ARTICLE OPEN ACCESS

Inflammatory myopathy with myasthenia gravisThymoma association and polymyositis pathology

Naohiro Uchio MD Kenichiro Taira MD Chiseko Ikenaga MD Masato Kadoya MD Atsushi Unuma MD

Kenji Yoshida MD Setsu Nakatani-Enomoto MD PhD Yuki Hatanaka MD PhD Yasuhisa Sakurai MD PhD

Yasushi Shiio MD PhD Kenichi Kaida MD PhD Akatsuki Kubota MD PhD Tatsushi Toda MD PhD

and Jun Shimizu MD PhD

Neurol Neuroimmunol Neuroinflamm 20196e535 doi101212NXI0000000000000535

Correspondence

Dr Shimizu

jshimizu-tkyuminacjp

AbstractObjectiveTo provide evidence that idiopathic inflammatory myopathy (IM) with myasthenia gravis(MG) frequently shows thymoma association and polymyositis (PM) pathology and sharesclinicopathologic characteristics with IM induced by immune checkpoint inhibitors (ICIs)

MethodsWe analyzed the clinicopathologic features of 10 patients with idiopathic IM andMG identifiedin 970 consecutive patients with biopsy-proven IM

ResultsSeven patients (70) had thymoma IM and MG were diagnosed with more than 5-year timedifference in 6 thymomatous patients and within 1 year in 1 thymomatous and 3 non-thymomatous patients Seven thymomatous patients showed rhabdomyolysis-like features withrespiratory failure (47) dropped head (37) cardiac involvement (27) and positive antindashacetylcholine receptor (anti-AChR) and anti-titin antibodies (77 and 46 respectively) butrarely showed ocular symptoms (27) or decremental repetitive nerve stimulation (RNS)responses (17) at IM diagnosis Three nonthymomatous patients showed acute cardiore-spiratory failure with rhabdomyolysis-like features (13) positive anti-AChR and anti-titinantibodies (32 and 22 respectively) and fluctuating weakness of the skeletal muscle withoutocular symptoms (33) Muscle pathology showed a PM pathology with infiltration of CD8-positive CD45RA-negative T-lymphocytes (99) scattered endomysial programmed cell death1 (PD-1)ndashpositive cells (99) and overexpression of programmed cell death ligand 1 (PD-L1)on the sarcolemma of muscle fibers around the infiltrating PD-1ndashpositive cells (79)

ConclusionRhabdomyolysis-like features positive anti-AChR antibody without decremental RNSresponses and PD-L1 overexpression are possible characteristics shared by ICI-induced IMFrequent thymoma association in patients with idiopathic IM and MG may suggest thymoma-related immunopathogenic mechanisms including dysregulation of the immune checkpointpathway

From the Department of Neurology (NU KT CI AU AK TT JS) Graduate School of Medicine University of Tokyo Division of Neurology (MK KK) Department of InternalMedicine National Defense Medical College Saitama Division of Neurology (Y Shiio) Tokyo Teishin Hospital Department of Neurology (Y Sakurai) Mitsui Memorial HospitalDepartment of Neurology (YH) Teikyo University School of Medicine and Department of Neurology (KY SN-E) Fukushima Medical University Japan

Funding information and disclosures are provided at the end of the article Full disclosure form information provided by the authors is available with the full text of this article atNeurologyorgNN

The Article Processing Charge was funded by the authors

This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives License 40 (CC BY-NC-ND) which permits downloadingand sharing the work provided it is properly cited The work cannot be changed in any way or used commercially without permission from the journal

Copyright copy 2018 The Author(s) Published by Wolters Kluwer Health Inc on behalf of the American Academy of Neurology 1

Idiopathic inflammatory myopathies (IMs) are a heteroge-neous group of muscle disorders Myasthenia gravis (MG)is rarely associated with IM The reported clinical featuresof patients with both idiopathic IM and MG includedbrachio-cervical weakness or dropped head1ndash5 respiratorydecompensation3ndash10 muscle swelling with pain411 cardiacinvolvement459 and markedly elevated serum creatine kinase(CK) levels468 In addition patients with antindashacetylcholinereceptor (anti-AChR) antibody (Ab)-positive thymomatousIM without MG symptoms showing rapidly progressiveweakness and respiratory failure with markedly elevated se-rum CK levels1213 have been described Although previousreports suggested some characteristics of idiopathic IMpatientswith MG andor thymoma clinical features including thetemporal relationship between the onset of IM and MG andthe prevalence of clinical characteristics including thymomaassociation have not been well known because of the lack ofsystematical study conducted in a series of patients with IMFurthermore pathologic findings have not been studiedsystematically

The rare combination of IM andMGhas been recently reportedin patients with immune-related adverse events induced byimmune checkpoint inhibitors (ICIs) targeting programmedcell death 1 (PD-1) or cytotoxic T-lymphocytendashassociatedprotein 4 (CTLA-4)14ndash17 The reported clinical features ofthese patients include cardiac involvement1415 andrhabdomyolysis-like features with markedly elevated serumCK levels1415 suggesting similarities of the clinical featuresof IM between the 2 groups ICI induced and idiopathicTherefore we analyzed the clinicopathologic characteristics ofIM associated with MG in a series of patients with biopsy-proven IM to determine whether some characteristic featuresare shared by ICI-induced and idiopathic patients with both IMand MG

