- Email: [email protected]
Challenges in the Diagnosis of Anderson-Fabry Disease
JOURNAL OF THE AMERICAN COLLEGE OF CARDIOLOGY
VOL. 68, NO. 10, 2016
ª 2016 BY THE AMERICAN COLLEGE OF CARDIOLOGY FOUNDATION
ISSN 0735-1097/$36.00
PUBLISHED BY ELSEVIER
http://dx.doi.org/10.1016/j.jacc.2016.06.026
EDITORIAL COMMENT
Challenges in the Diagnosis of Anderson-Fabry Disease A Deceptively Simple and Yet Complicated Genetic Disease* Ali J. Marian, MD
A
nderson-Fabry disease (AFD), a pleiotropic
into the diagnostic work-up and clinical management
lysosomal storage disease, is a masquerader
of patients with AFD.
wearing
multiple
cloaks.
It
commonly
In accord with the aforementioned pleiotropic
mimics hypertrophic cardiomyopathy, expressing it-
manifestations, Favalli et al. (7) report in this issue of
self as left ventricular hypertrophy without outflow
the Journal the results of a 10-year multidisciplinary,
tract obstruction (1–3). It often manifests with angio-
multicenter evaluation of 2,034 patients for AFD.
keratoma, corneal deposits, renal insufficiency with
Phenotypic characterization included organ-specific
proteinuria, and less commonly with peripheral neu-
clinical testing and sequencing of the GLA gene to
ropathy and cerebrovascular events (4,5). The pleio-
detect pathogenic variants. Additionally, a -Gal enzy-
tropic manifestations often delay the diagnosis,
matic activity was measured in the majority of
emphasizing the need for a comprehensive multidis-
participants and organ histology examined in some.
ciplinary approach to evaluate and manage patients
In all, 37 probands (1.8%) and 64 family members
with or at high risk of AFD.
carried pathogenic variants in the GLA gene. Clini-
AFD is caused by loss-of-function (LoF) mutations
cally, cardiac involvement, manifesting as left ven-
in the GLA gene, which encodes alpha-galactosidase
tricular hypertrophy with a wall thickness of >13 mm
A ( a -Gal), an enzyme responsible for hydrolysis of
(hypertrophic cardiomyopathy phenocopy), was the
a-D-galactose residues in glycosphingolipids (6).
most common finding, occurring in about one-half of
Mutations
the affected individuals, followed by acroparesthesia
affect
synthesis,
trafficking,
folding,
degradation, and enzymatic activity of a -Gal, result-
and renal insufficiency. The findings emphasized the
ing in accumulation of globotriaosylceramide (GB3),
need for a systemic approach to increase the diag-
the main a -Gal substrate, in the lysosomes of multiple
nostic yield of screening tests in AFD.
organs. Consequently, genetic, biochemical, and his-
SEE PAGE 1037
tological testing (whenever possible) are incorporated
AFD is the diagnostic epitome of single-gene disorders, as various diagnostic tools—including genetic testing,
biochemical
assays,
and
histological
*Editorials published in the Journal of the American College of Cardiology
examination—are available to validate the clinical
reflect the views of the authors and do not necessarily represent the
diagnosis. Yet, an accurate pre-clinical diagnosis of
views of JACC or the American College of Cardiology.
this seemingly straightforward genetic disease, orig-
From the Center for Cardiovascular Genetics, Brown Foundation Institute
inally described more than a century ago, has
of Molecular Medicine, The University of Texas Health Science Center
remained challenging (8). Genetic screening by
and Texas Heart Institute, Houston, Texas. Dr. Marian is supported in part by grants from National Institutes of Health; National Heart, Lung,
deoxyribonucleic acid sequencing to identify patho-
and Blood Institute (R01 HL088498, 1R01HL132401, and R34 HL105563);
genic variants offers the most pragmatic and desir-
Leducq Foundation (14 CVD 03); Roderick MacDonald Foundation
able first approach. Yet, one has to recognize that
(13RDM005); TexGen Fund from Greater Houston Community Founda-
unambiguous identification of the causal variant in a
tion; Texas Heart Institute Foundation; and George and Mary Josephine Hamman Foundation. P.K. Shah, MD, served as Guest Editor for this
proband or a small family is exceedingly challenging,
paper.
