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

Research

Original Investigation

Clinical Features and Diagnostic Usefulness of Antibodies to Clustered Acetylcholine Receptors in the Diagnosis of Seronegative Myasthenia Gravis Pedro M. Rodríguez Cruz, MD, MSc; Michal Al-Hajjar, MD; Saif Huda, MB, ChB; Leslie Jacobson, DPhil; Mark Woodhall, PhD; Sandeep Jayawant, FRCPH; Camilla Buckley, MRCP, DPhil; David Hilton-Jones, MD, FRCP; David Beeson, PhD; Angela Vincent, FMedSci, FRS; Maria Isabel Leite, MD, DPhil; Jacqueline Palace, DM Editorial 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 ⱕ .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 ⱕ .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.

JAMA Neurol. doi:10.1001/jamaneurol.2015.0203 Published online April 20, 2015.

Author Affiliations: Nuffield Department of Clinical Neurosciences, University of Oxford, John Radcliffe Hospital, Oxford, England (Rodríguez Cruz, Al-Hajjar, Huda, Jacobson, Woodhall, Buckley, Hilton-Jones, Vincent, Leite, Palace); Neurosciences Group, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, England (Rodríguez Cruz, Beeson); Department of Paediatric Neurology, Children’s Hospital, John Radcliffe Hospital, Oxford, England (Jayawant). Corresponding Author: Jacqueline Palace, DM, Nuffield Department of Clinical Neurosciences, University of Oxford, John Radcliffe Hospital, Level 3, West Wing, Headley Way, Headington OX3 9DU, England ([email protected]).

(Reprinted) E1

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Research Original Investigation

Antibodies to Clustered Acetylcholine Receptors and Myasthenia Gravis

M

yasthenia gravis (MG) is an antibody-mediated autoimmune disease of the neuromuscular junction. Approximately 80% of patients with generalized MG have autoantibodies against the muscle nicotinic acetylcholine receptor (AChR) measured by radioimmunoprecipitation assay (RIPA).1,2 The AChR antibodies are predominantly IgG1 and IgG3 subclasses and produce severe loss of AChRs by complementmediated damage to the postsynaptic membrane, receptor endocytosis, and occasionally direct AChR block.3 These patients are classically referred to as having seropositive MG. Other patients with generalized MG and 50% of those with ocular MG lack detectable AChR antibodies by RIPA.4 Autoantibodies to muscle-specific tyrosine kinase (MuSK) measured by RIPA have been reported in a variable proportion of patients with seronegative MG (SNMG), ranging from 0 to 70%.5-12 Autoantibodies to MuSK are mainly of the IgG4 subclass, which does not activate complement. They prevent the interaction of MuSK with lipoprotein receptor–related protein 4 (LRP4) and therefore inhibit agrin-dependent AChR clustering.13 MuSK autoantibodies identified patients with distinctive clinical features.14,15 Patients with MG lacking detectable AChR and MuSK antibodies by RIPA are referred to as having SNMG. A cell-based assay (CBA) was established for the improved detection of AChR antibodies in patients previously seronegative by RIPA.16 Cell-based assays also measure AChR antibodies detected by RIPA,16 although this is not performed routinely because of the high cost and time-consuming nature compared with the RIPA. The CBA involves expressing AChRs on the surface of a human embryonic kidney (HEK) cell and clustering by coexpression with the intracellular anchoring protein rapsyn. This is performed by transfecting the live HEK cells with the appropriate complementary DNAs (cDNAs) encoding these proteins. The binding of AChR antibodies can be scored visually using indirect immunofluorescence. Unlike most other diagnostic antibody tests, this CBA allows detection of antibodies binding to AChRs in a natural membrane environment, where they adopt native conformational states and appropriate glycosylation levels and are clustered as they are at the neuromuscular junction. The proportion of patients with SNMG with autoantibodies to clustered AChRs ranges from 16% to 60%.16-18 These antibodies are mainly of the complement-fixing IgG1 subtype and have pathogenic mechanisms similar to those detected by RIPA.17 Subsequently, several groups have reported autoantibodies against other components of the neuromuscular junction, namely, agrin, LRP4, and collagen Q in a variable and generally low proportion of patients with SNMG.19-23 We aimed to assess the clinical usefulness of clustered AChR antibodies in MG. We also sought to describe the clinical features of patients seen within the Division of Clinical Neurology at the John Radcliffe Hospital in Oxford, England, who had been tested for clustered AChR antibodies since the implementation of the assay.

tibodies against clustered AChRs between November 2009 and November 2013, all of whom had been negative by RIPA for AChR and MuSK antibodies (Figure 1). Myasthenia gravis was diagnosed by clinical and electromyographic criteria. In patients with normal neurophysiology and lacking detectable autoantibodies, MG was diagnosed based on the presence of fatigable weakness and a positive response to treatment with cholinesterase inhibitors or immunosuppression by experienced myasthenia physicians (S.J., C.B., D.H.-J., M.I.L., and J.P.). This study was approved by the Clinical Audit Team at the John Radcliffe Hospital. Patients with MG lacking detectable AChR antibodies by RIPA or CBA or lacking MuSK antibodies by RIPA were referred to as having SNMG. All available samples from these patients were retrospectively tested for LRP4 antibodies using a CBA. The patients in whom a final diagnosis was not achieved were referred to as having an uncertain diagnosis. If a final diagnosis other than MG was made, the patients were referred to as having other diagnoses. The clinical features of patients with SNMG were compared with those of patients with MG with antibodies only to clustered AChRs. Data on the individual patients with MG, including demographic data, were collated. Race/ethnicity is self-assessed on a routine basis in all our patients as part of a general questionnaire using options defined by the investigators and was reported because of the unexpected high proportion of nonwhite patients. Clinical features were graded on a scale of 0 to 3 (0 is absent, 1 is mild, 2 is moderate, and 3 is severe). The distribution and severity of myasthenic weakness were classified according to the Myasthenia Gravis Foundation of America (MGFA) grading system.24 Clinical state of patients with MG after institution of treatment was classified according to the MGFA postintervention status.24

CBAs for Clustered AChR and LRP4 Antibodies For the clustered AChR assay, HEK cells were transfected with cDNAs expressing human AChR α, β, δ, and ε/γ subunits and rapsyn-enhanced green fluorescent protein in a ratio 2:1:1:1:1. Figure 1. Flowchart Showing Patients Included in the Study 138 Total 45 Other diagnoses 51 Uncertain diagnosis 42 MG

AChR RIPA negative MuSK RIPA negative Clustered AChR antibody positive

16 Patients

Methods Clinical Material We assessed all 138 patients seen within the Division of Clinical Neurology at the John Radcliffe Hospital who were tested for anE2

AChR RIPA negative MuSK RIPA negative Clustered AChR antibody negative LRP4 negative

21 Patients

AChR RIPA negative MuSK RIPA negative Clustered AChR antibody negative LRP4 not tested

5 Patients

The clustered acetylcholine receptor (AChR) cell-based assay detected antibodies in 16 of 42 (38.1%) patients with radioimmunoprecipitation assay (RIPA)–negative myasthenia gravis (MG). LRP4 indicates lipoprotein receptor–related protein 4; MuSK, muscle-specific tyrosine kinase.

JAMA Neurology Published online April 20, 2015 (Reprinted)



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For the LRP4 assay, HEK cells were transfected with cDNAs expressing human LRP4 and a chaperone protein (low-density lipoprotein receptor–related protein–associated protein 1) to enhance cell surface expression. Measurement of antibody binding was performed by indirect immunofluorescence as previously described.16 Results were measured by 2 observers (S.H., L.J., M.W., and M.I.L.) on a nonlinear visual scale from 0 to 4 (0 is no signal, 0.5 is unclear, 1 is weak positive, 2 is moderate positive, 3 is strong positive, and 4 is very strong positive). When the score differed, the mean result was given. Unclear results were repeated until consensus was achieved.

Results

Original Investigation Research

clinical diagnosis of MG, 51 (37.0%) had an uncertain diagnosis, and 45 (32.6%) had other diagnoses (Table 1). The patients with a clinical diagnosis of MG represented all the AChR antibody–negative and MuSK-negative MG cases by RIPA encountered during the study period.

Antibody Results on RIPA and CBA in the Total Cohort All patients with uncertain diagnosis and other diagnoses were negative for clustered AChR antibodies. In the MG group, 16 of 42 (38.1%) were positive for clustered AChR antibodies (Figure 1). One patient with an unclear CBA result was designated as having SNMG. All patients with SNMG who were tested for LRP4 antibodies (21 of 26) were negative by CBA.

Patients Included in the Study

Patients With Positive CBA and Negative Results on RIPA

In total, 138 patients were tested for clustered AChR antibodies during the period (Figure 1). Forty-two (30.4%) had a final

The clinical characteristics of 16 patients with clustered AChR antibodies (CBA score range, 1-3; median score, 2) are listed in Table 2 and Table 3. Ten of 16 were female. The age range was wide (age range, 1-52 years), and 62.5% were children (age range, 1-10 years), with 8 of 10 children initially seen before age 5 years. Seven patients (43.8%) were of black British, Caribbean, or African race/ethnicity (4 individuals) or mixed white Asian (3 individuals). The rest were white British or white other. Neurophysiology at diagnosis was abnormal in 8 patients (50.0%), normal in 6 patients (37.5%), and not assessed in 2 patients (12.5%). Chest imaging was normal in the 3 patients who were imaged. The presentations were predominantly ocular (62.5%) with ptosis and restricted eye movements, with no generalization during the follow-up period. Bulbar symptoms were scarce and mild (25.0%), and no patients had respiratory weakness. The maximum MGFA grades related to age at onset are shown in Figure 2A.

Table 1. Patients With Other Diagnoses Included in the Study No. of Patients (n = 45)

Diagnosis Acquired cranial nerve palsy

4

Brainstem pathology

2

Chronic fatigue syndrome

4

Congenital cranial dysinnervation syndrome Congenital myasthenic syndrome

5 13

Functional disorder

3

Mitochondrial myopathy

2

Motor neuron disease

1

Oculopharyngodistal myopathy

1

Other

6

Primary ocular pathology

4

Table 2. Clinical Features of 16 Patients With Antibodies Only to Clustered Acetylcholine Receptors (Negative on Radioimmunoprecipitation Assay) Patient No./ Sex/ Age, y

Race/ Ethnicity

Onset

Repetitive Nerve Stimulation

Single-Fiber EMG

CBA Score

1/F/1

Black

Ptosis, ophthalmoparesis, limb weakness

Normal

Not done

2.5a

2/F/1

White

Ptosis, limb weakness

Abnormal

Abnormal

1

3/F/2

Asian

Ptosis, ophthalmoparesis

Abnormal

Not done

3a

4/F/2

White

Ptosis

Not done

Not done

2a

5/F/2

Black

Ptosis

Not done

Not done

2a

6/M/3

White

Ptosis

Normal

Not tolerated

2

7/F/3

Asian

Ptosis, facial weakness

Normal

Not done

3

8/M/4

White

Ptosis, facial weakness, bulbar weakness

Abnormal

Not done

3

9/M/7

Black


Not done

Abnormal

1

10/F/10

White


Normal

Normal

2

11/M/18

Asian


Normal

Abnormal

1.5

12/F/23

Black

Ophthalmoparesis

Not done

Normal

3 1

13/M/25

White

Limb weakness

Abnormal

Abnormal

14/F/42

White


Normal

Normal

1.5

15/F/47

White

Limb weakness, ophthalmoparesis

Abnormal

Abnormal

2a

16/M/52

White


Not done

Abnormal

1.5

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Abbreviation: CBA, cell-based assay. a

Five patients were receiving immunosuppressive treatment at the time of antibody testing.

(Reprinted) JAMA Neurology Published online April 20, 2015



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Table 3. Clinical Features of 16 Patients With Antibodies Only to Clustered Acetylcholine Receptors (Negative on Radioimmunoprecipitation Assay) Patient No./ Sex/ Age, y

Features During Follow-up on a Scale of 0 (Absent) to 3 (Severe)

Ptosis

Ophthalmoparesis Diplopia

RespiMaximum ratory Facial Bulbar Neck Limb MGFA Weakness Weakness Weakness Weakness Weakness Grade Treatment

Postintervention Status

1/F/1

2

3

0

1

1

1

1

0

IIA

Pyridostigmine, prednisolone, azathioprine

Pharmacological remission, improved

116

2/F/1

1

0

1

1

1

0

1

0

IIA

Complete Pyridostigmine, stable prednisolone, 3, 4-diaminopyridine remission, improved

147

3/F/2

2

2

0

0

0

0

0

0

I

Pyridostigmine, prednisolone

No remission, improved

22

4/F/2

2

1

0

0

0

0

0

0

I

Pyridostigmine, prednisolone, intravenous immunoglobulin

Minimal manifestations (MM-3), improved

24

5/F/2

2

0

0

0

0

0

0

0

I



69

6/M/3

2

0

0

0

0

0

0

0

I

No

Complete stable remission, improved

12

7/F/3

1

0

1

1

0

1

1

0

IIA

Pyridostigmine


84

8/M/4

2

0

0

1

1

1

0

0

IIA



47

9/M/7

2

2

2

0

0

0

0

0

I

Pyridostigmine


24

10/F/10

2

2

1

0

0

0

0

0

I

Pyridostigmine


72

11/M/18 2

3

2

0

0

0

0

0

I


a

12/F/23

2

2

1

0

0

0

0

0

I



84

13/M/25 2

1

0

1

0

0

1

0

IIA

Pyridostigmine

Minimal manifestations (MM-2), unchanged

74

14/F/42

2

2

1

1

0

0

0

0

I


a

15/F/47

3

0

1

1

2

0

3

0

IVA

Pyridostigmine, prednisolone, azathioprine, intravenous immunoglobulinb


37

16/M/52 3

1

1

0

0

0

0

0

I



48

Abbreviations: EMG, electromyography; MGFA, Myasthenia Gravis Foundation of America. a

6

6

the disease duration was less than 12 months. b

This patient also had thymectomy.

The MGFA postintervention status was not applicable to 2 patients in whom

Most of the cases were successfully treated with pyridostigmine bromide and prednisolone or with pyridostigmine alone, and only 2 of 16 patients (12.5%) needed further E4

Follow-up, mo

immunosuppression with azathioprine. Five patients were receiving immunosuppressive treatment at the time of antibody testing. Thymectomy was performed only in 1 patient,




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Figure 2. Age at Onset, Maximum Myasthenia Gravis Foundation of America (MGFA) Grade, and MGFA Postintervention Status in Patients With Myasthenia Gravis (MG) A Clustered AChR Antibody–Positive MG

Green: AChRs + rapsyn-EGFP Red: Serum + antihuman IgG

10 >50 y 31-50 y 8

11-30 y

No. of Patients

0-10 y 6

4

2

0

I

II

III

IV

V

MGFA Grade B

SNMG Green: AChRs + rapsyn-EGFP Red: Serum + antihuman IgG

No. of Patients

15

10

5

0

I

II

IV

III

V

MGFA Grade C

Clustered AChR Antibody–Positive MG (left) and SNMG (right) 10

20

8

No. of Patients

No. of Patients

15 6

4

10

5 2

0

CSR

PR

MM

NR

0

CSR

MGFA Postintervention Status

PR

MM

NR

MGFA Postintervention Status

Patients with antibodies only to clustered acetylcholine receptors (AChR) have younger age at onset and a milder MGFA grade (A) compared with patients with seronegative MG (SNMG) (B). Patients with clustered AChR antibodies went into remission in a higher proportion compared with patients with SNMG (C). The images represent the binding of IgG antibodies to AChRs clustered by

rapsyn-EGFP on HEK cells surface (green) and detected by antihuman IgG (red). Positive binding of serum from a clustered AChR antibody-positive MG patient (A). Negative binding of serum from a SNMG patient (B). CSR indicates complete stable remission; MM, minimal manifestations; NR, no remission; and PR, pharmacological remission.

who had the highest MGFA grade (IVA). Pathological examination showed an atrophic thymus, with no evidence of thymoma or lymphocytic infiltrations.

The mean (SD) follow-up period was 54.5 (40.6) months. Among 14 patients with adequate follow-up, the outcome was good in all but 3 patients, with complete stable remission in

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Table 4. Clinical Features of 42 Patients With Seronegative Myasthenia Gravis (SNMG) Compared With Patients With Antibodies Only to Clustered Acetylcholine Receptors (AChR)

Variable

SNMG (n = 26)

Clustered AChR Antibody–Positive MG (n = 16)

M-F ratio

1:5.5

1:1.7

Age at onset, median (range), y

38 (2-72)

6 (1-52)a

3 (11.5)

10 (62.5)a

Ocular

15 (57.7)

11 (68.8)

Bulbar

5 (19.2)

1 (6.3)

Limb

0

1 (6.3)

Generalized

6 (23.1)

3 (18.8)

Ocular, No. (%)

11 (42.3)

10 (62.5)

Generalized, No. (%)

15 (57.7)

6 (37.5)

Maximum MGFA grade

I (n = 11) IIA (n = 4) IIB (n = 2) IIIA (n = 3) IIIB (n = 4) IVB (n = 2)

I (n = 10) IIA (n = 5) IVA (n = 1)

Bulbar symptoms, No. (%)

12 (46.2)

4 (25.0)

Prepubertal age, No. (%) Clinical presentation, No. (%)

Clinical subtype

Respiratory symptoms, No. (%) Abnormal neurophysiology, No. (%)

6 (23.1) 16 (61.5)

0 8/14 (57.1)

Treatment, No. (%) Prednisolone

15 (57.7)

Other immunosuppression

11 (42.3)

11 (68.8) 2 (12.5)

Thymectomy

5 (19.2) Including 1 patient with thymoma

1 (6.3)

Postintervention statusb Remission, including CSR or PR Follow-up, mean (SD), mo

2/24 (8.3)

7/14 (50.0)a

61.27 (79.96)

54.50 (40.60)

Abbreviations: CSR, complete stable remission; MGFA, Myasthenia Gravis Foundation of America; PR, pharmacological remission. a

P ⱕ .05.

b

The MGFA postintervention status was not applicable to 2 patients with SNMG and 2 patients with clustered AChR antibody–positive MG in whom the disease duration was less than 12 months.

4, pharmacological remission in 3, and minimal manifestations in 4. The mean (SD) time from onset of symptoms to complete resolution was 78.75 (55.33) months. However, 3 children with ocular or mild generalized disease did not attain remission.

Comparison of Patients With MG Without Clustered AChR Antibodies The clinical characteristics of 26 patients with SNMG with negative CBA results are summarized in Table 4 and Figure 2B and C. Overall, the patients with clustered AChR antibodies had younger age at onset (P = .02, Mann-Whitney test) and a trend toward milder disease (P = .06, Fisher exact test), and a higher proportion attained clinical remission (P = .03, Fisher exact test) compared with the patients with SNMG who did not have clustered AChR antibodies. E6

Discussion We confirm that the clustered AChR CBA improves diagnostic sensitivity for MG. We show that patients with these antibodies have a milder phenotype than patients without any detectable AChR antibodies, with younger onset age and complete or pharmacological remission in 7 of 14 patients with adequate follow-up. Cell-based assays were recently established for the improved detection of AChR antibodies in patients who were previously seronegative. Herein, we show that the CBA is helpful in the diagnostic workup of MG, detecting AChR antibodies in 38.1% (16 of 42) of negative patients with MG with 100% specificity. On the other hand, none of the patients with SNMG who were tested for LRP4 antibodies were positive. This is in keeping with our ongoing observations that the incidence of this antibody is low. Autoantibodies detected by CBA have pathogenic mechanisms similar to those detected by RIPA and include complement-mediated lysis.16,17 Complement deposition was noted in a passive transfer model of clustered AChR antibody MG.17 Although not formally described, it is likely that the IgG1 antibodies detected by CBA cross-link the clustered AChRs on the cell surface by divalent binding, thus explaining the increased sensitivity of the CBA. Alternatively, a less likely explanation could be the loss of antigenic determinants in the solubilized AChRs used in the RIPA. Cell-based assays require the use of live cells, tissue culture facilities, and expertise with performing and interpreting the assay, which are scored visually using indirect immunofluorescence. Because of these limitations, their current use is mainly confined to specialized research centers. However, if these difficulties were overcome, fluorescence-activated cell sorting analysis16 could be a suitable option for an automated and objective analysis. Patients with antibodies only to clustered AChRs had early onset and overall milder disease, with predominantly isolated ocular symptoms, low generalization rate, better prognosis, and no need for thymectomy. These results differ from those of patients with SNMG in the present study and differ from the classic AChR MG phenotype seen in large series with respect to age at onset, MGFA grades at maximum severity, and prognosis.25,26 However, because of the small numbers of patients included herein, their low MGFA grades were not significantly different compared with patients with SNMG (P = .06). Our group previously found that 50% of patients with RIPA-seronegative ocular MG had complement-fixing IgG clustered AChR antibodies.17 Results from 2 other studies18,27 have suggested that individuals with antibodies to clustered AChRs have a milder disease, but those studies did not include pediatric populations. Juvenile MG, especially in prepubertal children, is reported to differ from adult MG, with more ocular myasthenia, lower rates of generalization, better prognosis with a higher probability of achieving remission, and a lower frequency of AChR antibodies by RIPA.28-31 However, the previous observation of similar female and male prevalence28,29 differs from our 2:1 ratio in those with clustered AChR positivity. Serone-




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gative MG was more common in prepubertal (44%) compared with peripubertal (18%) and postpubertal (0%) onset of MG.32 Although it is possible that some of those individuals had an undiagnosed congenital myasthenic syndrome, we also found a high proportion of prepubertal onset of MG (66.6%) in the group of patients with autoantibodies against clustered AChRs who were previously negative on RIPA. Therefore, the usefulness of the CBA is shown with respect to the differential diagnosis with congenital myasthenic syndrome because this is the major group with an alternative diagnosis and both are rare disorders in childhood.33 Thymectomy should be considered as a treatment option early in the course of generalized AChR antibody–positive juvenile MG.34 However, indications for thymectomy within the prepubertal and seronegative groups are less clear.35 In our cohort of pediatric patients with antibodies only to clustered AChRs, thymectomy was not performed, probably related to the mildness of disease, high rate of pharmacological remission, and young age of some patients. In the French study18 cited earlier, thymectomy was performed in 2 individuals (ages 14 and 18 years) with antibodies to clustered AChRs and revealed thymus hyperplasia and thymoma. A further study16

ARTICLE INFORMATION Accepted for Publication: February 17, 2015. Published Online: April 20, 2015. doi:10.1001/jamaneurol.2015.0203. Author Contributions: Drs Vincent and Palace had full access to all the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. Drs Vincent, Leite, and Palace contributed equally to this work. Study concept and design: Rodríguez Cruz, Al-Hajjar, Beeson, Vincent, Leite, Palace. Acquisition, analysis, or interpretation of data: All authors. Drafting of the manuscript: Rodríguez Cruz, Al-Hajjar, Vincent, Palace. Critical revision of the manuscript for important intellectual content: All authors. Statistical analysis: Rodríguez Cruz. Administrative, technical, or material support: Huda, Jacobson, Woodhall, Beeson. Study supervision: Vincent, Palace. Conflict of Interest Disclosures: Dr Rodríguez Cruz reported being supported by the National Health Service National Specialised Commissioning Group for Congenital Myasthenia. Dr Huda reported being supported by a Watney, Myasthenia Gravis Association, and Oxford Biomedical Research Centre fellowship. Ms Buckley reported receiving research support from the Medical Research Council, United Kingdom. Dr Vincent reported serving on scientific advisory boards for the Patrick Berthoud Charitable Trust, the Brain Research Trust, and the Myasthenia Gravis Foundation of America; reported receiving funding for travel and a speaker honorarium from Baxter International, Inc, and Biogen, Idec; reported being an associate editor for Brain; reported earning royalties from the publication of Clinical Neuroimmunology (Blackwell Publishing, 2005) and Inflammatory and Autoimmune Disorders of the Nervous System in Children (Mac Keith Press, 2010); reported obtaining research support from the European jamaneurology.com


in adult patients reported evidence of thymic involvement, with typical lymphocytic infiltrates and germinal centers less striking but in a fashion similar to the classic AChR MG. The reasons underlying the clinical differences between prepubertal juvenile MG and adult MG are unknown. We found a female predominance and a high proportion of nonwhite patients (43.7%) within our cohort, especially black British individuals. This agrees with the observation that juvenile MG is rare in patients of white race/ethnicity.33 In Asian populations, juvenile MG and ocular MG have been reported to represent almost half of the MG cases.36 Chinese racial/ethnic groups have also been reported to have lower antibody titers on RIPA, which could be related to a higher frequency of ocular cases or milder disease.37

Conclusions In conclusion, we have shown that measuring antibodies to clustered AChRs by CBA is helpful for the diagnosis of SNMG. This is particularly true in children, who tend to have early onset, milder disease, and increased chance of spontaneous remission.

Union, National Institute for Health Research Oxford Biomedical Research Centre, Euroimmun AG, and the Sir Halley Stewart Trust; and reported receiving MuSK antibody royalties and consulting fees from Athena Diagnostics, Inc. Dr Leite reported being supported by the National Health Service National Specialised Commissioning Group for Neuromyelitis Optica and by the National Institute for Health Research Oxford Biomedical Research Centre and reported receiving speaking honoraria from Biogen Idec and travel grants from Novartis. Dr Palace reported being partly funded by highly specialized services to run a national congenital myasthenia service and a neuromyelitis optica service; reported receiving support for scientific meetings and honoraria for advisory work from Merck Serono, Biogen Idec, Novartis, Teva, Chugai Pharma, and Bayer Schering and unrestricted grants from Merck Serono, Novartis, Biogen Idec, and Bayer Schering; reported that her hospital trust receives funds for her role as a clinical lead for the United Kingdom Department of Health risk-sharing scheme; reported receiving grants from the National Multiple Sclerosis Society and The Guthie-Jackson Charitable Foundation for unrelated research studies; and reported serving as a board member for the charitable European MS foundation “The Charcot Foundation” and on the steering committee for the European collaborative multiple sclerosis imaging group “Magnetic Resonance Imaging in Multiple Sclerosis.” No other disclosures were reported. REFERENCES 1. Lindstrom JM, Seybold ME, Lennon VA, Whittingham S, Duane DD. Antibody to acetylcholine receptor in myasthenia gravis: prevalence, clinical correlates, and diagnostic value. Neurology. 1976;26(11):1054-1059. 2. Vincent A, Newsom-Davis J. Acetylcholine receptor antibody as a diagnostic test for myasthenia gravis: results in 153 validated cases

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