Molecular Genetics and Metabolism 113 (2014) 8491

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Molecular Genetics and Metabolism

journal homepage: www.elsevier.com/locate/ymgme

Methods of diagnosis of patients with Pompe disease: Data from the

Pompe Registry
Priya S. Kishnani a,, Hernn M. Amartino b, Christopher Lindberg c, Timothy M. Miller d,
Amanda Wilson d, Joan Keutzer d
a
Division of Medical Genetics, Department of Pediatrics, Duke University Medical Center, Box 103856 DUMC, 4th Floor GSRBI, 595 LaSalle Street, Durham, NC 22710, USA
b
Division of Child Neurology, Department of Pediatrics, Austral University Hospital, Juan Domingo Peron 1500, Pilar (B16641NZ), Buenos Aires, Argentina
c
Neuromuscular Centre, Sahlgrenska University Hospital, S-413 45 Gothenburg, Sweden
d
Genzyme, a Sano company, 500 Kendall Street, Cambridge, MA 02142, USA

a r t i c l e i n f o a b s t r a c t

Article history: Pompe disease is a rare, autosomal recessive disorder characterized by deciency of lysosomal acid alpha-
Received 27 May 2014 glucosidase and accumulation of lysosomal glycogen in many tissues. The variable clinical manifestations,
Received in revised form 10 July 2014 broad phenotypic spectrum, and overlap of signs and symptoms with other neuromuscular diseases make
Accepted 11 July 2014
diagnosis challenging. In the past, the diagnosis of Pompe disease was based on enzyme activity assay in skin
Available online 16 July 2014
broblasts or muscle tissue. In 2004, methods for measuring acid alpha-glucosidase activity in blood were
Keywords:
published. To compare how diagnostic methods changed over time and whether they differed by geographic
Pompe disease region and clinical phenotype, we examined diagnostic methods used for 1059 patients enrolled in the
Diagnosis Pompe Registry in three onset categories (Group A: onset of signs/symptoms 12 months of age with car-
GAA enzyme assays diomyopathy; Group B: onset 12 months without cardiomyopathy and onset N1 year to 12 years;
Dried blood spots Group C: onset N 12 years). Enzyme activity-based assays were used more frequently than other diagnostic
Lysosomal storage disorder methods. Measuring acid alpha-glucosidase activity in blood (leukocytes, lymphocytes, or dried-blood
Registry spot) increased over time; use of muscle biopsy decreased. The increased use of blood-based assays for
diagnosis may result in a more timely diagnosis in patients across the clinical spectrum of Pompe disease.
2014 Elsevier Inc. All rights reserved.

1. Introduction
Pompe disease is a rare, autosomal recessive, neuromuscular disorder
caused by a deciency of the lysosomal enzyme acid alpha-glucosidase
(GAA) that results in an accumulation of glycogen in lysosomes of
many tissues, most notably skeletal, cardiac, and smooth muscle.
Progressive accumulation of glycogen in these tissues leads to clinical
debilitation, organ and system failure, and often death. Pompe disease
manifests as a broad clinical spectrum with considerable variation in
age of symptom onset, presenting signs and symptoms, and degree of se-
verity and organ involvement, including cardiomegaly, hepatomegaly,
and macroglossia [1,2]. The most severe form, classic infantile-onset
Pompe disease, has an onset of signs and symptoms in the rst few
weeks to months of life and is characterized by progressive, hypertrophic
cardiomyopathy, hypotonia, and death due to rapid cardiorespiratory
Abbreviations: ACP, acid phosphatase; CRIM, cross-reactive immunologic material;
DBS, dried blood spot; ECG, electrocardiogram; ERT, enzyme replacement therapy; GAA,
acid alpha-glucosidase; IRB/EC, Institutional Review Board or Ethics Committee; MRI,
magnetic resonance imaging; NBS, newborn screening; PAS, periodic acid-Schiff.
Corresponding author. Fax: +1 919 684 8944.
E-mail address: kishn001@mc.duke.edu (P.S. Kishnani).
failure, typically before 2 years of age [1,3,4]. Late-onset Pompe disease
presents after 12 months of age (or as late as the 6th decade of life)
and is characterized by a progressive, predominantly proximal limb
and respiratory muscle weakness that is associated with signicant mor-
bidity and mortality [1,47]. While cardiac involvement is considered a
dening feature of classic infantile-onset Pompe disease, it also has
been reported in some late-onset patients [1,2,8]. However, more recent
reports note cardiac dysfunction, suggesting that its presence has been
underreported in late-onset patients. The reported cardiac involvement
in late-onset patients is variable and includes rhythm disturbances,
such as WolffParkinsonWhite syndrome, electrocardiogram (ECG) ab-
normalities, and ascending aorta dilation [915]. Increased natural histo-
ry data and the publication of various novel clinical presentations have
expanded the known clinical spectrum of late-onset Pompe disease to in-
clude features such as arterial aneurysms, lingual weakness, oropharyn-
geal dysphagia, ptosis, and scoliosis [8,14,1620]. Enzyme replacement
therapy (ERT) is available as a specic treatment for Pompe disease. Pa-
tient outcomes improved in patients with the classic infantile-onset form
of the disease when treatment with ERT is started early for these very
young patients. Studies in patients with late-onset Pompe disease sug-
gest that treatment with ERT can lead to improved outcomes or stabiliza-
tion of disease progression [2131]. With more widespread use of