if not impossible (9). Of course, no gene is perfect and
1052
Marian
JACC VOL. 68, NO. 10, 2016 SEPTEMBER 6, 2016:1051–3
Diagnosis of Fabry Disease
the GLA gene is no exception, even though it is highly
activity overlaps with normal values, diminishing the
intolerant to the LoF variant. Considering this intol-
diagnostic utility of a -Gal activity assays. Genetic
erance, identification of such a variant—defined as
testing, performed by whole exome sequencing and
nonsense, splice acceptor, and splice donor variants
targeted analysis of the GLA gene, could increase
caused by single nucleotide changes—in the GLA gene
confidence in the biochemical and clinical diagnosis if
should isolate the genetic cause of AFD.
it leads to identification of a pathogenic variant.
In contrast, GLA also carries several missense and
Nevertheless, the findings of a missense variant
typically rare variants that are present in the general
should not be considered an unequivocal validation
population and, hence, should not be considered
of the diagnosis.
causal
variants.
For
example,
the
p.Asp313Tyr
Alpha-Gal activity in female subjects, who carry a
variant, which has been reported as a causal variant
heterozygous pathogenic variant in the GLA gene, is
for AFD, has a frequency of 0.4% in the non-Finnish
subject to X chromosome inactivation, which is
European population and is several orders of magni-
typically random, cell type–dependent, and often
tude greater than prevalence of AFD in that popula-
nonuniform across the silenced chromosome (12). The
tion. Therefore, the p.Asp313Tyr variant is not
process could lead to mosaic inactivation and escape
considered a disease-causing variant, even though
of the GLA gene from inactivation in the germ or
bioinformatics tools predict it to be “probably
somatic cells. The complexity of X chromosome
damaging” and “deleterious” (10). Further compli-
inactivation further compounds interpretation of
cating the genetic landscape is the potential influence
GLA-variant functional data, relevance of a -Gal ac-
of genetic backgrounds on functional and phenotypic
tivity measured in the blood or white blood cells to
consequences of a specific variant, which might
other organs, and, hence, clinical significance of the
render the variant pathogenic in some but not other
findings. Likewise, it complicates correlations among
genetic backgrounds. Whereas detection of a LoF
the genetic variants, functional data, and organ
variant strongly indicates causality, detection of a
involvement. Nevertheless, as a group, a -Gal activity
rare missense variant is insufficient to conclude cau-
is higher in female subjects with the pathogenic GLA
sality or evidence of genetic validation for the clinical
variants than the corresponding values in male sub-
diagnosis of AFD. Consequently, overreliance on ge-
jects. Consequently, a -Gal activity in female subjects
netic screening alone, even when focused on patho-
who carry GLA variants often overlaps with normal
genic variants, risks overdiagnosing AFD. Commonly used with genetic testing, biochemical
values in about one-third of cases, posing significant diagnostic challenges (5).
assays test the activity of the a -Gal enzyme in the
Accumulation of a -Gal substrates in various cells
blood or white blood cells and measure plasma levels
and organs in a heterogeneous manner produces the
of the storage products GB3 and globotriaosyl-
clinical phenotype in AFD. Consequently, measure-
sphingosine (Lyso-GB3), the latter a GB3 degradation
ment of GB3 and Lyso-GB3 levels in biological speci-
product. Using genetic testing with biochemical
mens could facilitate AFD diagnosis (13). Plasma (and
assays also helps discern functional significance of
urinary) levels of GB3 and Lyso-GB3 are sensitive
genetic variants, increasing each assay’s overall
biomarkers, albeit not without limitations, which are
diagnostic utility. Biochemical assays, however, have
more pronounced in female subjects and in the early
their own limits (11); factors such as pH, temperature,
stages of AFD (8). Likewise, the assays may be
endogenous substances including free hemoglobin,
insufficiently specific (8). Nevertheless, these bio-
and additional galactosidases (5) can affect the
markers are emerging as useful diagnostic tools as
in vitro assay readouts of a -Gal activity. Thus, perti-
well
nent laboratories must establish and report assay
replacement therapy in AFD.