MethodsPatientsClinical records and biopsy reports were reviewed for 970consecutive patients who were referred to our department forpathologic diagnosis between April 1986 andDecember 2017IM diagnosis was based on the criteria proposed by Bohan andPeter1819 in addition both (1) elevated serum CK levels and(2) muscle biopsy findings of inflammatory changes withmajor histocompatibility complex (MHC) class I expressionon non-necrotic muscle fibers and sometimes with necrotic

andor regenerating fibers18ndash20 were required Exclusion ofmuscular dystrophy by immunohistochemistry and clinicalfeatures was also required Inclusion body myositis (IBM)was excluded using the 188th European NeuromuscularCentre IBM criteria21 Sarcoid myopathy was excluded onthe basis of clinical and pathologic findings22 Patients whodeveloped IM as an adverse effect of drugs including ICIswere excluded from this study MG diagnosis was basedon clinical features of weakness with increased fatigabilityof skeletal muscles and one or more of the following 3 cri-teria (1) a positive edrophonium infusion test (2) decre-mental repetitive nerve stimulation (RNS) responses and (3)increased jitter or blocking on a single-fiber electromyogram(SFEMG) Thymoma diagnosis was made on the basis of CTof the anterior mediastinum tumor and confirmed by patho-logic examination

In recruiting patients with both IM and MG patients withIM who had a past or subsequent history of MG were in-cluded Because the patients with thymoma sometimes haveMG or subclinical MG or may develop MG during follow-upperiods the patients with IM who had a past or subsequenthistory of thymoma were also included By these methodsamong 970 consecutive patients with IM we found 10patients with both IM and MG (10) and 1 non-MG thy-momatous patient with IM (figure 1) The 11 patients withboth IM and MG included 3 patients who have been pre-viously reported in separate reports (no 11 Yamaguchiet al 2000 no 5 Maekawa et al 2014 and no 9 Suzukiet al 2014)23ndash25

Definition of onset of IM and MGIM onset was defined as the time of development of sustainedskeletal muscle weakness with elevated serum CK levels MGonset was defined as the earlier of 2 dates (1) the date ofdevelopment of weakness with increased fatigability of skel-etal muscles (eg fluctuating diplopia ptosis or skeletalmuscle power) or (2) the date of detection of abnormality onRNS or an SFEMG in weakened muscles

Histochemical andimmunohistochemical analysisStored frozen muscle samples were processed into 10-μmsections for routine histochemistry and immunohistochem-istry Immunohistochemistry were performed with a conven-tional method using the substrate 33rsquo-diaminobenzidinetetrahydrochloride (DAB) Primary antibodies against MHCclass I (Dako Agilent Technologies CA) MHC class II (Dako)

Glossaryanti-AChR = antindashacetylcholine receptor CK = creatine kinase CTLA-4 = cytotoxic T-lymphocytendashassociated protein 4IBM = inclusion body myositis ICI = immune checkpoint inhibitor IM = inflammatory myopathyMG = myasthenia gravisMHC = major histocompatibility complexMSA = myositis-specific autoantibody PD-1 = programmed cell death 1 PD-L1 =programmed cell death ligand 1 PM = polymyositisPSL = prednisoloneRNS = repetitive nerve stimulation SFEMG = single-fiber electromyogram

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CD4 (Dako) CD8 (Nichirei Biosciences Tokyo Japan)CD20 (Abcam Cambridge UK) CD21 (Dako) CD45leukocyte common antigen (Dako) CD45RA (Dako)CD68 (Dako) myxovirus resistance protein A (M x AAbcam) PD-1 (Abcam) programmed cell death ligand 1(PD-L1 Abcam) and CTLA-4 (Abcam) were used

Detection of autoantibodiesMyositis-specific autoantibodies (MSAs) including antindashJo-1antindashPL-7 antindashPL-12 antindashMi-2 and antindashsignal recogni-tion particle 54-kDa protein Abs were detected by dot-blotting using recombinant proteins (Diarect AG FreiburgGermany) Antindashtranscriptional intermediary factor 1 gammaand antindashmelanoma differentiation-associated gene 5 Abswere detected by immunoprecipitation and antindash3-hydroxy-3-methylglutaryl-CoA reductase Ab was detected by ELISAas described in our earlier study26 Anti-titin Ab was alsodetected by ELISA (DLD Diagnostika Hamburg Germany)

Standard protocol approvals registrationsand patient consentsThe study was approved by the institutional ethics commit-tees of the University of Tokyo Hospital Written informedconsent was obtained from each patient at the time of biopsyand serum sampling

Data availabilityAnonymized data will be shared by request from any qualifiedinvestigators

ResultsClinical features of patients at IM diagnosisTable 1 shows the clinical features and histopathologic find-ings of the skeletal muscles of 10 patients with both IM andMG the patientsrsquo detailed clinical features are described inappendix e-1 linkslwwcomNXIA95

The mean age at IM diagnosis was 60 years (range 47ndash78years) Six patients were women None of the patients hadskin rash or interstitial lung disease Seven patients were as-sociated with invasive thymoma and the other 3 had nothymoma The temporal relationship between the onset of IMandMG are as follows (1) 5 thymomatous patients (nos 1 2and 4ndash6) had preexisting MG (diagnosed more than 9 yearsbefore their IM diagnosis) (2) 1 thymomatous patient (no7) had preexisting IM (diagnosed with MG 5 years after theIM diagnosis) and (3) 1 thymomatous patient (no 3) and 3nonthymomatous patients (nos 8ndash10) had concurrent IMand MG (diagnosed with IM and MG within 1 year) Clinicalfeatures related to MG at IM diagnosis included fluctuatingocular symptoms (210) abnormal RNS responses onweakened extremity muscles (410) positive edrophoniuminfusion test (23) and positive antindashAChR Ab (910)