http://dx.doi.org/10.1016/j.ymgme.2014.07.014
1096-7192/ 2014 Elsevier Inc. All rights reserved.
 P.S. Kishnani et al. / Molecular Genetics and Metabolism 113 (2014) 8491
newborn screening (NBS) programs, more will be learned about the
early initiation of ERT in patients across the disease spectrum. Some
challenges do remain because of unrecognized factors in a subset of
patients who may not do well despite early start of ERT. In most cases,
however, clinical benets are maximal when treatment is initiated
early. Therefore, with the availability of ERT and reported treatment
benets, accurate and early diagnosis of Pompe disease is important for
all patients.
Demonstrating deciency of GAA enzyme activity remains a stan-
dard for diagnosing Pompe disease. Current biochemical testing
assays measure GAA enzyme activity in cultured skin broblasts,
muscle biopsy tissue, or blood samples using dried blood spots
(DBS) of whole blood on lter paper, puried lymphocytes, and
mixed leukocytes [2,3239]. Although measuring GAA activity is
relatively easy, non-invasive, and inexpensive, signicant delays in
diagnosis remain [2,9,4044]. The rarity of the disease, its variable
clinical manifestations and phenotypes, and signicant overlap of
signs and symptoms with other neuromuscular diseases, make con-
sidering Pompe disease and ordering the diagnostic test a challenge
[2,5,7,32,39].
Presented here is an overview of the diagnostic practices reported to
the Pompe Registry. The methods used to diagnose Pompe disease, the
change in use of the various diagnostic testing methods over time, and
differences in methods used in the various geographic regions and pa-
tient subgroups for patients enrolled in the Registry will be reviewed.
The ways in which these new technologies and clinical approaches are
potentially leading to earlier and more accurate diagnosis of Pompe dis-
ease specically for patients in the Registry and possibly for patients in
general are discussed.
2. Methods
2.1. The Pompe Registry
The Pompe Registry, a long-term, multinational, observational
program started in 2004 and sponsored and administered by Genzyme,
a Sano company (Cambridge, MA), was designed to develop a better
understanding of the natural history and outcomes of patients with
Pompe disease. The Registry contains the largest collection of data on
patients diagnosed with the disease. All patients with a conrmed
diagnosis (documented GAA deciency from any tissue source and/or
documentation of two GAA gene mutations) of Pompe disease, regard-
less of age, clinical manifestations, or treatment status, can be enrolled
by physician investigators worldwide. Patient participation in the
Pompe Registry is voluntary. Clinical information is reported voluntarily
to the Registry by participating physicians and healthcare team mem-
bers involved in treating enrolled patients.
Each independent site obtains a patient's informed consent to
submit his/her health information to the Registry and to use and
disclose this information in subsequent aggregate analyses (journal
articles, annual reports, education materials, and public health reports).
Historically, each independent site was responsible for determining
whether site-specic Institutional Review Board or Ethic Committee
(IRB/EC) review is required for participation in the Registry in accor-
dance with institutional policies and local laws and regulations. As of
February 2013, the Pompe Registry protocol requires sites to submit to
an IRB/EC.
2.2. Analysis description
We examined the methods used to diagnose Pompe disease in
patients enrolled in the Pompe Registry and explored how use of diag-
nostic methods has changed over time and how they differ across
geographic regions and by clinical phenotype, using a descriptive,
cross-sectional analysis. All patients enrolled in the Registry were eligi-
ble for inclusion. The time to diagnosis from onset of symptoms of
85
Pompe disease was not determined for this analysis, but is reported
elsewhere [44].
A recommended Schedule of Assessments, developed by the Pompe
Registry Board of Advisors, is provided as guidance. However, because
