Prevalence of Late-Onset Pompe Disease in Portuguese Patients with Diaphragmatic Paralysis - DIPPER Study

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www.revportpneumol.org

ORIGINAL

ARTICLE

Prevalence

of

late-onset

pompe

disease

in

Portuguese

patients

with

diaphragmatic

paralysis

---

DIPPER

study

M.J.

Guimarães

a,∗

,

J.C.

Winck

b

,

B. Conde

c

,

A. Mineiro

d

,

M. Raposo

e

,

J. Moita

f

,

A. Marinho

g

_,

_J.M.

_Silva

h

_,

_N.

_Pires

i

_,

_S.

_André

j

_,

_C.

_Loureiro

f

a_Hospital_Guimarães,_Centro_Hospitalar_do_Alto_Ave,_Guimarães,_Portugal b_Universidade_do_Porto,_Porto,_Portugal

c_Centro_Hospitalar_de_{Trás-os-Montes}_e_Alto_Douro,_Vila_Real,_Portugal

d_Hospital_de_Santa_Marta,_Centro_Hospitalar_de_Lisboa_Central,_Lisboa,_Portugal e_Hospital_Egas_Moniz,_Centro_Hospitalar_de_Lisboa_Ocidental,_Lisboa,_Portugal f_Centro_Hospitalar_e_{Universitário}_de_Coimbra,_Coimbra,_Portugal

g_Centro_Hospitalar_de_São_João,_Porto,_Portugal

h_Hospital_Sousa_Martins,_Unidade_Local_de_Saúde_da_Guarda,_Guarda,_Portugal i_Hospital_de_Santa_Maria_Maior,_Barcelos,_Portugal

j_Centro_Hospitalar_do_Porto,_Porto,_Portugal

Received26July2016;accepted15February2017 Availableonline9May2017

KEYWORDS

Diaphragmatic; Paralysis; Pompe

Abstract Pompediseaseisarareautosomalrecessiveneuromusculardisordercausedbyacid ␣-glucosidaseenzyme(GAA)deficiencyanddividedinto twodistinctvariants,infantile-and late-onset.Thelate-onsetvariantischaracterizedbyaspectrumofphenotypicvariationthat may rangefromasymptomatic,toreduced musclestrengthand/ordiaphragmaticparalysis. Since musclestrength lossis characteristic ofseveraldifferent conditions, whichmay also causediaphragmaticparalysis,aprotocolwas createdtosearchfor thediagnosis ofPompe diseaseandexcludeotherpossiblecauses.

Methods:Wecollectedasamplesizeof18patients(10females,8males)withamedianage of60yearsanddiagnosisofdiaphragmaticparalysisofunknownetiology,followedinthe Pul-monologyoutpatientconsultationof9centersinPortugal,overa24-monthstudyperiod.We evaluateddatafrompatient’sclinicalanddemographiccharacteristicsaswellas complemen-tarydiagnostictestsincludingbloodtests,imaging,neurophysiologicandrespiratoryfunction evaluation.AllpatientswereevaluatedforGAAactivitywithDBS(driedbloodtest)orserum quantificationandpositiveresultsconfirmedbyserumquantificationandsequencing.

Results:ThreepatientswerediagnosedwithPompe’sdiseaseandrecommendedforenzyme replacementtherapy.TheprevalenceofPompe,araredisease,inourdiaphragmaticparalysis patientsamplewas16.8%.

∗_{Corresponding}_author.

E-mailaddress:zezaguimaraes@gmail.com(M.J.Guimarães). http://dx.doi.org/10.1016/j.rppnen.2017.02.004

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Conclusion: WeconcludethatDBStestforGAAactivityshouldberecommendedforallpatients withdiaphragmaticparalysiswhich,despitelookingatallthemostcommoncauses,remains ofunknownetiology;thiswouldimproveboththetimingandaccuracyofdiagnosisforPompe diseaseinthispatientpopulation.Accuratediagnosiswillleadtoimprovedcareforthisrare, progressivelydebilitatingbuttreatableneuromusculardisease.

http://creativecommons.org/licenses/by-nc-nd/4.0/).