Thymomatous IM patients rarely showed fluctuating ocularsymptoms (27) and abnormal RNS responses on extremities(17) By contrast 3 nonthymomatous IM patients showedabnormal RNS responses (33) and positive edrophoniuminfusion test (22) but showed no ocular symptoms (03)

Before IM diagnosis 2 patients had a history of transientelevation of serum CK levels without limb weakness (4209IUL in no 1 and 518 IUL in no 8) Progression of weak-ness in IMwas rapid in 5 patients Four thymomatous patientswith normal RNS tests on extremities presented with rapidprogression of weakness with acute markedly elevated serumCK levels (1008ndash10226 IUL) at IM development(rhabdomyolysis-like features) and required ventilator sup-port One nonthymomatous patient (no 9) developed re-spiratory muscle weakness with relatively preserved limbmuscle strength cardiac failure and acute elevation of serumCK levels (1140 IUL)

The clinical features of the 10 patients included respiratoryfailure requiring ventilator support (510) severe weakness ofextremities (410) dropped head (410) myalgia (410)dysphagia (59) and cardiac involvement (310) One thy-momatous patient (no 7) had initial symptoms of muscleswelling and pain in the forearms

The mean serum CK level was 2712 IUL The serum testingfor autoantibodies demonstrated anti-AChR Ab (910)MSAs (09) and anti-titin Ab (68) A needle EMG per-formed for 9 patients showed spontaneous activities at rest inmost of them (89) Three patients (310) showed cardiacinvolvements A cardiac muscle biopsy performed in 1 patient

Figure 1 Flowchart of patient inclusion

Clinical and pathologic features of 1146 patients were evaluated based onBohan and Peterrsquos myositis criteria and serum CK levels and then 970patients were identified as more than possible polymyositis or dermato-myositis with an elevated serum CK level Among these patients 10 patientswith a history of MG in flow A and 8 patients with a history of thymoma inflow Bwere identified Finally 10 patients with MGwere recruited excludingthe 7 overlapping patients between flow A and flow B In addition 1 non-MGpatient with a history of thymoma was also recruited CK = creatine kinaseDM = dermatomyositis PM = polymyositis

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Table 1 Clinical and pathologic features of IM in patients with both IM and MG

Patient 1 2 3 4 5 6 7 8 9 10

Age at IMMG diagnosis (y) 5950 7356 5352 5038 5743 4936 4752 7878 7070 6060

Sex Male Female Male Female Male Female Female Male Female Female

MG diagnosis before IMdiagnosis (y)

+9 +17 lt1f +12 +14 +13 minus5 lt1 lt1 lt1

Thymoma + + + + + + + minus minus minus

WHO classification B2 B2 B2 ND B3 B2 B2 NA NA NA

Age at thymoma diagnosis(y)

42 56 53 38 43 36 47 NA NA NA

Thymoma status at IMdiagnosis

Under CHT No recurrence Untreated No recurrence Recurrence No progression Untreated NA NA NA

Disease activity of MG

Ocular symptomsfluctuating weakness atIM diagnosis

minusminus ++ minusminusf ++ minusminus minusminus minusminus minusminus minusminus minusminus

Decremental responses onRNS at IM diagnosis

minus minus minus + minus minus minus + + +

Positive edrophoniuminfusion test at IMdiagnosis

NE NE NE NE NE NE minus + NE +

Anti-AChR Ab at MGIMdiagnosis (nmolL)

ND22 11540 100ND 80300 1637 4110 ND12 477i 65i minusi

CK level at MG diagnosis(IUL)

ND ND 231 ND 185 41 ND 263 1140 282

Clinical features of IM at IMdiagnosis

History of transientlyelevated CK levels

+ minus minus minus minus minus minus + minus minus

Progression of weakness atIM diagnosis

Rapid No Rapid Chronic Chronic Rapid Rapid Chronic Rapid Chronic

Limb weakness Severe Nondashmild Severe Moderate Nondashmild Severe Severe Nondashmild Nondashmild Moderate

Dropped head + minus + minus + minus minus + minus minus

Dysphagia + minus + + + + minus minus ND minus

Continued

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Table 1 Clinical and pathologic features of IM in patients with both IM and MG (continued)

Patient 1 2 3 4 5 6 7 8 9 10

Respiratory failure + (NIPPV) minus + (MV) minus minus + (MV) + (MV) minus + (MV) minus

Myalgia + minus minus minus minus + + (swelling)h + minus minus

Fever minus Low grade minus minus minus Remittent Low grade minus minus minus

Cardiac involvement minus minus minus Anteroseptalhypokinesis

minus VT and diffusehypokinesis

minus minus VT and diffusehypokinesis

minus

Other complications minus minus DG DG Alopecia DG DPBand AIC

minus DG and DPB minus Pure red cell aplasia minus

Laboratory findings at IMdiagnosis

CK level (IUL) 10226 376 1008 341 1059 5102 7315 263 1140 292

CRP level (mgdL) 26 006 004 007 183 104 332 lt03 47 minus

ESR (mmh) 20 26 4 23 NE 19 73 30 NE NE

Myositis-specificautoantibodies

minus minus minus minus minus minus minus minus minus NE

Anti-titin Ab minus NE + + minus + + + + NE

Spontaneous activities atrest on nEMG

+ + + + + + + + NE minus

Histopathologic findings

Necrotic andorregenerating fibersa

+ + + plusmn plusmn + +++ + plusmn +

Perimysial inflammationb plusmn plusmn +++ ++ plusmn plusmn + +++ plusmn plusmn

Endomysial inflammationb + + + plusmn plusmn + +++ + + +

CD8+ T cells surroundinginvading non-necroticfibers

++ ++ ++ +minus +minus +minus ++ +minus +minus ++

CD45RA+ cell infiltration minus minus +g minus minus minus NE +g minus minus

MHC class I expressionc ++ +++ +++ + + +++ +++ +++ ++ +++

MHC class II expressionc + + + + + + + minus + +++

Granulomatous lesions minus minus minus minus minus minus + minus minus minus

Continued

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Table 1 Clinical and pathologic features of IM in patients with both IM and MG (continued)

Patient 1 2 3 4 5 6 7 8 9 10

CD20+ lymphoidaggregatesd

minus minus + minus minus minus NE + minus minus

MxA+ capillaries and fibers minus minus minus minus minus minus NE minus minus minus

PD-1+ cell infiltration + + + + + + NE + + +

PD-L1 upregulation onfiberse

++ minus + minus + + NE + ++ +

CTLA-4+ cell infiltration + minus + + minus minus NE minus minus +

Classification based onENMC criteria

DefinitePM

Definite PM DefinitePM

Probable PM Probable PM Probable PM Definite PM ProbablePM

Probable PM DefinitePM

Abbreviations AIC = autoimmune cholangitis CHT = chemotherapy CK = creatine kinase CRP = C-reactive protein DG = dysgeusia DPB = diffuse panbronchiolitis ESR = erythrocyte sedimentation rate MV = mechanicalventilation NA = not applicable ND = no data NE = not examined NIPPV = noninvasive positive-pressure ventilation nEMG = needle electromyogram VT = ventricular tachycardia WHO = World Health Organizationa Necrotic and regenerating fibers in 1 fascicle plusmn sparse + 1 to lt5 ++ 5 to lt20 and +++ ge20b Density of inflammatory cells minus 0 plusmn 1 to lt5 + 5 to lt20 ++ 20 to lt100 and +++ ge100c MHC class I or class II antigen on the sarcolemma of non-necrotic fibers minus 0 + 1 to lt25 ++ 25 to lt50 and +++ 50ndash100d CD20+ lymphoid aggregate was defined when more than 100 CD20+ cellsplace were observede PD-L1 upregulation on the sarcolemma of non-necrotic fibers minus none + invaded fibers limitedly and ++ invaded fibers and non-necrotic fibers around invaded fibersf The patient who was diagnosed with ocular MG developed IM 3 months after the introduction of immunotherapy for MGg CD45RA+ cells were observed only at perimysial lymphoid aggregates in these patientsh The patient showed myalgia with muscle swellingi Data at IM diagnosis

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(no 6) showed abundant T-lymphocyte infiltration consis-tent with lymphocytic myocarditis

Pathologic findingsWe observed various amounts of necrotic fibers and in-flammatory cells in both the perimysium and the endomysiumAll patients showed different levels of overexpression of MHCclass I on muscle fibers with various extent levels 1+ n = 2 2+n = 2 and 3+ n = 6 (table 1) Overexpression of MHC class IIon muscle fibers was observed in 9 patients (910) All patientsshowed the findings of endomysial CD8+ T-lymphocytes sur-rounding (1010) and sometimes invading (510) non-necrotic muscle fibers expressing MHC class I antigen andtherefore met the pathologic criteria of polymyositis (PM)20

(definite PM n = 5 probable PM n = 5 figure 2A)

Immunohistochemistry for CD45RA which was performedin 9 frozen sample available patients showed no endomysialCD45RA+ cells indicating that the endomysial CD8+

T-lymphocytes surrounding and sometimes invading non-necrotic muscle fibers were all CD45RAminus cells (99) Twopatients (nos 3 and 8) showed perimysial accumulation ofCD45RA+ cells (figure e-1 linkslwwcomNXIA95)

One thymomatous patient (no 7) showed granulomatousinflammatory cell invasion with multinucleated giant cells(figure 2B) The granulomas and giant cells were positive forCD68 Two patients (nos 3 and 8) showed focal perimysialaccumulation of mononuclear cells many of which werepositive for CD20 (figure 2 C and D)

Immunohistochemistry for PD-1 and PD-L1 showed scat-tered endomysial PD-1+ cells (99) and overexpression ofPD-L1 on the sarcolemma of muscle fibers near PD-1+ cells(79) Of interest 2 patients (nos 1 and 9) showed PD-L1overexpression extending on non-necrotic fibers around thefibers invaded by the PD-1+ cells (figure 2 E and F) Im-munohistochemistry for CTLA-4 demonstrated CTLA-4+

cells in endomysial infiltration (49) (figure e-2 linkslwwcomNXIA95)

Associated thymomaThe World Health Organization classifications of invasivethymoma in 7 patients were B2 (n = 5) B3 (n = 1) and nodata available (n = 1) (table 1) The mean age at thymomadiagnosis was 45 years (range 36ndash56 years) Thymoma di-agnosis was preceded to IM diagnosis more than 12 years(range 12ndash17 years) in 5 patients (nos 1 2 and 4ndash6) andboth thymoma and IM were codiagnosed in 2 patients (nos 3and 7) The disease statuses of thymoma at IM diagnosis wereas follows no recurrence or progression without therapy (n =3) untreated (n = 2) recurrence (n = 1) and no progressionunder chemotherapy (n = 1)

Thymomatous IM patient withoutMG symptomsOur series included 1 thymomatous IM patient withoutMG symptoms (no 11 details in appendix e-1 linkslww

comNXIA95) This 51-year-old woman had moderate tosevere weakness in her limbs and respiratory muscles re-quiring ventilator support Laboratory data showed an el-evated serum CK level (695 IUL) and positive anti-AChRand anti-titin Abs Needle EMG showed spontaneous ac-tivities but RNS test was normal Chest CT showed an an-terior mediastinum tumor (1 cm in size) which was revealedto be necrotic tissues Histopathologic diagnosis of musclebiopsy was definite PM20 She was successfully treated withprednisolone (PSL) Her anterior mediastinum tumor in-creased in size without neurologic symptoms Five years afterthe diagnosis of IM the tumor was resected and pathologi-cally confirmed as combined B2B3 thymoma Her 11-yearfollow-up showed no exacerbation of IM or thymoma ordevelopment of MG

Figure 2 Histopathologic findings of patients with both in-flammatory myopathy and myasthenia gravis

(A) Immunohistochemical staining for CD8 showing CD8-positive cells in-vading non-necrotic muscle fibers in patient 1 (B) Hematoxylin-eosin (HE)staining showing granulomatous lesion with multinucleated giant cells inpatient 7 (C and D) Serial sections with HE staining and immunohisto-chemical staining for CD20 showing perimysial aggregation of CD20-positivecells in patient 8 (E and F) Serial sections with immunohistochemicalstaining for PD-1 and PD-L1 showing endomysial PD-1ndashpositive cells and PD-L1 upregulation on non-necrotic fibers in patient 9 Scale bar = 50 μm PD-1 =programmed cell death 1 PD-L1 = programmed cell death ligand 1

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Clinical courses and response to treatmentsOne patient (no 8) showed normalized serumCK levels duringbed rest and refused immunotherapy He showed no exacer-bation of symptoms for 26 years until final follow-up Theremaining 9 patients including 2 patients who showed nor-malization of serumCK levels during bed rest were treated withimmunotherapy (table 2) The treatments included immuno-suppressants alone in 1 patient (no 4) and increasing doses oforal PSL (20ndash60 mg) in the other 8 patients in combinationwith various immunosuppressive therapies oral immunosup-pressants (n = 4) high-dose IV methylprednisolone (n = 4)high-dose IV immunoglobulin (n = 2) and immunoabsorption(n = 1) Of interest the elevated serum CK levels normalizedwithin 3weeks in all the treated patients including the 3 patientswith very high serum CK levels (nos 1 6 and 7)

The mean follow-up period was 38 years (range 05ndash109years) During the follow-up period 3 patients died Thecauses of death were sepsis following diffuse panbronchiolitis(n = 2 patients 5 and 7) and bronchopneumonitis compli-cated by cardiac dysfunction and pure red cell aplasia (n = 1patient 9) Themodified Rankin Scale scores at final follow-upof the 7 patients who were alive were grade 0 (n = 1) grade 1(n = 4) and grade 3 (n = 2)

DiscussionThe prevalence of thymoma association in patients with bothIM and MG (70) was much more than that in patients withonly MG (10)27 Thymoma is known to be associated withvarious autoimmune diseases The frequent thymoma associ-ation in IM patients with both IM andMG should indicate thatthe association of these 2 diseases is not coincidental but in-duced by thymoma-related immunopathogenic mechanisms

In 7 thymomatous patients with both IM and MG ocularsymptoms or decremental RNS responses were rarely observedat IM diagnosis (27 or 17 respectively) Among them 4patients with normal RNS responses developed rapidly pro-gressive muscle weakness and respiratory failure with acuteelevation of serum CK to very high levels (rhabdomyolysis-likefeatures) Previous studies reported patients with similar clin-ical features IM and antindashAChR Ab-positive thymomatousMG presenting with normal RNS responses28 and rapidlyprogressive weakness and respiratory failure in conjunctionwith markedly elevated serum CK levels712 Our findingsdemonstrated that thymomatous patients with both IM andMG frequently showed nonsimultaneous progression of IMand MG and often included those with severe progression andmarkedly elevated serum CK levels The nonsimultaneousprogression of IM and MG suggests that distinctly differentthymoma-related immunopathogenic mechanisms on skeletalmuscles are involved in the development of each disease

Three nonthymomatous IM patients in our study sharedthe following characteristic features with thymomatous IM

patients none showed ocular symptoms 1 patient showedacute respiratory and cardiac failure with acute serum CKlevel elevation 2 patients (22) showed positive anti-titinAb and all 3 patients showed PM pattern pathology charac-terized by CD8+ T-lymphocytendashmediated cellular autoimmu-nity PM pattern pathology is a rare pathologic finding amongpatients with IM2930 and is suggested as a secondary pathologicpattern associated with other systemic autoimmune diseasesincluding connective tissue diseases viral infections cancerand drug toxicity31 The overlapping of clinical serologic andpathologic characteristics between thymomatous and non-thymomatous IM patients may suggest a common immunemechanism underlying the IM development in patients withboth IM and MG irrespective of thymoma association How-ever the immunemechanical background of nonthymomatousIM patients may be heterogeneous considering that 1 non-thymomatous IM patient (no 10) was seronegative for anti-AChR Ab

We found 1 non-MG thymomatous IM patient (no 11) withfeatures similar to those of thymomatous patients with bothIM and MG except for the presentation of MG A previousreport described 1 non-MG thymomatous IM patient withsimilar clinical features positive anti-AChR Ab normal RNSresponses a very high serum CK level (7271 IUL) anda pathologic finding of endomysial CD8+ cells with CD68 +

giant cells13 The clinical features in both patients supportedthe idea that thymoma-related immunopathogenic mecha-nisms can affect skeletal muscle toward the development ofIM even without MG