assessments are performed according to individual patient needs and
abilities at assessment time points and may be affected by regional clin-
ical practices and standards of care, capabilities, and availability of test-
ing resources, all patients may not have all recommended assessments
completed. Also, data may not be recorded in the Registry for all mea-
surements or assessments done for patients by their healthcare team.
These assessment and reporting practices thus account for differences
in the number of patients reported for various measures. Percentages
reported for individual measures therefore reect the percentages of
patients who have the specic data reported in the Registry for individ-
ual measures and are not percentages of the total number of patients in
the analysis.
Safety data are not collected by, or reported through, the Pompe
Registry. Healthcare providers are advised to report adverse events
and matters related to the safety of ERT directly to the Genzyme Global
Pharmacovigilance and Epidemiology Department.
Data collected through the Pompe Registry are entered into a data-
base, analyzed, and reviewed for missing data points, incomplete infor-
mation, and discrepancies with previously submitted data by members
of the Registry staff. If necessary, issues are resolved with the site. Site
visits are conducted periodically to review the quality of the data
entered into the Registry database. All data management and analyses
occur in a validated computing environment.
For this analysis, three onset categories based on patient age at rst
sign or symptom onset and evidence of cardiomyopathy (as reported
by echocardiogram, or as an enlarged heart on chest X-ray or 1
enlarged atria or ventricles on an electrocardiogram in the absence of
echocardiogram results) were identied: Group A (onset of signs or
symptoms 12 months of age with cardiomyopathy); Group B (onset
of signs or symptoms 12 months without cardiomyopathy and onset
of signs or symptoms N 1 year to 12 years); and Group C (onset of
signs or symptoms N12 years). Age at sign or symptom onset was report-
ed in months for patients in Group A and in years for Groups B and C. The
year of diagnosis for patients was grouped into pre-2000 and 2-year stra-
ta after 2000 to evaluate changes in diagnostic methods used over time.
The presenting sign or symptom class was determined by the
reported signs or symptoms occurring within 1 month of the rst
recorded symptom in patients in Group A, and within 1 year for patients
in Groups B and C.
The diagnostic methods used were categorized as follows: all possi-
ble combinations of DNA, enzyme activity, and other testing, or as
unknown/missing for patients without a reported method of diagnosis
or for whom the method was unknown. The enzyme diagnostic cate-
gory includes GAA enzyme activity testing methods that are reported as
DBS, other blood-based (lymphocyte and leukocyte), broblast, muscle,
or unknown/missing. Other includes all write-in responses that could
not be recoded to DNA or enzyme analysis, such as muscle biopsy for
histologic examination, which involves the microscopic evaluation of
muscle tissue to identify histologic characteristics of abnormal glycogen
accumulation. This differs from enzyme assays that measure GAA
enzyme activity in a sample of muscle tissue [2]. Patients may have
had more than one type of assay used in reaching a diagnosis and, there-
fore, could have more than one assay type reported.
For analysis of single versus multiple methods of diagnosis, single
refers to use of a single enzyme assay method, DNA analysis only, or a
single reported other method only, and multiple includes any com-
bination of enzyme activity assay, DNA analysis, or other methods or
more than one enzyme methods.
We used descriptive statistics to analyze data according to demo-
graphic and clinical characteristics of patients in the Registry who
were included in this analysis. Percentages were calculated for the
diagnostic methods used among patients for each independent
86 P.S. Kishnani et al. / Molecular Genetics and Metabolism 113 (2014) 8491