as
for
assessing
effectiveness
of
enzyme
performance metrics and consider various factors
Consistent with effects of genetic variants on
that might influence the readouts, including biolog-
downstream phenotypes (14–16), GLA variant effects
ical sample quality.
range from negligible to severe reduction of a -Gal
As GLA is located on Xq22.1 chromosome locus, sex
activity (17). Typically, a -Gal activity of less than one-
chromosomes compound the effect of the genetic
third of the mean normal value is considered patho-
variants on a -Gal activity. Male individuals with a LoF
genic in AFD (8). However, a true biologic threshold
variant typically exhibit very low or undetectable
may not exist and each functional variant might
a-Gal activity and, thus, a phenotype well in accord
impose phenotypic effects, whether biological, his-
with classic AFD. Consequently, diagnostic accuracy
tological, or clinical. Detecting the phenotypic effects
of exceedingly low and undetectable a -Gal activity is
depends on the resolution of the screening tools.
quite high. In some male subjects, however, a -Gal
Similarly, one might speculate that a -Gal activity
Marian
JACC VOL. 68, NO. 10, 2016 SEPTEMBER 6, 2016:1051–3
Diagnosis of Fabry Disease
measured in vitro correlates closely with its substrate
seemingly simple yet complicated storage disease
utilization in vivo and in various organs. Whether
requires a multifaceted approach integrating various
there is a critical threshold for reduced a -Gal activity
available diagnostic modalities. Large datasets, such
in a given organ beyond which substrate utilization is
as the one reported by Favalli et al. (7), offer glimpses
impaired, producing substrate accumulation, remains
into the power of an integrated approach in detecting
to be determined. A linear correlation between
and evaluating patients at risk of AFD.
reduced a -Gal activity and substrate accumulation in vivo in a specific organ would pose the intriguing
REPRINT REQUESTS AND CORRESPONDENCE: Dr.
hypothesis that phenotypic expression of AFD is a
A.J. Marian, Center for Cardiovascular Genetics, The
continuum, wherein clinically manifested pheno-
Brown Foundation Institute of Molecular Medicine,
types reflect the spectrum’s extreme end.
The University of Texas Health Sciences Center, 6770
Considering each specific diagnostic test’s shortcomings, an accurate and early diagnosis of this
Bertner Street, Suite C900A, Houston, Texas 77030. E-mail: [email protected].
REFERENCES 1. Kozor R, Grieve SM, Tchan MC, et al. Cardiac involvement in genotype-positive Fabry
7. Favalli V, Disabella E, Molinaro M, et al. Genetic screening of Anderson-Fabry disease in probands
13. Aerts JM, Groener JE, Kuiper S, et al. Elevated globotriaosylsphingosine is a hallmark of Fabry
disease patients assessed by cardiovascular MR. Heart 2016 [Epub ahead of print].
referred from multispecialty clinics. J Am Coll Cardiol 2016;68:1037–50.
disease. Proc Natl Acad Sci U S A 2008;105: 2812–7.
2. Linhart A, Kampmann C, Zamorano JL, et al., for the European FOS Investigators. Cardiac manifestations of Anderson-Fabry disease: results from the international Fabry outcome survey. Eur Heart J 2007;28:1228–35.
8. Schiffmann R, Fuller M, Clarke LA, Aerts JM. Is it Fabry disease? Genet Med 2016 May 19 [E-pub ahead of print].
14. Marian AJ. Nature’s genetic gradients and the clinical phenotype. Circ Cardiovasc Genet 2009;2: 537–9.
9. MacArthur DG, Manolio TA, Dimmock DP, et al. Guidelines for investigating causality of sequence
3. Monserrat L, