk of MG deterioration at the start of treatment) are well known.[71] Nonetheless, these agents still represent the first option in MG whereby immunosuppression is required on account of their efficacy (70–80% of response rate) and rapid-onset effect, that is advantageous in severely affected cases.[72] Prednisone and prednisolone are the preparations most commonly used, starting with high doses or with a progressively increasing regimen,[71] up to 1 mg/kg/day in patients with generalized MG; an alternate-day schedule is generally preferred in chronic administration.[54] High-dose steroids, in association with PLEX, are the first-choice treatment for respiratory crises, while lower doses can be sufficient in patients with mild or purely ocular symptoms.[73] Different classes of immunosuppressants, including cytostatic drugs (azathioprine, mycophenolate mofetil and methotrexate), the alkylating agent cyclophosphamide and calcineurin inhibitors (cyclosporine A and tacrolimus), are used in MG treatment. These agents can be used in monotherapy, but because of delayed-onset effect, they are frequently administered in association with prednisone (to reduce its dosage and treatment duration) and can replace steroids in long-term treatment. Positive clinical evidence, though from studies based on small sample size is available for azathioprine (as steroidsparing agent), cyclosporine A and iv. pulse cyclophosphamide.[55] On the other hand, three RCTs, two on mycophenolate mofetil [74,75] and one on tacrolimus [76] failed to demonstrate significant efficacy (in these trials, due to the study design, the investigational drug effect was probably masked by the concomitant steroid treatment). Clinical benefit of mycophenolate [77] and tacrolimus [78] was reported in retrospective, open label studies and small trials, and, also in view of their favorable side-effect profile, these agents are used as second- or third-line therapy. In many countries, azathioprine (2–3 mg/kg body weight (b.w.)/day as starting dose, 1 mg/kg as maintenance dose) is the first-choice immunosuppressant in MG patients. As patients with thiopurine methyl transferase deficiency may develop severe bone-marrow toxicity, thiopurine methyl transferase activity should be measured before treatment. Mycophenolate mofetil (at a standard dose of 2–2.5 g/daily) is generally used as second-choice; cyclosporine A (at an initial dose of 4–6 mg/kg b.w. and a maintenance dose 80% patients with generalized AChR-MG and in nearly all MuSK-MG cases, with an overall response rate around 90% and an outcome of minimal manifestations or better [80] in 66% of patients.[72] However, disease relapses are frequent on dose tapering or after treatment withdrawal, and most patients need long-term therapy. Patient candidates to immunosuppression should be screened for chronic infections like viral hepatitis and tuberculosis. Bone mineral density assessment and calcium, vitamin D and bisphosphonate supplement should be started concurrently with corticosteroids to prevent osteoporosis, and other acute and long-term steroid complications should be promptly diagnosed and treated. In patients taking immunosuppressants, side-effect monitoring through specific and periodic blood tests is necessary. As patients’ cooperation is of the utmost importance, patients and relatives must be fully informed of potential risks and expected benefit.

Refractory MG MG is defined as refractory to conventional treatment, when patients complain of disabling weakness or severe disease relapses in spite of adequate immunosuppressive treatment, and/or they require too high doses of steroids or other immunosuppressive agents with serious side effects.[81] The frequency of refractory MG was found to be around 10%, with the highest prevalence rate among patients with thymoma-associated AChR-MG and MuSK-MG.[82] These findings are not surprising, considering that these patient groups are generally affected with severe disease and tend to remain dependent on immunosuppressive therapy.[72] The reasons why some patients, at some point of their disease, become resistant to treatment and exactly when the autoimmune response goes out of control are largely unknown. Multiple factors are likely to be involved, including failure of regulatory signals, dysfunction of B cell maturation and survival, generation of long-lived autoreactive plasma cells, changes in Ab subclass and epitope specificity.[83–86]