Introduction

Pompediseaseisarareautosomalrecessiveneuromuscular disorder caused byacid ␣-glucosidase enzyme (GAA) defi-ciency, resulting in the accumulation of glycogen in the lysosomesofnumerous,butprimarilymuscular,tissues.1_It

isoftentermedglycogenstoragediseasetypeII(GSDII)or acidmaltasedeficiency.2,3

Depending onthe age of onset,Pompe is divided into infantileandlate-onset4_disease._Infantile_onset_Pompe

dis-easerepresentsthemostsevereformandalmostinvariably leads to death, due to cardio-respiratory failure, within oneyear.Thelate-onsetvariantpresentsatanytimeafter the age of one year, and is characterized by a spectrum of phenotypicvariation. It mayrange fromasymptomatic patients with increased creatine kinase (CK) to muscle crampsandpainsyndromeorrigid-spinesyndrome.5---7_The

lowerlimbsandparaspinalmusclesarefrequentlyaffected first,followedbytherespiratorymuscles, particularlythe diaphragm, intercostal and accessory muscles. Respira-tory failure is themain causeof increased morbidity and mortality8,9_and_the_main_cause_of_respiratory_failure_is

dia-phragmaticweakness.10

Thediaphragmisthemajormuscleofventilation consti-tutedby a dome-shaped structure of tendonsand muscle innervated by the phrenic nerves which provide sensory, sympathetic and motor function. Diaphragm contraction expands the chest increasing pleural negative pressure and promotingair flow into the lungs.Whenever thereis diaphragmatic weakness the diaphragm fails to contract appropriately,causingreducedinspiratoryvolumeand pos-sibledyspnea.Anextremeformofdiaphragmaticweakness isunilateral orbilateraldiaphragmaticparalysis.11 _Several

different conditions may present with respiratory failure and distinguishingamongthem is important todetermine thecorrecttherapeuticoptions.12 _Pompe _disease_is_in _the

differential diagnosis of a wide variety of myopathies, and therefore accurate diagnostic tools are needed for an effectivescreeningof thiscondition.Particularly when the progressive involvement and weakness of respiratory muscles anddiaphragm, characteristicofthe disease,are closely related to respiratory dysfunction which affects sleep,dailylifeactivities,andoverallqualityoflife.

The prevalence of this condition in the general pop-ulation is unknown, and varies according to clinical presentationandethnicity.Thelate-onsetformhasan esti-mated incidence of 1/57000.13 _In _Portugal, _there _is _no

accuratedataontheprevalenceofPompedisease,butitis estimatedthatthereare27casesofpatientsunderenzyme replacementtherapy(ERT).14---16

Therefore, the primary objective of this study was to estimatethe prevalence of this disorder in patients with adiagnosisofdiaphragmaticparalysisofunknownetiology followedintheoutpatientsettingofaPulmonologyclinic, and to characterize the clinical and socio-demographic profileofthesepatients.Asecondaryobjectivewasto cre-atean algorithm for the accurate diagnosis of late-onset Pompe disease in patients with diaphragmatic paralysis of unknown etiology, which will lead to better manage-ment and treatment of patients with this neuromuscular disease.

Materials

and

methods

This was a national, multicenter, epidemiological study of patientswith a diagnosis of diaphragmatic paralysis of unknownetiology.Thestudypopulationwasidentifiedfrom the Pulmonology outpatient consultations of 9 centers in Portugalovera24-monthstudyperiod.Patientswere con-secutively enrolled if they were ≥18 years of age, gave informedconsenttoparticipation,andfulfilledoneofthe followinginclusioncriteria:diagnosisof(unilateralor bilat-eral)diaphragmaticparalysis of unknowncause; diagnosis of restrictive lung disease (both FVC [forced vital capac-ity] and TLC [total lung capacity]<80% of predicted, or ≥12% decrease in VC [vital capacity] in the supine posi-tion), decreased maximal inspiratory pressure (IPmax) or sniffnasal inspiratorypressure (SNIP) (at least −10cm of H2O than predicted) of unknown origin; diagnosis of

pro-gressive chronic myopathy with respiratory involvement, particularlyofthediaphragm,witheitherinconclusive mus-clebiopsyoradiagnosisbasedonthegenericdefinitionof inclusionbodymyositis.

Patients were excluded from the study if they were unableorunwillingtoprovideinformedconsent,were tra-cheostomized or pregnant, required invasive mechanical ventilation, or suffered from one of the following con-ditions: neuromuscular junction diseases (Eaton---Lambert syndrome, myasthenia gravis), myopathies (polymyositis and other mixed connective tissue diseases, dystrophy mitochondrialmyopathies,amyloidosis),spinalcord myelo-pathy (cervical spine injury, sarcoidosis, syringomyelia, polyomyelitis, amyotrophic lateral sclerosis), peripheral neuropathy [cervical spine injury, mediastinal tumor, Guillain---Barrésyndrome,nutritionalneuropathies(vitamin B12deficiency)and leadneuropathy] or activeneoplastic disease.

Information retrieved from patients included socio-demographicandclinicalhistory,aswellasphysicalexams

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Table1 Patients’socio-demographiccharacteristics.

n(%)

Totalno.ofpatients 18(100)

Male 8(44)

Female 10(56)

Medianage,years(range) 60(43---81)

Race Caucasian 18(100) Region Braga 7(38.9) Porto 4(22.0) Lisbon 3(16.7) Coimbra 1(5.6) Guarda 1(5.6) Leiria 1(5.6) VilaReal 1(5.6)

and diagnostic parameters (including respiratory function tests,polysomnography,ultrasoundscan,chest radiograph andbloodbiochemistry),allcollectedaspartoftheroutine clinicalpracticefordiaphragmaticparalysisdiagnosis.Acid ␣-glucosidaseactivitywasdeterminedindried-bloodspots (DBS) for thediagnosis ofPompe disease,17 _and _a_further

confirmationthroughGAAgenesequencingwasdoneifthe blood-basedGAAenzymeassayshowedreducedactivity.18

Respiratoryfunctiontests

Respiratory function tests were conducted using body plethysmography with vital capacity determination in seated versus supine position. The single-breath method wasusedtomeasure thealveolo-capillary diffusion ofCO (DLCO-SB)and thoracic pressures wereascertained either by maximal inspiratory pressure (MIP) or by sniff nasal inspiratorypressure(SNIP)followedbymaximalexpiratory pressure(MEP).

Statisticalanalysis

Statistical analysis was performed using SPSS 15.0 soft-ware. Continuous variables were described as medians and interquartile ranges, and categorical variables were describedasnumbersand(proportionswith)percentages.A multivariateanalysiswasperformedtoevaluatethe associ-ationbetweendiaphragmaticparalysisandmusclestrength losscharacteristicofPompedisease.

Results

A total of 18 patients with diaphragmatic paralysis were includedintheDIPPERstudybythecollaborativePortuguese centers.Patients’ socio-demographicbaseline characteris-ticsarereportedinTable1.Eight(44%)patientsweremale and10(56%)werefemale,themedianagewas60.8±11.1 (range43---81)years,allpatientswereCaucasianandmainly fromtheNorthofPortugal.

PatientclinicalhistoryissummarizedinTable2.Briefly, mostpatients(27.8%)weregrade3(walksslowerthanmost

Table2 Patients’clinicalcharacteristics.

n(%) MRCdyspneascale 1 4(22.2) 2 3(16.7) 3 5(27.8) 4 2(11.1) Daytimesymptoms Anxiety 8(44.4) Irritability 5(27.8) Headache 6(33.3) Nocturnalsymptoms Anxiety 6(33.3) Snoring 8(44.4) Insomnia 6(33.3) Non-restorativesleep 7(38.9) Personalhistory Comorbidities(hypertension, depression,diabetes,dyslipidemia, hypothyroidism) 14(77.8) Historyoftrauma 5(27.8) Previoussurgeries 8(44.4) Familyhistory Neuromuscularconditions 2(11.1) Metabolicdiseases(diabetes,

hypertriglyceridemia,hypothyroidism, dyslipidemia)

4(22.2)

Obstructivesleepapneasyndrome 1(5.6) Arterialhypertension 3(16.7) Suddendeath 2(11.1)

peopleonthelevel,stopsafteramileorso,orstopsafter 15minwalkingatownpace)intheMRCdyspneascale,and 57.1%experienced anxietyasthemaindaytimesymptom. Excessivedaytime sleepiness(EDS)wasmeasured through theEpworthsleepinessscale(ESS)withamedianvalue of 7 andaninterquartilerangeof 4---13,the maximum value fortheEpworthscaleattainedbyanypatientwas19. Sev-eralnocturnalsymptomswerereported,snoring(61.5%)and non-restorativesleep(53.8%)beingthemostfrequent. Arte-rialhypertensionwasthemostfrequenthealthprecedent, andmetabolicdiseasesthemostfrequentfamilyprecedent (n=4,22.2%).

Physicalexamination

Overallphysicalstatuswasassessedin14patientsby empir-icalobservationwith9(64.3%)patientsreportedashaving goodphysicalfunction,and5(35.7%)reportedtohave mod-estphysicalfunction.Anthropometricsofthesepatientsare depictedinTable3.

TheMallampatiscore,usedtopredicttheeaseof intu-bation,wasassessedin13patientsandshowedthatmostof them(n=5,38.5%)wereclass2(softpalate,fauces,portion of uvula), 4 (30.8%) patients were class 3 (soft palate, baseofuvula),3(23.1%)patientswereclass1(softpalate, fauces,uvula,pillars),andonly1(7.7%)patientwasclass4 (hardpalateonly).

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Table3 Anthropometricsofthe14patientswhoperformed thephysicalexamination.

Median(range)

Weight(kg) 72.5(48.0---119.0)

Height(cm) 160.0(144.0---175.0)

Bodysurface(m2₎ _1.7_(1.4---2.0)

Bodymassindex,BMI(kg/m2₎ _29.0_{(19.0---37.0)}

Bloodpressure(mmHG)

Max 129.0(105.0---158.0) Min 80.0(53.0---104.0)

Oxygensaturation(%) 94.0(84.0---97.0)

Fractionofinspiredoxygen(FiO2) 21.0(21.0---28.0)

Cervicalperimeter(m) 35.5(18.0---45.0)

Neurologic evaluation revealed that most patients had difficultygettingup(n=8,61.5%)andsitting(n=7,53.8%). Concerning upper limbs, cases of muscular atrophy were observed in 3 (23.1%) patients, mainly in arms, forearms and hands, 8 (72.7%) patients had sustained muscular strength,and10(76.9%)patientshadnormalreflexes. Con-cerning lower limbs, muscular atrophy wasobserved in 3 (23.1%) patients,9 (69.2%)patients had sustained muscu-larstrength,and9(69.2%)patientshadnormalreflexes.An abnormalgaitwasidentifiedin 7(58.3%)patients, mainly duetoataxiaandweaknessinbothfeet(n=2,28.6%,each), but alsodue to Trendelenburgwalk in onepatient, para-paresisinonepatient,andasaconsequenceofbimalleolar osteoporoticfractureinonepatient.Nine(69.2%)patients wereunable torun,with main causes beingfatigue, dys-pnea,andsequelsfrombimalleolarosteoporoticfracture.

Complementarydiagnostictests

Fifteenofthe18patientsincludedinthisstudyperformed complementarydiagnostictests,includingcompleteblood count(CBC)andbloodbiochemistry(Table4),blood sero-logy andthyroid function assessment. For88.9% of these patients, thyroid function was within the normal range. OnepatienthadpositiveserologyforHBV,andanotherfor HCV.Immunoglobulinswerewithinthenormalrangefor13 patientsandwereabnormalfortwopatients,andabnormal levelsofautoantibodieswerealsoobservedintwopatients. Creatine kinase (CK) levels were elevated in these patients,witha meanCK of249.8 (SD=332.9)U/L,and a medianof152(range:24---1290)U/L.

ThechestX-rayrevealedabnormalresultsin13patients, dueto(right,leftorboth)hemidiaphragmelevation.Inall patientsdiaphragmaticparalysiswereconfirmedbythoracic CTscan(Fig.1)orthoracicultrasound(Fig.2)andchanges indiaphragmaticmovementascertained.

Respiratoryfunctiontests

Respiratoryfunctiontestsweremandatorytoassesswhether there was a deterioration of patients’ respiratory condi-tion.Theymaybeindicative,forinstance,ofdiaphragmatic weakness by showing a difference between forced vital capacity(FVC)insittingandsupineposition---i.e.postural

Table4 Complementarydiagnostictestresultsforthe15 patientswhoperformedthem.

Median(range) CBC Hemoglobin(g/100ml) 13.6(10.1---16.8) Leukocytecount 6725(1160---11400) Neutrophilcount 3820(824---4964) Lymphocytecount 1425(209---1750) Plateletcount 237(160---2390) Bloodbiochemistry Sodium(mmol/L) 141(135---147) Creatinine(mg/dl) 0.7(0.5---5.5) Potassium(mmol/L) 4.4(3.4---5.0) Glucose(mg/DL) 102.5(81---156) Protein(g/L) 70(58---75.4) Albumin(g/L) 42(35---62.2) Creatinekinase(CK)(IU/L) 152(24---1290) Calcium(mmol/L) 8.8(2.3---9.9) Phosphorus(mmol/L) 3.2(0.9---34) Alkalinephosphatase 63.5(34---117) Gamma-glutamyltransferase (GGT)(IU/L) 28(8---209) Alanineaminotransferase(ALT)

(IU/L)

32(6---105) Aspartateaminotransferase

(AST)(IU/L)

25.5(15---94) Totalbilirubin(␮mol/L) 9.2(3.4---23.7) Directbilirubin(␮mol/L) 0.2(0.18---0.21) Totalcholesterol(mg/dL) 228.5(119---319) Thyroglobulin(ng/dL) 105.5(2.3---291) Sedimentationvelocity(S) 10(5---80) Angiotensin-convertingenzyme (ACE)(unit/L) 15(3---67)

IU/L=internationalunits/liters;S=svedbergunit.

drop---orbyadecreasedmeaninspiratorypressure(MIP). Spirometrywasusedtomeasurehowmuchandhowquickly patientscouldmoveairoutoftheirlungs,andlungfunction valuesobtainedareshowninTable5,aswellasthenumber ofpatientsinwhichthisevaluationwascollected.

Arterialbloodgasanalysiswasperformed atrest(FiO2:

21%). Median PaO2 was 64.5 (52.0---92.0)mmHg, median

PaCO2 was 45.0 (30.0---65.2)mmHg, median O2

satura-tion was 92.5 (84.5---97.1)%, and median H2CO3 was 28.8

(23.5---36.0)meq/L.

Sevenpatientsperformedthe6-min walk(6MWDT)test usingpulseoxymetry(SpO2)monitors.Six-minwalkdistance

test(6MWDT)wasdefinedasthelongestdistancepossiblein 6MWDTwithoutencouragement.Overall,patientswalkeda meandistanceof334(SD=129.8)mandhadameanfallof oxygensaturationof7.1%[91.4%beforeand84.3%afterthe test].

Polysomnography(PSG)test

PSGresultsareshowninTable6.

Electromyography was preformed in 6 patients and revealednormal results for 4patients and alteredresults fortheother2.

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Figure1 ThoracicCTscanshowingrighthemidiaphragmaticparalysis.

Figure2 ThoracicultrasoundofpatientwithrightdiaphragmaticparalysislaterconfirmedasPompe.Thereisdecreasedright diaphragmmovement(A)andnormalleftdiaphragmaticmovementamplitude(B).

Table5 Respiratoryfunctiontests(PFTs)results. Median(range) n FEV1,l/s 1.5(0.48---2.67) 12 FEV1,% 47(30.0---84.0) 12 FVC 1.7(0.72---3.25) 12 FVC,% 46.9(29.0---90.0) 13 DropinFVCfrom seatedtosupine position,% 30.0(13.0---60.0) 10 TLC,l 3.3(2.55---4.27) 5 TLC,% 72.0(52.9---90.0) 9 RV 1.9(1.12---2.0) 7 RV,% 92.2(35.0---104.0) 8 Ratio 82.0(69.0---98.4) 9 DLCO-SB,% 67.9(27.8---120.0) 10 IPmax,% 35.0(32.8---50.6) 11 EPmax,% 76.0(30.1---135.0) 11 SNIP,% 30.5(20---41) 2

FEV1=forced expiratory volume in one second; FVC=forced vital capacity; TLC=total lung capacity; RV=residual vol-ume; DLCO-SB=single-breath diffusing capacity of the lung for CO; IP max=decreasedmaximal inspiratory pressure; EP max=decreasedmaximalexpiratorypressure;SNIP=sniffnasal inspiratorypressure;l/s=liters/second;l=liters.

Ref.46.

Table6 Polysomnography(PSG)testresults.

Median(range) n

RDI 7.1(3.1---11.0) 2 Desaturationindex,% 12.0(9.3---17.5) 5 TimewithO2saturation<90%,% 24.5(0.5---100.0) 8

Centralsleepapnea,% 15.0(10.0---20.0) 2 Obstructivesleepapnea,% 49.5(0.05---60.0) 4 Mixedapneas,% 10.0(10.0---10.0) 2 Hypopnea 15.0(9.4---61.1) 4 RERAs 0.2(0.2---0.2) 1

RDI=respiratory disturbance index; RERA=respiratory effort-relatedarousal.

Out of the 16 samples measured for dried blood spot (DBS) acid ␣-glucosidase, four tested positive for Pompe disease, a fifth patient evidenced a small reduction of acid␣-glucosidaseunconfirmedbyserumGAAactivity mea-surement, and the remaining 11 tested negative. The 2 remainingpatientsweretestedforserumGAAactivity(not DBS)andalsotestednegativeforPompe.

Discussion

Thelate-onsetformofPompediseasehasaheterogeneous presentation, mimicking other neuromuscular diseases, leadingtodiagnosticchallenge.Clinicalmanifestationsvary in terms oforganinvolvement, ageat onsetand severity,

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andsymptomsareoftenunspecificandmayremainmildfor decades, so that neitherthe patientnor the doctor con-sideredgoingfurtherwithdiagnosticprocedures.Anearly diagnosiscouldberelevantduetothechanceofimproving or at leaststabilizing thecourse of diseasethrough enzy-maticreplacementtherapy(ERT).19,20

Although Pompe disease is a rare condition, it has beenreportedinanumberofdifferentethnicpopulations, namely Caucasian, Taiwanese, Korean and Japanese.21---31

This studyisthe firsttoinvestigate theincidenceof late-onset Pompe disease in Portugal, based on respiratory signs/symptoms. One clinical study had previously inves-tigated and identified four cases of the juvenile form of the disease in Portuguese patients, raising awareness of the fact that Pompe disease should be suspected in pro-gressivemyopathiesat anyage,especiallythoseinvolving limb-girdleandrespiratorymusclesandinsmallinfantswith cardiomyopathy.29

In the present study, 18 patients with a mean age of 60.8±11.1yearsanddiaphragmatic paralysiswere identi-fiedand investigatedfor thelate-form of Pompe disease. InPortugal, thediagnosis recommendationsfor late-onset Pompediseasesuggestthatpatientswithaprogressive limb-girdleweakness,fatigue,crampsandmusclepainshouldbe evaluatedwithcreatinine kinase(CK)levels, electromyog-raphy,dynamicspirometryandmusclebiopsyininconclusive cases.16 _Suspected_cases_and_those_in_which_muscle_biopsy

couldnotallowotherdiagnosisshouldbescreenedfor lyso-somal acid-␣-glucosidasedeficiency withdried blood spot (DBS), andthe diagnosis shouldbeconfirmed by determi-nationoflysosomalacid-␣-glucosidaseactivityinasecond sample and lysosomal acid-␣-glucosidase gene sequenc-ing. According to this, 5 of the 18 patients investigated evidencedabsenceor reductionofGAAactivityintheDBS test.

Muscular weakness was a common symptom in these patients,with61.5%ofpatientsshowingdifficultyingetting upand53.8%insittingdown,inkeepingwithotherstudies thathavereportedmuscularweaknessasthemostcommon initialsymptominlate-onsetPompepatients.8,32

Laboratory analysisrevealed ameanCKvalue of249.8 (±332.9;range:24---1290)U/L,whichissuggestiveofPompe disease,since CKvaluesare typicallyelevated (>250U/L, equivalent to 1.5---15 times the upper limit of normal in adults)inallformsofthisdisease.

The exercise capacity(normallymeasured usingthe 6-min walk distance test [6MWDT]) andpulmonary function (forced vitalcapacity [FVC]) areoutcome measures com-monly used in studies of late-onset Pompe disease, by registeringchangesinthe6MWDTdistanceandinVC (mea-suredaspercentagechangeinpredicted FVC[%predicted FVC]). In this study, patients walked a mean distance of 334±129.8m, whichis inaccordancewiththe246---340m distance reported by other studies of late-onset Pompe patients.33---37_Although_the_6MWDT_is_a_valuable_and_widely

used functional measure, it is associated with consider-ableinter- andintra-investigator variability,and does not elucidate themechanismsbehind thecompromised physi-calfunctionthatitmeasures.38 _Furthermore,_the_distance

walked can be affected by factors such as patient moti-vation, age, sex, height, and weight as well as skeletal

problems,39_which_can_affect_gait_and_thereby_influence_the

distancewalked,aswenoticedinthisstudy.

FVCprovides asimple measureof pulmonary function, andisawidelyusedoutcomemeasureinstudiesofpatients withneuromusculardisorders.Becausediaphragmweakness oftenoccursearlyinthediseaseprocessandmuscle weak-nessmaynotbe evidentuntillaterin thediseasecourse, respiratoryevaluationis extremely importantfor patients who may have Pompe disease. Spirometric measurement ofFVC in both the seatedand supinepositions is manda-tory.A greater than 10% drop in FVC fromthe seated to thesupinetestingpositionissuggestiveofdiaphragm weak-nessandshouldraisethepossibilityofPompedisease.40_Both

measurementsshouldbemaderegardlessofwhether respi-ratorysymptomsarepresent.Becausepatientsmayhavea normal seated FVC, the presence of diaphragm weakness can be missed if supine FVC is not measured.13,41 _In _this

study,spirometryresultsrevealeda30%mediandropinFVC whenpatientsswitchedfromtheuprighttothesupine posi-tion,which isconcordantwiththediagnosisof diaphragm dysfunction.

Althoughnotyetevaluatedinlate-onsetPompedisease, thesniffnasalinspiratory pressure(SNIP) testallows ear-lier detection of changes in respiratory muscle strength thanFVC, andmight be a useful measureto assessthese patients.42

Inthisstudy,wecouldnotfindastatisticallysignificant correlationbetweenanyofthefunctionaltest results,for eitherpulmonaryfunctionormuscleweakness,andPompe disease,butdiaphragm involvement wasconfirmed inthe resultsforallthepatientsincluded.

The gold standard for diagnosis in Pompe disease is the determination of partial or complete deficiency of GAAenzyme activityin blood or fibroblasts. Inthis study, 18 patients with diaphragmatic paralysis were screened for Pompe disease, 2 through ␣-glucosidase serum activ-itymeasurements, 16 throughthe dried blood spot (DBS) ␣-glucosidase test, withpositive results for 5patients. In one patient Pompe disease was not confirmed by subse-quentserumactivity quantifications(whichwerenormal). InthreeofthefourpositivepatientsthediagnosisofPompe diseasewasestablishedbygene sequencing,andallwere recommendedfor enzymatic replacement therapy by the NationalCouncilforLysosomalStorageDiseases.Thefourth patient revealed only one previously identified mutation (D91N)relatedtoapseudodeficiencyofGAA,andthe diag-nosisforPompediseaseremainsunclear.

Due to the low prevalence and wide range of symp-toms,Pompe diseaseis often unrecognized byphysicians. Therefore,the development of a diagnostic methodology representsa valuable tool to enhance the recognition of Pompe disease by pulmonologists and other physicians. The American Association of Neuromuscular and Electro-diagnosticMedicine has developed an algorithm to aid in the diagnosis of Pompe disease,13 _and _other _algorithms

havebeen developed in recent yearstoallow for an ear-lier detection of late-onset Pompe disease cases.6,13,43---45

Inpatientspresenting withdiaphragmparalysisor respira-torysymptomssuggestiveofdiaphragmmuscleinvolvement, Pompe disease should be included in the differential diagnosis and patients should undergofurther laboratory,

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electrodiagnosticandpulmonary evaluations, and imaging studies,asappropriate.

AsimpleDBStestcanimproveboththetimingand accu-racyofthediagnosis,whichwillleadtoanimprovementin thecareofpatientswhohavethisprogressivelydebilitating neuromusculardisease.

Limitations

Wewerenotabletoestablishaspecific algorithmforthe accuratediagnosisoflate-onsetPompediseaseinthis pop-ulation.

AnotherlimitationofourstudywasthatthenonPompe disease patients with diaphragm paralysis did not have a standardized neurologicalevaluation;these patientswere followedbyPulmonologistonlyanditmaybethecasethat amultidisciplinaryevaluationiswarrantedforbetter char-acterization

Consideringourprimaryobjective,inthisstudywewere abletodiagnosePompediseasein3ofourpoolof18patients (16.7%).

Although results from thisstudy shouldbe interpreted withcautiondue tothesmallsample size, we wouldlike to highlight that Pompe’s Disease is rare, with only 27 estimatedcasesundergoingERTinanoverallnational popu-lationofapproximately10million,andisourfirmbeliefthat Pompeshould be suspectedin all patients with diaphrag-matic paralysis of unknown etiology and that DBS should alwaysbeperformedinthesepatients.

Ethical

responsibilities

Protectionofpeopleandanimals.Theauthorsstatethat nohumanoranimalexperimentshavebeen performedfor thisinvestigation.

Confidentialityofthedata

The authors declare that no patient data appears in this article.

Righttoprivacyandwrittenconsent

The authors declare that no patient data appears in this article.

Conflicts

of

interest

Dr.Guimarãeshavereceivedfinancialsupportforthis inves-tigationproject,travelgrantsandhonorariumsforlecturing fromSanofiGenzyme® ref. GZ-2010-10243--- for DIPPER ---DIaphragmParalysisinPompeInvestigation.

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