variable. All analyses were conducted using SAS 9.1 (SAS Institute
Inc., Cary, NC, USA).
3. Results
3.1. Study population
As of March 2012, 1059 patients were enrolled in the Pompe Regis-
try (Table 1). Nearly one-third (29.3%, n = 310) of the patients in the
Registry were diagnosed with Pompe disease prior to 2000. The number
of Registry patients diagnosed each year since 2000 showed a general
upward trend. Most patients were from Europe (58.8%, 623/1059) and
the United States (27.5%, 291/1059); 75.3% (797/1059) of all patients
were Caucasian. The distribution of patients by gender was nearly
equal. Of the 1059 enrolled patients, 14.3% (151/1059) were categorized
into onset category Group A, 13.9% (147/1059) into Group B, and 51.1%
(541/1059) into Group C. Onset category was unknown/missing for
20.8% (220/1059) of patients. Musculoskeletal signs and symptoms
were the most frequent presenting sign and symptom class and were
reported for 45.2% (479/1059) of patients. Only 8.5% (90/1059) of all
patients enrolled in the Registry rst presented with respiratory signs
and symptoms.
3.2. Methods used for diagnosis and geographic distribution
Method of diagnosis was reported for 98.4% (1042/1059) of the
patients in the Registry. Overall, enzyme-based assays were used for
diagnosis in 83.1% (880/1059) of patients. More than one-third (34.5%,
365/1059) of patients were diagnosed with enzyme assays alone.
Table 1
Characteristics of patients enrolled in the Pompe Registry.
Enzyme assays were used for diagnosis for the majority of patients,
regardless of geographic region (Fig. 1), ranging from 79.1% (493/623)
for patients enrolled in Europe to 94.7% (44/47) for patients in the
Asia-Pacic region. DNA methods were used for diagnosis in 45.2%
(479/1059) of all patients and were the second most frequently used di-
agnostic method for all geographic regions. Muscle biopsy for histologic
evaluation was reported less often than enzyme- and DNA-based
methods for all patients, although this may be an artifact of how the
data were collected and recorded in the Pompe Registry, possibly
reecting the fact that data for assessments are not always or consis-
tently entered into the Registry and may not always accurately report
actual clinical practices.
While all methods of diagnosis were used in all years, those measur-
ing enzyme activity were always the most frequently used (Fig. 2).
There was a general increasing trend over time for the use of enzyme
activity assays, with the exception of 20042005 (81.3% of patients)
compared with 20022003 (87.0%). Enzyme activity assays were used
in 94.5% (154/163) of patients diagnosed in 20082009. The use of
DNA diagnostic methods also showed a slight upward trend over time.
DNA assays remained the second most frequently used method of diag-
nosis for all years after 2000 and were used for 59.5% of the patients
diagnosed during 20082009. As the use of DNA and enzyme methods
increased over time, the use of histologic evaluation of muscle biopsy
specimens for diagnosis showed a general decreasing trend.
No substantial differences were seen in the use of enzyme or DNA
methods to diagnose patients across the three onset categories
(Fig. 3). Enzyme assays were the most frequently used diagnostic
method in all three groups. DNA testing was done for more than
one-third of patients in each onset category. Patients in Group A,
the youngest onset category, were less likely to have undergone
muscle biopsy for histologic examination for diagnosis (10.6%) than
patients in Group B (36.7%) or Group C (25%).

Patients 3.3. Use of single methods versus multiple methods for diagnosis
N %

Total 1059 100.0 Multiple methods were used for diagnosis in some patients in all
Region onset categories, which is in keeping with general recommendations
Asia Pacic 57 5.4 that more than one diagnostic be used to conrm and ensure accurate
Europe 623 58.8
diagnosis [2,32]. In recent years (20082009), multiple methods of
Latin America 88 8.3
USA 291 27.5 diagnosis were used more often in Groups A (68.4%) and C (68.3%)
Year of diagnosis compared with Group B (53.3%). The combined use of both DNA and
Pre-2000 310 29.3
20002001 68 6.4
20022003 92 8.7
20042005 128 12.1
20062007 162 15.3
20082009 163 15.4
20102011 67 6.3
Unknown/missing 69 6.5
Onset category
Group A: 12 months with cardiomyopathy 151 14.3
Group B: 12 months without cardiomyopathy, and N12 months 147 13.9
to 12 years
Group C: N12 years 541 51.1
Unknown/missing 220 20.8
Presenting symptom class
Respiratory 90 8.5
Musculoskeletal 479 45.2
Non-respiratory/non-musculoskeletal 42 4.0
Respiratory and musculoskeletal concurrently 222 21.0
Unknown/missing 226 21.3
Gender
Male 541 51.1
Female 518 48.9
Ethnicity
Caucasian 797 75.3
Black 36 3.4
Hispanic 45 4.2
Asian 67 6.3
Fig. 1. Diagnostic methods used for patients in the Pompe Registry by geographic region.
Other 37 3.5
(Note: Patients may have more than one method of diagnosis reported in the Pompe
Unknown/missing 77 7.3
Registry.)
 P.S. Kishnani et al. / Molecular Genetics and Metabolism 113 (2014) 8491
Fig. 2. Diagnostic methods used for patients in the Pompe Registry based on year of diagnosis. (Note: Patients may have more than one method of diagnosis reported in the Pompe
Registry.)
enzyme assays together in patients for diagnosis showed a general up-
ward trend over time, from 25.5% (pre-2000) to 54% (20082009)
(Fig. 4).
3.4. Enzyme activity diagnostic methods
In patients who had been diagnosed with an enzyme activity assay,
use of blood-based assays (leukocyte/lymphocyte and DBS combined)
generally increased over time starting in 2000, from 35.6%
(20002001) to 77.9% (20082009) of patients (Fig. 5). Use of DBS in-
creased during recent years, most notably during 20062007 (26.2%)
and 20082009 (31.8%) as compared with pre-2000 (5.1%). Conversely,
use of both skin broblasts and muscle tissue for diagnostic enzyme ac-
tivity assays showed overall decreasing trends.
Fig. 3. Diagnostic methods used for patients in the Pompe Registry by onset category.
(Group A: onset of signs and symptoms 12 months of age with cardiomyopathy;
Group B: onset of signs and symptoms 12 months of age without cardiomyopathy
and onset N12 months to 12 years of age; and Group C: onset of signs and
symptoms N12 years of age. Note: Patients may have more than one method of
diagnosis reported in the Pompe Registry.)
87
Among the 880 patients diagnosed with methods based on enzyme
activity, blood-based assays using leukocytes/lymphocytes were the
most frequently used method in all three onset categories (Fig. 6). DBS
was used more often in Group A (27.8%) compared with Groups B
(9.1%) and C (16.2%). Enzyme assays using muscle tissue were used
more often in patients in later onset categories (29.8% in Group B and
34.5% in Group C) compared with only 8.3% in Group A. Enzyme assays
using broblasts, which require prolonged time for tissue culturing,
were used more frequently in Group A (33.1%) than in Group B
(26.4%) and Group C (22.4%). Some regional differences in the types of
blood-based assays used were identied. For patients diagnosed with
blood-based methods, assays measuring GAA activity in leukocytes
and lymphocytes were used more than twice as often as DBS assays
for diagnosing patients in the Asia-Pacic region (67.4% vs. 32.6%,
respectively) and Latin America (67.3% vs. 32.7%, respectively), and
nearly nine times more frequently for patients in Europe (89.6% vs.
10.4%, respectively), whereas in the United States, DBS was used in
72% of patients diagnosed with blood-based assays compared with
28% of patients diagnosed with leukocyte and lymphocyte assays.
Fig. 4. Combined use of both DNA and enzyme assays together for patients in the Pompe
Registry for diagnosis by year of diagnosis. (Note: Patients may have more than one meth-
od of diagnosis reported in the Pompe Registry.)
88 P.S. Kishnani et al. / Molecular Genetics and Metabolism 113 (2014) 8491

90 Pre-2000 (n=217) 2000-2001 (n=59) 2002-2003 (n=80)

81.2
80 77.9 2004-2005 (n=104) 2006-2007 (n=149) 2008-2009 (n=154)

70

Percent of Patients
60 52.6
53
51.9
50 47.5
42.4 41.9
39
40 35.6
28.8 30
28.1
30 26
22.8 22.8
20.1
20
11.7 11.5 10.4
9.2
10 5.1 5.4 6.8 6.7
2.8 2.5 3.8 3.8
0
0
Blood-Based* Fibroblast Muscle Other Unknown/Missing
Diagnostic Method

Fig. 5. Use of enzyme activity-based assays for diagnosis over time. (Note: Patients may have more than one method of diagnosis reported in the Pompe Registry. *Blood-based methods
include both leukocyte/lymphocyte and DBS assay methods. The percentage breakdowns of patients who had each method [either leukocyte/lymphocyte assay or DBS assay, respectively],
for each time period in the gure are as follows: Pre-2000: 47.9% and 5.1%; 20002001: 30.5% and 5.1%; 20022003: 46.3% and 6.3%; 20042005: 47.1% and 4.8%: 20062007: 55.0% and
26.2%; 20082009: 46.1% and 31.8%.)

4. Discussion have been conrmed [38,45]. The raised awareness of the availability
of these tests that resulted from educational symposia and initiatives
Results of this analysis indicate that physicians who have enrolled supported by Genzyme also may have contributed to the increased
patients in the Pompe Registry used enzyme activity assays for diagnos- use of blood-based assays and the sequence of diagnostic tests. In
ing patients more frequently than other available diagnostic methods. recognition of the need for more widespread adoption of blood-
The use of blood-based assays that measure GAA activity in leukocytes, based testing to provide for more accurate and timely diagnosis of pa-
lymphocytes, or DBS increased for Registry patients over time. tients with Pompe disease and potential limited availability, resources,
As the overall use of blood-based assays increased, the use of skin and training for accurate testing, particularly DBS, Genzyme requested
broblast and muscle tissue for enzyme testing decreased, which is that all patients with Pompe disease in Genzyme-sponsored clinical
not unexpected in light of the more convenient, less invasive, and trials voluntarily donate blood samples for DBS assay development
less expensive nature of the blood-based assays compared to the purposes. Patient consent was nearly 100%. Detailed protocol and
tissue-based enzyme methods. In the past, GAA enzyme activity technical assistance, and in some instances grant support, were
assays using cultured skin broblasts from skin biopsies or tissue made available to participating laboratories worldwide by Genzyme.
from muscle biopsy samples were commonly used for diagnosis DBS methods provide samples that are relatively stable for ship-
because reliable methods using blood samples were not available. ping and thus offer a diagnostic means for patients with limited
However, results from collecting skin and muscle samples, which access to specialty laboratories. Test results can be obtained relative-
requires biopsy and tissue culture in the case of skin, can take several ly quickly and are inexpensive [36,43,46]. Skin broblast testing
weeks [2,3335,42,45]. The increased use of blood-based assays continues to remain important, however, especially for infantile
measuring GAA enzyme activity may reect the acceptance of these patients, where this is the only available method to determine if
assays as reliable methods of diagnosis, since validity and accuracy patients produce any molecular GAA species, termed cross-reactive