Refractory MG can been managed with intermittent PLEX/IA or IVIg treatments, in association with immunosuppressive therapy.[55] This option provides shortterm benefit, has a significant impact on patient’s lifestyle, is costly and not exempt from side effects. Intravenous cyclophosphamide, both on high-doses (200 mg/kg in 4 consecutive days) [81] and as pulse treatment (at 0.75 mg/m2 every 4 weeks for 6 cycles) followed by conventional immunotherapy,[87] resulted in significant improvement in all forms of MG. However, its use is limited by relevant toxicity. The use of biologics as monoclonal Abs (-mAbs) and therapeutic fusion proteins (-cepts) is rapidly expanding in the treatment of autoimmune diseases. So far, B cell depletion and complement inhibition have proved beneficial in patients with refractory MG. Rituximab, a chimeric anti-CD20 mAb, causes a profound depletion of circulating pre-B and B lymphocytes without reducing total IgG levels. To date, 170 MG patients treated with rituximab have been reported in the English literature. The results of a recent meta-analysis showed a higher response rate in MuSK-MG (88.8%) than in AChR-MG (80.4%),[88] thus confirming previous reports,[89,90] together with a trend toward an inverse correlation between disease duration and response to rituximab; treatment was generally well tolerated.[88] Rituximab efficacy and tolerability in MG have been evaluated, so far, in single-case reports and small openlabel studies, characterized by great variability in treatment schedule, result evaluation criteria and follow-up duration. A phase-II RCT is currently under way. Decrease in specific Ab levels [89] and reconstitution of T and B regulatory cells have been proposed as biomarkers of response to rituximab.[90,91] Eculizumab, a humanized mAb, inhibits the Complement component 5 preventing the formation of the terminal C cascade. It proved effective and well tolerated in a small Phase II RCT in AChR-MG patients. [92] A phase III RCT is ongoing. Cyclophosphamide and rituximab, as well as conventional immunosuppressants, do not deplete long-lived plasma cells, that, having entered the immunological niche, can persistently secrete Abs. Different agents targeting plasma cells and niche survival factors, like chemokines and their receptors, cytokines such as interleukin-6, B-cell activating factor (BAFF) and a proliferation-inducing ligand (APRIL), are being evaluated in the treatment of autoimmune diseases.[84] Currently, belimumab, a human anti-BAFF mAb, and the proteasome inhibitor bortezomib that eliminates both short-lived and long-lived plasma cells are the object of RCTs in MG. Improving our understanding of mechanisms involved in treatment resistance and the availability of

EXPERT REVIEW OF CLINICAL IMMUNOLOGY

biomarkers predictive of MG flares could optimize the use of disease-modifying medications.

Downloaded by [University of New England] at 09:05 11 November 2017

Expert commentary and five-year view Recently, the effects of MuSK Abs have been defined in experimental studies and new Ab targets have been identified in MG patients. These developments have expanded our understanding of the pathophysiology of the disease and improved its management. However, new Ab assays require specific facilities and expertise, and are currently performed in few laboratories. In clinical practice, the diagnosis of MG can be challenging in patients with negative results on AChR and MuSK Ab standard assays. In these cases, diagnostic confirmation requires the exclusion of conditions that can mimic MG and should rely more on the clinical context than on positive results on a single test. Current treatment is mostly based on unspecific immunosuppression and has considerable limitations. RCTs are difficult in a rare disease like MG, but recently completed trials have provided evidence for therapeutic approaches that had long been in clinical use. Complement inhibition and B-cell targeting therapies seem to be promising approaches as rescue treatment for refractory disease. A better understanding of the mechanisms leading to nonresponsiveness to immunosuppressants may justify a more extensive and earlier use of biologic agents. However, growing expenditures on extensive use of off-label treatments is already a cause for concern in payers and patients, and a major challenge in the next future will be how to grant effective and safe therapies at sustainable costs. Biomarkers that can predict the disease course and response to treatment have long been sought after in

165

MG. Possible candidates, as subsets of regulatory cells, cytokines and microRNAs are currently under investigation.[66,93] Epigenetics and exosomes are promising research fields for biomarker discovery.[94,95] In particular, the proteomic and microRNA profiling of thymus- or muscle-derived exosomes may provide useful insights for the monitoring of MG course. Epitope mapping can elucidate the pathogenic effects of Abs [86] and, together with changes in IgG subclasses, may correlate more closely with clinical changes that total Ab titer. Possible candidate targets for new therapies are T follicular helper cells, a subset of T lymphocytes that plays a key role in B cell maturation. Compelling evidence exists in mice and in humans that aberrant generation and/or activation of T follicular helper cells may contribute to the pathogenesis of autoimmune diseases.[85] MAbs targeting molecules expressed by this lymphocytes subset (e.g. ICOS) are currently evaluated in the treatment of systemic lupus erythematosus.[96] Different forms of antigen-specific immunotherapy [54,97] have been successfully developed in animal models, and, the next years will see more efforts to fill the gap between experimental studies and human disease.

Financial & competing interests disclosure The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

Key issues ● Acetylcholine receptor (AChR) antibodies (Abs) are the first to be tested when myasthenia gravis (MG) is ● ● ●

● ● ●

suspected. Muscle-specific tyrosine kinase receptor (MuSK) Abs should be tested in all AChR negative patients. MuSKMG is more likely in young adult women with predominant weakness of bulbar and neck muscles. Clustered-AChR Abs seem to be more common in young patients (including children) with ocular or mild generalized disease. Further studies are needed to assess the specificity of low-density lipoprotein receptor-related protein 4 Abs for MG. These Abs should be assayed in AChR and MuSK negative patients. The associated phenotype appears to be similar to AChR-MG. The frequency of triple-negative (AChR/MuSK/low-density lipoprotein receptor-related protein 4 negative) MG could be estimated at 3–4%. Most MuSK-MG patients are unresponsive to or intolerant of acetylcholinesterase inhibitors. 3,4-diaminopyridine and albuterol might be beneficial in these cases. In uncontrolled studies, rituximab proved effective in MG, especially in MuSK-MG.

166

A. EVOLI ET AL.

Downloaded by [University of New England] at 09:05 11 November 2017

References Papers of special note have been highlighted as: * of interest ** of considerable interest 1. Keesey JC. Clinical evaluation and management of myasthenia gravis. Muscle Nerve. 2004;29:484–505. 2. Gomez AM, Van Den Broeck J, Vrolix K, et al. Antibody effector mechanisms in myasthenia gravis pathogenesis at the neuromuscular junction. Autoimmunity. 2010;43:353–370. 3. Marx A, Pfister F, Schalke B, et al. The different roles of the thymus in the pathogenesis of the various myasthenia gravis subtypes. Autoimmun Rev. 2013;12:875–884. 4. Burden SJ, Yumoto N, Zhang W. The role of MuSK in synapse formation and neuromuscular disease. Cold Spring Harb Perspect Biol. 2013;5:a009167. 5. Huijbers MG, Zhang W, Klooster R, et al. MuSK IgG4 autoantibodies cause myasthenia gravis by inhibiting binding between MuSK and Lrp4. Proc Natl Acad Sci USA. 2013;110:20783–20788. ** The authors demonstrate that human IgG4 musclespecific tyrosine kinase receptor (MuSK) antibodies bind to the first Ig-like domain in MuSK, thus disrupting MuSK–low-density lipoprotein receptor-related protein 4 interaction and inhibiting agrin-stimulated MuSK phosphorylation. 6. Zhang W, Coldefy AS, Hubbard SR, et al. Agrin binds to the N-terminal region of Lrp4 protein and stimulates association between Lrp4 and the first immunoglobulin-like domain in muscle-specific kinase (MuSK). J Biol Chem. 2011;286:40624–40630. 7. Zisimopoulou P, Evangelakou P, Tzartos J, et al. A comprehensive analysis of the epidemiology and clinical characteristics of anti-LRP4 in myasthenia gravis. J Autoimmun. 2014;52:139–145. * A multinational survey of acetylcholine receptor/muscle-specific tyrosine kinase receptor negative patients reporting an overall low-density lipoprotein receptorrelated protein 4 antibody frequency of 18.7% with differences among populations. Both clinical presentation and response to therapy in low-density lipoprotein receptor-related protein 4-myasthenia gravis (MG) patients were more similar to those in acetylcholine receptor-MG than in MuSK-MG. 8. Meriggioli MN, Sanders DB. Muscle autoantibodies in myasthenia gravis: beyond diagnosis?. Expert Rev Clin Immunol. 2012;8:427–438. * A comprehensive review of antibodies to muscle antigens in myasthenia gravis. The authors thoroughly discuss the relevance of different antibodies in diagnosis and as measure of disease severity. 9 Howard JF Jr. Electrodiagnosis of disorders of neuromuscular transmission. Phys Med Rehabil Clin N Am. 2013;24:169–192. 10. Benatar M. A systematic review of diagnostic studies in myasthenia gravis. Neuromusc Disord. 2006;16:459–467. 11. Evoli A, Padua L. Diagnosis and therapy of myasthenia gravis with antibodies to muscle-specific kinase. Autoimmun Rev. 2013;12:931–935.

12. Oger J, Frykman H. An update on laboratory diagnosis in myasthenia gravis. Clinica Chimica Acta. 2015;444:126–131. 13. Chan KH, Lachance DH, Harper CM, et al. Frequency of seronegativity in adult-acquired generalized myasthenia gravis. Muscle Nerve. 2007;36:651–658. 14. Berrih-Aknin S, Frenkian-Cuvelier M, Eymard B. Diagnostic and clinical classification of autoimmune myasthenia gravis. J Autoimmun. 2014;48-49:143–148. 15. Chiang LM, Darras BT, Kang PB. Juvenile myasthenia gravis. Muscle Nerve. 2009;39:423–431. 16. Zisimopoulou P, Brenner T, Trakas N, et al. Serological diagnostics in myasthenia gravis based on novel assays and recently identified antigens. Autoimmun Rev. 2013;12:924–930. 17. Rodriguez Cruz PM, Huda S, López-Ruiz P, et al. Use of cellbased assays in myasthenia gravis and other antibodymediated diseases. Exp Neurol. 2015;270:66–71. 18. Evoli A, Tonali PA, Padua L, et al. Clinical correlates with anti-MuSK antibodies in generalized seronegative myasthenia gravis. Brain. 2003;126:2304–2311. 19. Deymeer F, Gungor-Tuncer O, Yilmaz V, et al. Clinical comparison of anti-MuSK- vs anti-AChR-positive and seronegative myasthenia gravis. Neurology. 2007;68:609–611. 20. Lauriola L, Ranelletti F, Maggiano N, et al. Thymus changes in anti-MuSK-positive and -negative myasthenia gravis. Neurology. 2005;64:536–538. 21. Leite MI, Ströbel P, Jones M, et al. Fewer thymic changes in MuSK antibody-positive than in MuSK antibody-negative MG. Ann Neurol. 2005;57:444–448. 22. Nikolic A, Djukic P, Basta I, et al. The predictive value of the presence of different antibodies and thymus pathology to the clinical outcome in patients with generalized myasthenia gravis. Clin Neurol Neurosurg. 2013;115:432–437. 23. Leite MI, Jacob S, Viegas S, et al. IgG1 antibodies to acetylcholine receptors in ‘seronegative’ myasthenia gravis. Brain. 2008;131:1940–1952. 24. Vincent A, Waters P, Leite MI, et al. Antibodies identified by cell-based assays in myasthenia gravis and associated diseases. Ann N Y Acad Sci. 2012;1274:92–98. 25. Jacob S, Viegas S, Leite MI, et al. Presence and pathogenic relevance of antibodies to clustered acetylcholine receptor in ocular and generalized myasthenia gravis. Arch Neurol. 2012;69:994–1001. 26. Rodríguez Cruz PM, Al-Hajjar M, Huda S, et al. Clinical features and diagnostic usefulness of antibodies to clustered acetylcholine receptors in the diagnosis of seronegative myasthenia gravis. JAMA Neurol. 2015;72:642–649. * This study reports the clinical correlates of clusteredacetylcholine receptor antibodies, showing a high rate of positive results in patients with childhood or ocular myasthenia gravis. 27. Devic P, Petiot P, Simonet T, et al. Antibodies to clustered acetylcholine receptors: expanding the phenotype. Eur J Neurol. 2014;21:130–134.

Downloaded by [University of New England] at 09:05 11 November 2017

EXPERT REVIEW OF CLINICAL IMMUNOLOGY

28. Zhao G, Wang X, Yu X, et al. Clinical application of clustered-AChR for the detection of SNMG. Sci Rep. 2015;5:10193. 29. Tsonis AI, Zisimopoulou P, Lazaridis K, et al. MuSK autoantibodies in myasthenia gravis detected by cell based assay–A multinational study. J Neuroimmunol. 2015;284:10–17. 30. Kim N, Stiegler AL, Cameron TO, et al. Lrp4 is a receptor for agrin and forms a complex with MuSK. Cell. 2008;135:334–342. 31. Yumoto N, Kim N, Burden SJ. Lrp4 is a retrograde signal for presynaptic differentiation at neuromuscular synapses. Nature. 2012;489:438–442. 32. Higuchi O, Hamuro J, Motomura M, et al. Autoantibodies to low-density lipoprotein receptor-related protein 4 in myasthenia gravis. Ann Neurol. 2011;69:418–422. 33. Zhang B, Tzartos JS, Belimezi M, et al. Autoantibodies to lipoprotein-related protein 4 in patients with doubleseronegative myasthenia gravis. Arch Neurol. 2012;69:445–451. 34. Shen C, Lu Y, Zhang B, et al. Antibodies against lowdensity lipoprotein receptor-related protein 4 induce myasthenia gravis. J Clin Invest. 2013;123:5190–5202. 35. Pevzner A, Schoser B, Peters K, et al. Anti-LRP4 autoantibodies in AChR- and MuSK-antibody-negative myasthenia gravis. J Neurol. 2012;259:427–435. 36. Marino M, Scuderi F, Samengo D, et al. Flow cytofluorimetric analysis of anti-LRP4 (LDL receptor-related protein 4) autoantibodies in Italian patients with myasthenia gravis. PLoS One. 2015;10:e0135378. 37. Tzartos JS, Zisimopoulou P, Rentzos M, et al. LRP4 antibodies in serum and CSF from amyotrophic lateral sclerosis patients. Ann Clin Transl Neurol. 2014;1:80–87. 38. Zhang B, Shen C, Bealmear B, et al. Autoantibodies to agrin in myasthenia gravis patients. PLoS One. 2014;9: e91816. 39. Gasperi C, Melms A, Schoser B, et al. Anti-agrin autoantibodies in myasthenia gravis. Neurology. 2014;82:1976–1983. 40. Cossin J, Belaya K, Zoltowska K, et al. The search for new antigens in myasthenia gravis. Ann N Y Acad Sci. 2012;1275:123–128. 41. Zoltowska Katarzyna M, Belaya K, Leite M, et al. Collagen Q - a potential target for autoantibodies in myasthenia gravis. J Neurol Sci. 2015;348:241–244. 42. Madhavan R, Gong ZL, Ma JJ, et al. The function of cortactin in the clustering of acetylcholine receptors at the vertebrate neuromuscular junction. PLos One. 2009;4: e8478. 43. Gallardo E, Martínez-Hernández E, Titulaer MJ, et al. Cortactin autoantibodies in myasthenia gravis. Autoimmun Rev. 2014;13:1003–1007. 44. Skeie GO, Aarli JA, Gilhus NE. Titin and ryanodine receptor antibodies in myasthenia gravis. Acta Neurol Scand. 2006;113(S183):19–23. 45. Choi Decroos E, Hobson-Webb LD, Juel VC, et al. Do acetylcholine receptor and striated muscle antibodies predict the presence of thymoma in patients with myasthenia gravis?. Muscle Nerve. 2014;49:30–34. 46. Suzuki S, Baba A, Kaida K, et al. Cardiac involvements in myasthenia gravis associated with anti-Kv1.4 antibodies. Eur J Neurol. 2014;21:223–230.

167

47. Romi F, Suzuki S, Suzuki N, et al Anti-voltage-gated potassium channel Kv1.4 antibodies in myasthenia gravis. J Neurol. 2011;259:1312–6. 48. Meriggioli MN, Sanders DB. Autoimmune myasthenia gravis: emerging clinical and biological heterogeneity. Lancet Neurol. 2009;8:475–490. 49. Mori S, Kubo S, Akiyoshi T, et al. Antibodies against muscle-specific kinase impair both presynaptic and postsynaptic functions in a murine model of myasthenia gravis. Am J Pathol. 2012;180:798–810. 50. Morsch M, Reddel SW, Ghazanfari N, et al. Pyridostigmine but not 3,4-diaminopyridine exacerbates ACh receptor loss and myasthenia induced in mice by muscle-specific kinase antibody. J Physiol. 2013;591:2747–2762. ** In this passive transfer study, pyridostigmine exacerbated the structural and functional alterations induced by muscle-specific tyrosine kinase receptor antibodies at motor endplate. In contrast, 3-4 diaminopyridine enhanced neuromuscular transmission. 51. Ghazanfari N, Morsch M, Tee N, et al. Effects of the β2adrenoreceptor agonist, albuterol, in a mouse model of anti-MuSK myasthenia gravis. PLoS ONE. 2014;9:e87840. 52. Skjei KL, Lennon VA, Kuntz NL. Muscle specific kinase autoimmune myasthenia gravis in children: a case series. Neuromuscul Disord. 2013;23:874–882. 53. Haran M, Schattner A, Mate A, et al. Can a rare form of myasthenia gravis shed additional light on disease mechanisms?. Clin Neurol Neurosurg. 2013;115:562–566. 54. Mantegazza R, Bonanno S, Camera G, et al. Current and emerging therapies for the treatment of myasthenia gravis. Neuropsychiatr Dis Treat. 2011;7:151–160. 55. Sathasivam S. Current and emerging treatments for the management of myasthenia gravis. Ther Clin Risk Manag. 2011;7:313–323. 56. Gronseth GS, Barohn RJ. Practice parameter: thymectomy for autoimmune myasthenia gravis (an evidence-based review). Neurology. 2000;55:5–15. 57. Sonett JR, Jaretzki AIII. Thymectomy for nonthymomatous myasthenia gravis. Ann N Y Acad Sci. 2008;1132:315–328. 58. Dìaz A, Black E, Dunning J. Is thymectomy in non-thymomatous myasthenia gravis of any benefit?. Interact Cardiovasc Thorac Surg. 2014;18:381–389. 59. Barnett C, Katzberg HD, Keshavjee S, et al. Thymectomy for non-thymomatous myasthenia gravis: a propensity score matched study. Orphanet J Rare Dis. 2014;9:214. 60. Baggi F, Andreetta F, Maggi L, et al. Complete stable remission and autoantibody specificity in myasthenia gravis. Neurology. 2013;80:188–195. 61. Newsom-Davis J, Cutter G, Wolfe GI, et al. Status of the thymectomy trial for nonthymomatous myasthenia gravis patients receiving prednisone. Ann N Y Acad Sci. 2008;1132:344–347. 62. Reddel SW, Morsch M, Phillips WD. Clinical and scientific aspects of muscle-specific tyrosine kinase-related myasthenia gravis. Curr Opin Neurol. 2014;27:558–565. 63. El-Salem K, Yassin A, Al-Hayk K, et al. Treatment of MuSKassociated myasthenia gravis. Curr Treat Options Neurol. 2014;16:283. 64. Ito A, Sasaki R, Ii Y, et al. A case of thymoma-associated myasthenia gravis with anti-MuSK antibodies. Rinsho Shinkeigaku. 2013;53:372–375.

Downloaded by [University of New England] at 09:05 11 November 2017

168

A. EVOLI ET AL.

65. Gajdos P, Chevret S, Clair B, et al Clinical trial of plasma exchange and high-dose intravenous immunoglobulins in myasthenia gravis. Ann Neurol. 1997;41:789–796. 66. Barth D, Nabavi Nouri M, Ng E, et al. Comparison of IVIg and PLEX in patients with myasthenia gravis. Neurology. 2011;76:2017–2023. * This study provides Class I evidence that intravenous immunoglobulin and plasma-exchange have comparable efficacy in patients with moderate to severe myasthenia gravis. 67. Dhawan PS, Goodman BP, Harper CM, et al. IVIG versus PLEX in the treatment of worsening myasthenia gravis: what is the evidence? A critically appraised topic. Neurologist. 2015;19:145–148. 68. Antozzi C. Immunoadsorption in patients with autoimmune ion channel disorders of the peripheral nervous system. Atheroscler Suppl. 2013;14:219–222. 69. Lazaridis K, Evaggelakou P, Bentenidi E, et al. Specific adsorbents for myasthenia gravis autoantibodies using mutants of the muscle nicotinic acetylcholine receptor extracellular domains. J Neuroimmunol. 2015;278:19–25. 70. Berger M. Subcutaneous IgG in neurologic diseases. Immunotherapy. 2014;6:71–83. 71. Jani-Acsadi A, Lisak RP. Myasthenia gravis. Curr Treat Options Neurol. 2010;12:231–243. 72. Sanders DB, Evoli A. Immunosuppressive therapies in myasthenia gravis. Autoimmunity. 2010;43(5–6):428–435. 73. Kerty E, Elsais A, Argov Z, et al. EFNS/ENS guidelines for the treatment of ocular myasthenia. Eur J Neurol. 2014;21:687–693. 74. Muscle Study Group. A trial of mycophenolate mofetil with prednisone as initial immunotherapy in myasthenia gravis. Neurology. 2008;71:394–399. 75. Sanders DB, Hart IK, Mantegazza R, et al. An international, phase III, randomized trial of mycophenolate mofetil in myasthenia gravis. Neurology. 2008;71:400–406. 76. Yoshikawa H, Kiuchi T, Saida T, et al. Randomised, double-blind, placebo-controlled study of tacrolimus in myasthenia gravis. J Neurol Neurosurg Psychiatry. 2011;82:970–977. 77. Burns TM, Sanders DB, Kaminski HJ, et al. Two steps forward, one step back: mycophenolate mofetil treatment for myasthenia gravis in the United States. Muscle Nerve. 2015;51:635–637. 78. Cruz JL, Wolff ML, Vanderman AJ, et al. The emerging role of tacrolimus in myasthenia gravis. Ther Adv Neurol Disord. 2015;8:92–103. 79. Heckmann JM, Rawoot A, Bateman K, et al. A singleblinded trial of methotrexate versus azathioprine as steroid-sparing agents in generalized myasthenia gravis. BMC Neurol. 2011;11:97. 80. Jaretzki A 3rd, Barohn RJ, Ernstoff RM, et al. Myasthenia gravis: recommendations for clinical research standards. Task Force of the Medical Scientific Advisory Board of the Myasthenia Gravis Foundation of America. Neurology. 2000;55:16–23. 81. Drachman DB, Adams RN, Hu R, et al. Rebooting the immune system with high-dose cyclophosphamide for treatment of refractory myasthenia gravis. Ann N Y Acad Sci. 2008;1132:305–314.