At the time of IM diagnosis 5 patients had a preexistingthymoma and among them 2 patients showed no signs ofrecurrence In the literature we found a total of 4 patients withboth IM and MG with preexisting thymoma for 4ndash17 yearsbefore IM diagnosis4932 The thymoma status of the 4patients at IM diagnosis was not described except in 1 patientwho was autopsied and showed no thymoma recurrenceHowever some studies have described thymoma recurrencelong after thymectomy33 and microscopic thymoma (smallerthan 1 mm) which cannot be detected by imaging studies34

Considering the suggested thymoma-related immunopatho-genic mechanisms in the development of IM we recommendnot only careful checkup for thymoma recurrence in patientswith preexisting thymoma but also regular checkup for thy-moma development in nonthymomatous patients with bothIM and MG

The presence of endomysial infiltration of T-lymphocytes(lymphorrhage) in the skeletal muscle tissue of patients withMG has been known for decades particularly in patients withthymomatousMG3235 One suggestion is that the lymphocyticinfiltration does not represent autoimmune IM but rather aninfiltration consisting of CD8+CD45RA+-naive T-lymphocytesderived from lymphocyte-rich thymoma However in our se-ries CD8+CD45RAminus cells but not CD8+CD45RA+ naive cellswere the lymphocytes infiltrating the endomysium and the

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Table 2 Treatment of IM in patients with both IM and MG

Patient 1 2 3 4 5 6 7 8 9 10

Treatment

Immunotherapy after IMdiagnosis

IVMP PSL 55 mgand TCR 3 mg

PSL 30 mg PSL 50 mg andTCR 3 mg

CyA 200 mgand PB

PSL 20 mg IVMP PSL 50 mg IVIgIAPP and TCR 15 mg

IVMP PSL 60 mg IVIgand MTX 125 mg

PB IVMP andPSL 40 mg

PSL 60 mgand PB

CK normalization by rest beforeimmunotherapya

minus + (8 d) minus minus + (20 d) minus minus + (18 d) minus minus

Period between treatmentintroduction and CKnormalization

2 w NA ND 1 w NA lt3 w 2 w NA 4 d 6 d

Prognosis

Follow-up period (y) 13 30 27 29 13 05 61 26 02 17

MGFA-PIS at final follow-up MM MM Improved MM Died of MG PR Died of MG MM Died of MG PR

IM recurrence minus minus minus minus minus minus + minus minus minus

MG recurrence minus minus minus minus minus minus NA minus minus minus

Modified Rankin Scale change(IM diagnosisrarrfinal follow-up)

3rarr0 1rarr1 3rarr3 2rarr1 5rarr6 5rarr3 5rarr6 2rarr1 5rarr6 2rarr1

Abbreviations CK = creatine kinase CyA = cyclosporin A d = day IAPP = immunoadsorption plasmapheresis IVIg = IV immunoglobulin IVMP = IV methylprednisolone MGFA-PIS = Myasthenia Gravis Foundation of Americapostintervention status MM = minimal manifestation MTX = methotrexate NA = not applicable ND = no data PB = pyridostigmine bromide PR = pharmacologic remission PSL = prednisolone TCR = tacrolimus w = weeka Period in brackets refers days between peak and normalization of CK levels before treatment

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extensive expression of MHC class I was observed in allpatients (99 and 1010 respectively) suggesting a true cell-mediated PM pattern of inflammation as a pathologic pheno-type in our patients

In addition to PM pattern pathology massive collections ofCD20+ B-lymphocytes were observed in 2 patients andCD68+ granulomas in 1 patient These pathologic findingshave already been reported in patients with both idiopathicIM and MG39ndash111328 Because the thymus gland is a centrallymphatic organ that provides various immunologic functionsmultifactorial autoimmune dysregulation in thymic disordersincluding thymoma has been reported36 Further studies areneeded to elucidate the overall immune mechanisms thatcause these complicated pathologic features

Some immune-related adverse events induced by antitumortherapy with ICIs have been recently reported to cause theassociation of MG with IM141617 A recent study of 12nonthymomatous patients who developed MG after anti-tumor therapy with nivolumab (an antindashPD-1 monoclonalAb) reported that 4 patients had IM and 3 patients hadmyocarditis including 1 patient with both IM and myocar-ditis15 This study also showed the following clinical featuresof patients with nivolumab-induced MG markedly elevatedCK levels (1012 83) positive anti-AChR Ab (101283) positive edrophonium infusion test (47 57) andabnormal decremental RNS responses (27 29) Patientswith ICI-induced IM have been often reported to haverhabdomyolysis-like clinical features with myocarditis orrespiratory failure37 Therefore the assumption that patientswith ICI-induced IM share similar immunopathogenicmechanisms with patients with both idiopathic IM and MGshould be reasonable Moreover we recently found PD-1+

cell infiltration and extended PD-L1 (a ligand of PD-1)overexpression on non-necrotic fibers in patients withantindashPD-1 Ab-induced IM16 The similarity of clinical fea-tures between patients with ICI-induced IM and those withidiopathic IM and MG prompted us to examine PD-1 PD-L1 and CTLA-4 expressions in our patients Subsequently wefound PD-L1 overexpression on non-necrotic muscle fibersaround PD-1+ cells in 7 patients including 2 patients withextended PD-L1 overexpression and endomysial infiltrationof CTLA-4+ cells in 4 patients CTLA-4+ cells in endomysialinfiltrates with invasion of muscle fibers have been reportedin patients with idiopathic PM or IBM38 Both PD-1PD-L1and CTLA-4B7 pathways are involved in the induction ofimmune tolerance on activated cytotoxic T-lymphocytesalthough the 2 pathways operate at different stages of animmune response39 The expression of PD-1 PD-L1 andCTLA-4 in IM pathology may suggest dysregulation ofimmune checkpoint molecules in the microenvironmentof IM in patients with IM and MG The backgrounds ofthe dysregulation of the immune checkpoint molecules inour patients remain unclear However a previous reporthas shown that genetic polymorphisms of CTLA-4 are as-sociated with thymomatous MG patients40 The finding

indicates involvement of the immune checkpoint pathwayin the development of autoimmune diseases in thymoma-tous patients

Our study has some limitations First the number of patientswith both IM and MG is small because of the rarity of si-multaneous occurrence of these 2 diseases Second we didnot always follow the clinical courses in patients with IM fora long time enough to detect MG association Third MGactivity may be underestimated because SFEMG or edro-phonium infusion test had not been performed in all of ourpatients Despite these limitations our study showed thatidiopathic IM with MG frequently shows thymoma associ-ation and PM pathology In addition this study demon-strated that idiopathic IM with MG shares clinicopathologiccharacteristics with IM induced by ICIs Further studiesshould be conducted to determine the essential mechanismsof thymoma-related autoimmune dysregulation in patientswith both IM and MG particularly focusing on the roles ofimmune checkpoint molecules

Author contributionsN Uchio study design and concept acquisition of dataanalysis and interpretation and drafting of the manuscriptK Taira C Ikenaga A Unuma M Kadoya and K Yoshidaacquisition of data S Nakatani-Enomoto Y HatanakaY Sakurai Y Shiio and K Kaida acquisition of data andcritical revision of the manuscript A Kubota critical re-vision of the manuscript T Toda study supervision JShimizu study design and concept acquisition of dataanalysis and interpretation and critical revision of themanuscript

AcknowledgmentThe authors thank Naoko Tokimura Masako Nishizawa andYuki Ogiwara for technical assistance

Study fundingThis work was supported by the Health and Labour SciencesResearch Grant on Rare and Intractable Diseases (Validationof Evidence-based Diagnosis and Guidelines and Impacton QOL in Patients with Neuroimmunological Diseases)from the Ministry of Health Labour and Welfare of Japanand a Grant-in-Aid for Scientific Research (KAKENHI15K09347) from JSPS

DisclosureN Uchio K Taira C Ikenaga M Kadoya A Unuma KYoshida S Nakatani-Enomoto Y Hatanaka Y Sakuraiand Y Shiio report no disclosures K Kaida is an associateeditor of Clinical and Experimental Neuroimmunology AKubota and T Toda report no disclosures J Shimizu re-ceived research support from Grants-in-Aid for ScientificResearch Full disclosure form information provided bythe authors is available with the full text of this article atNeurologyorgNN

10 Neurology Neuroimmunology amp Neuroinflammation | Volume 6 Number 2 | March 2019 NeurologyorgNN

Publication historyReceived by Neurology Neuroimmunology amp NeuroinflammationSeptember 4 2018 Accepted in final form October 24 2018

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neoplasia infection and autoimmune disorders Acta Neuropathol 198257221ndash2292 van de Warrenburg BP Hengstman GJ Vos PE Boerman RH ter Laak HJ van

Engelen BG Concomitant dermatomyositis and myasthenia gravis presenting withrespiratory insufficiency Muscle Nerve 200225293ndash296

3 Pestronk A Kos K Lopate G Al-Lozi MT Brachio-cervical inflammatory myopathiesclinical immune and myopathologic features Arthritis Rheum 2006541687ndash1696

4 Suzuki S Utsugisawa K Yoshikawa H et al Autoimmune targets of heart and skeletalmuscles in myasthenia gravis Arch Neurol 2009661334ndash1338

5 Venna N Gonzalez RG Zukerberg LR Case 39-2011 a woman in her 90s withunilateral ptosis N Engl J Med 20113652413ndash2422

6 Hassel B Gilhus NE Aarli JA Skogen OR Fulminant myasthenia gravis and poly-myositis after thymectomy for thymoma Acta Neurol Scand 19928563ndash65

7 Otton SH Standen GR Ormerod IEC T cell lymphocytosis associated with poly-myositis myasthenia gravis and thymoma Clin Lab Haematol 200022307ndash308

8 Antoine JC Camdessanche JP Absi L Lassabliere F Feasson L Devic disease andthymoma with anti-central nervous system and antithymus antibodies Neurology200462978ndash980

9 Kon T Mori F Tanji K Miki Y Kimura T Wakabayashi K Giant cell polymyositisand myocarditis associated with myasthenia gravis and thymoma Neuropathology201333281ndash287

10 Shah A Pace A Hilton D Househam E Weatherby S Giant cell myositis responsiveto combined corticosteroids and immunoglobulin Pract Neurol 201515456ndash459

11 SetonMWuCC Louissaint A Case 26-2013 a 46-year-old woman with muscle painand swelling N Engl J Med 2013369764ndash773

12 Gidron A Quadrini M Dimov N Argiris A Malignant thymoma associated with fatalmyocarditis and polymyositis in a 32-year-old woman with a history of hairy cellleukemia Am J Clin Oncol 200629213ndash214

13 Sasaki H Yano M Kawano O Hikosaka Y Fujii Y Thymoma associated with fatalmyocarditis and polymyositis in a 58-year-old man following treatment with carbo-platin and paclitaxel a case report Oncol Lett 20123300ndash302

14 Kimura T Fukushima S Miyashita A et al Myasthenic crisis and polymyositis in-duced by one dose of nivolumab Cancer Sci 20161071055ndash1058

15 Suzuki S Ishikawa N Konoeda F et al Nivolumab-related myasthenia gravis withmyositis and myocarditis in Japan Neurology 2017891ndash8

16 Uchio N Taira K Ikenaga C et al Granulomatous myositis induced by antindashPD-1monoclonal antibodies Neurol Neuroimmunol Neuroinflamm 20185e464

17 Liao B Shroff S Kamiya-Matsuoka C Tummala S Atypical neurological complica-tions of ipilimumab therapy in patients with metastatic melanoma Neuro Oncol201416589ndash593

18 Bohan A Peter J Polymyositis and dermatomyositis (first of two parts) N Engl J Med1975292344ndash347

19 Bohan A Peter JB Polymyositis and dermatomyositis (second of two parts) N Engl JMed 1975292403ndash407

20 Hoogendijk JE Amato AA Lecky BR et al ENMC international workshop trialdesign in adult idiopathic inflammatory myopathies with the exception of inclusionbody myositis 10ndash12 October 2003 Naarden The Netherlands Neuromuscul Dis-ord 200414377ndash345

21 Rose MR Ibm E Group W 188th ENMC international workshop inclusion bodymyositis 2ndash4 December 2011 Naarden The Netherlands Neuromuscul Disord2013231044ndash1055

22 Iannuzzi MC Rybicki BA Teirstein AS Sarcoidosis N Engl J Med 20073572153ndash2165

23 Yamaguchi Y Sakurai Y Mannen T Shimizu J Rapidly progressive polymyositis withelevated antiacetylcholine receptor antibody activity Intern Med 2000391108ndash1110

24 Maekawa R Shibuya H Hideyama T Shiio Y A case of myasthenia gravis withinvasive thymoma associated with diffuse panbronchiolitis alopecia dysgeusiacholangitis and myositis Clin Neurol 201454703ndash708

25 Suzuki S Baba A Kaida K et al Cardiac involvements in myasthenia gravis associatedwith anti-Kv14 antibodies Eur J Neurol 201421223ndash230

26 Hida A Yamashita T Hosono Y et al Anti-TIF1-γ antibody and cancer-associatedmyositis a clinicohistopathologic study Neurology 201687300ndash308

27 Gilhus NE Myasthenia gravis N Engl J Med 20163752570ndash258128 Stefanou MI Komorowski L Kade S Bornemann A Ziemann U Synofzik M A case

of late-onset thymoma-associatedmyasthenia gravis with ryanodine receptor and titinantibodies and concomitant granulomatous myositis BMCNeurol 201616172ndash174

29 van der Meulen MF Bronner IM Hoogendijk JE et al Polymyositis an over-diagnosed entity Neurology 200361316ndash321

30 Ikenaga C Kubota A Kadoya M et al Clinicopathologic features of myositis patientswith CD8-MHC-1 complex pathology Neurology 2017891060ndash1068

31 Vilela VS Prieto-Gonzalez S Milisenda JC Selva-O Callaghan A Grau JM Poly-myositis a very uncommon isolated disease clinical and histological re-evaluationafter long-term follow-up Rheumatol Int 201535915ndash920

32 Zamecnik J Vesely D Jakubicka B et al Muscle lymphocytic infiltrates in thymoma-associated myasthenia gravis are phenotypically different from those in polymyositisNeuromuscul Disord 200717935ndash942

33 Evoli A Minisci C Di Schino C et al Thymoma in patients with MG characteristicsand long-term outcome Neurology 2002591844ndash1850

34 Fukuhara M Higuchi M Owada Y et al Clinical and pathological aspects of mi-croscopic thymoma with myasthenia gravis and review of published reports J ThoracDis 201791592ndash1597

35 Nakano S Engel AGMyasthenia gravis quantitative immunocytochemical analysis ofinflammatory cells and detection of complement membrane attack complex at theend-plate in 30 patients Neurology 1993431167ndash1172

36 Shelly S Agmon-levin N Altman A Shoenfeld Y Thymoma and autoimmunity CellMol Immunol 20118199ndash202

37 Johnson DB Balko JM ComptonML et al Fulminant myocarditis with combinationimmune checkpoint blockade N Engl J Med 20163751749ndash1755

38 Murata K Dalakas MC Expression of the costimulatory molecule BB-1 the ligandsCTLA-4 and CD28 and their mRNA in inflammatory myopathies Am J Pathol 1999155453ndash460

39 Buchbinder EI Desai A CTLA-4 and PD-1 pathways similarities differences andimplications of their inhibition Am J Clin Oncol 20163998ndash106

40 Chuang WY Strobel P Gold R et al A CTLA4high genotype is associated withmyasthenia gravis in thymoma patients Ann Neurol 200558644ndash648

NeurologyorgNN Neurology Neuroimmunology amp Neuroinflammation | Volume 6 Number 2 | March 2019 11

DOI 101212NXI000000000000053520196 Neurol Neuroimmunol Neuroinflamm

Naohiro Uchio Kenichiro Taira Chiseko Ikenaga et al polymyositis pathology

Inflammatory myopathy with myasthenia gravis Thymoma association and

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