Fig. 6. Use of enzyme activity based assays for diagnosis by onset category. (Group A: onset of signs and symptoms 12 months of age with cardiomyopathy; Group B: onset of signs and
symptoms 12 months of age without cardiomyopathy and onset N12 months to 12 years of age; and Group C: onset of signs and symptoms N12 years of age. Note: Patients may have
more than one diagnostic method reported in the Pompe Registry.)
 P.S. Kishnani et al. / Molecular Genetics and Metabolism 113 (2014) 8491
immunological material (CRIM) positive, which may be signicant in
determining treatment response. An assay that can determine CRIM
status using blood has been developed [47].
Blood-based methods, particularly DBS, also allow for testing of large
numbers of samples from patients with variable clinical ndings. This is
important because many late-onset patients remain undiagnosed or
labeled with the non-specic diagnosis of limb girdle muscle dystrophy
for many years [34]. Although blood-based assays using DBS were 100%
sensitive and specic when the samples were collected in a clinical trial
[36], the validity of DBS testing as the sole method of routine diagnosis
continues to be investigated [45,48]. The possibility of false-positive
results with DBS should also be considered. The c.1726 G N A
[p.G576S] pseudodeciency allele is prevalent in Asia and has led to
false-positive results in NBS pilot programs in Asia [4951]. This may
complicate results of enzymatic diagnostic assays as a sole means of
establishing the diagnosis [49]. Results of blood-based testing using
DBS therefore should be interpreted carefully and within the context
of clinical symptoms. A number of studies have demonstrated the valid-
ity of using blood-based assays for diagnosis of patients of all ages. How-
ever, conrmatory testing with more than one diagnostic method is
generally recommended to ensure accurate diagnosis [2,22,32,36,39,
42,43,46,49,52,53]. The analysis here showed that multiple methods,
as recommended, are commonly used for diagnosing patients in the
Pompe Registry.
While enzyme activity-based assays were the most commonly
used diagnostic tools for Registry patients, other methods, including
DNA analysis and muscle biopsy for histologic examination, also
were reported. The increase in use of DNA methods for diagnosis
was somewhat surprising and may be indicative of an increasing
role of mutation analysis in the diagnosis of Pompe disease as the
number of laboratories capable of performing such testing increases.
Mutation analysis of the GAA gene may be helpful in conrming the
biochemical diagnosis, which is particularly important for patients
with late-onset Pompe disease who may have relatively high residu-
al GAA enzyme activity [35,46]. It also plays a very important role in
predicting CRIM status in infantile patients in cases where the geno-
type and correlation with CRIM status are known. However, DNA
analysis is not the preferred rst method for diagnosis because the
gene locus is heterogeneous. Often, only one previously identied
mutation is found coupled with a novel, unclassied sequence vari-
ant of unknown signicance [2,33,46,54,55].
There was a relatively frequent use of histologic evaluation of muscle
biopsy specimens for diagnosis, particularly in Group B at 36.7%. The fact
that patients in Group B have a reported symptom onset at a young age
but without cardiomyopathy may result in ambiguous presentations
necessitating broad differential diagnoses that lead to an increased use
of this more invasive diagnostic testing. Historically, histologic exami-
nation of muscle biopsy specimens has played a signicant role in the
diagnosis of patients presenting with limb girdle weakness and can be
useful in identifying many conditions within the differential diagnosis
of late-onset Pompe disease [39,56]. Unfortunately, several publications
have now identied that histologic evaluation of muscle biopsy samples
may not provide reliable diagnosis or exclusion of Pompe disease due to
several factors, including heterogeneous muscle involvement and lyso-
somal accumulation of glycogen between and within muscles (particu-
larly in juveniles and adult with Pompe disease), variation in the
location and visibility of abnormal glycogen by light microscopy, and
the possibility of error in preparing currently used staining techniques
[7,39,46]. Magnetic resonance imaging (MRI) before muscle biopsy
has been proposed as a means of identifying and selecting suitably af-
fected muscles for biopsy [57]. However, because abnormal muscle
characteristics seen on MRI images may be representative of other
neuromuscular disorders and not specic to Pompe disease, a correla-
tion between MRI ndings and Pompe disease has not been denitively
established and can potentially lead to unnecessary muscle biopsies.
Although disease progression and muscle changes in patients with a
89
conrmed diagnosis of Pompe disease can be monitored with MRI
over time, the value and usefulness of MRI as a diagnostic tool combined
with muscle biopsy needs to be studied further [58]. While potentially
useful for identifying many conditions included within the differential
diagnosis of patients presenting with late-onset Pompe disease, use of
histologic examination of muscle biopsy is not recommended as the
sole tool for conrming or excluding a diagnosis of Pompe disease, espe-
cially in late-onset patients, in whom a diagnosis of Pompe disease may
be missed [7,32,39,43,46]. Muscle biopsy performed in young patients
with Pompe disease, especially those with concomitant cardiac abnor-
malities, is cautioned as the procedure itself typically requires anesthe-
sia, posing additional risk to patients and adding to the burden of
disease [2,21,46]. It therefore is actively discouraged in patients with
this clinical presentation. Fortunately, newer, less invasive assays pro-
vide reliable, alternative diagnostic tools.
Several staining techniques have also shown promise as potential
screening methods for a diagnosis of Pompe disease [59,60]. Abnormal
cytoplasmic vacuolation of lymphocytes occurs in many lysosomal stor-
age disorders, including Pompe disease, most notably in classic
infantile-onset patients and less frequently in milder forms of the dis-
ease seen in juvenile and adult patients. These vacuolated lymphocytes
can be identied by routine blood lm examination (prepared from a
simple standard blood draw) and stain positive with periodic acid-
Schiff (PAS) reagent [61]. Because this staining appears to be specic
to Pompe disease and is relatively quick, inexpensive, and minimally in-
vasive, evaluating blood lms for PAS-positive lymphocyte vacuoles can
be a clinically useful screening tool in the differential diagnosis of
Pompe disease, especially for infants with the disease [59,61]. In older
patients with late-onset disease, characteristic vacuolated muscle bers
typically seen in Pompe disease may not always be present. Some pa-
tients with Pompe disease have been found to have unique cytoplasmic
globular inclusions that stain positive to acid phosphatase (ACP) and
not PAS, which also appear to be specic to Pompe disease. As suggested
by Tsuburaya et al. [60], these ACP-positive globular inclusions could be
potential hallmarks of Pompe disease and may be a useful diagnostic
marker of Pompe disease in adult-onset patients who lack typical vacu-
olated bers.
Findings from this analysis of patients enrolled in the Pompe Reg-
istry suggest that recent advances in diagnostic capabilities and lab-
oratory methodology are being used to diagnose patients with this
progressive, debilitating neuromuscular disorder. The increased use
of blood-based assays for diagnosis is encouraging and implementa-
tion of initiatives such as NBS programs will assist with a more time-
ly diagnosis in patients across the clinical spectrum of Pompe disease
[23,28,62]. Although not within the scope of this study, future analy-
ses of Registry data will be needed to evaluate if patients diagnosed
with blood-based assays have shorter gaps between symptom
onset and diagnosis of Pompe disease. The Pompe Registry contains
the largest collection of Pompe patients worldwide and is a valuable
resource for helping the medical community understand Pompe
disease. Clinicians' understanding of the natural history of Pompe
disease tends to be incomplete, and data obtained from the Registry
contribute substantially to the clinical understanding of this rare
condition, as evident from published analyses generated from the
Pompe Registry [17,63]. The Pompe Registry will continue to play
an important role as a means of collecting, understanding, and
disseminating clinical information about Pompe disease globally.
Disclosures
The Pompe Registry is sponsored by Genzyme, a Sano company
(Cambridge, MA).
Priya S. Kishnani has received research/grant support and honoraria
from Genzyme and is a member of the Pompe and Gaucher Registry
Advisory Boards for Genzyme.
90 P.S. Kishnani et al. / Molecular Genetics and Metabolism 113 (2014) 8491
Hernn M. Amartino has received honoraria and support for travel
from Genzyme and is a member of the Pompe Registry Advisory Board
for Genzyme.
Christopher Lindberg has received honoraria and support for travel
from Genzyme and is a member of the Pompe Registry Advisory Board
for Genzyme.
Joan Keutzer is an employee of Genzyme.
Timothy M. Miller and Amanda Wilson are former employees of
Genzyme.
Acknowledgments
The authors would like to thank the patients and their families for
volunteering their medical information to the Pompe Registry; the clini-
cians and their staffs who collect and enter data into the Registry; and
the following individuals for their assistance with the development of
this manuscript: Marianne B. Zajdel, a Senior Medical Writer contracted
by Genzyme, and Cinde Clatterbuck, Project Manager, Genzyme.
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