82. Suh J, Goldstein JM, Nowak RJ. Clinical characteristics of refractory myasthenia gravis patients. Yale J Biol Med. 2013;86:255–260. 83. Berrih-Aknin S, Ragheb S, Le Panse R, et al. Ectopic germinal centers, BAFF and anti-B-cell therapy in myasthenia gravis. Autoimmun Rev. 2013;12:885–893. 84. Winter O, Dame C, Jundt F, et al. Pathogenic long-lived plasma cells and their survival niches in autoimmunity, malignancy, and allergy. J Immunol. 2012;189:5105–5111. * This review describes how long-lived plasma cells survive in a protected microenvironment and discusses the strategies for their specific depletion. 85. Ueno H, Banchereau J, Vinuesa CG. Pathophysiology of T follicular helper cells in humans and mice. Nat Immunol. 2015;16:142–152. 86. Huijbers MG, Lipka AF, Plomp JJ, et al. Pathogenic immune mechanisms at the neuromuscular synapse: the role of specific antibody-binding epitopes in myasthenia gravis. J Intern Med. 2014;275:12–26. 87. Buzzard KA, Meyer NJ, Hardy TA, et al. Induction intravenous cyclophosphamide followed by maintenance oral immunosuppression in refractory myasthenia gravis. Muscle Nerve. 2015;52:204–210. 88. Iorio R, Damato V, Alboini PE, et al. Efficacy and safety of rituximab for myasthenia gravis: a systematic review and meta-analysis. J Neurol. 2015;262:1115–1119. 89. Díaz-Manera J, Martínez-Hernández E, Querol L, et al. Long-lasting treatment effect of rituximab in MuSK myasthenia. Neurology. 2012;78:189–193. 90. Sun F, Ladha SS, Yang L, et al. Interleukin-10 producingB cells and their association with responsiveness to rituximab in myasthenia gravis. Muscle Nerve. 2014;49:487–494. * This study demonstrates responsiveness in myasthenia gravis patients was associated with a rapid repopulation of B regulatory cells. 91. Sfikakis PP, Souliotis VL, Fragiadaki KG, et al. Increased expression of the FoxP3 functional marker of regulatory T cells following B cell depletion with rituximab in patients with lupus nephritis. Clin Immunol. 2007;123:66–73. 92. Howard JF Jr, Barohn RJ, Cutter GR, et al. A randomized, double-blind, placebo-controlled phase II study of eculizumab in patients with refractory generalized myasthenia gravis. Muscle Nerve. 2013;48:76–84. 93. Berrih-Aknin S, Le Panse R. Myasthenia gravis: a comprehensive review of immune dysregulation and etiological mechanisms. J Autoimmun. 2014;52:90–100. 94. Ngalamika O, Zhang Y, Yin H, et al. Epigenetics, autoimmunity and hematologic malignancies: a comprehensive review. J Autoimmun. 2012;39:451–465. 95. Vlassov AV, Magdaleno S, Setterquist R, et al. Exosomes: current knowledge of their composition, biological functions, and diagnostic and therapeutic potentials. Biochim Biophys Acta. 2012;1820:940–948. 96. Thanou A, Merrill JT. Treatment of systemic lupus erythematosus: new therapeutic avenues and blind alleys. Nature Rev Rheumatol. 2014;10:23–34. 97. Luo J, Lindstrom J. Antigen-specific immunotherapeutic vaccine for experimental autoimmune myasthenia gravis. J Immunol. 2014;193:5044–5055.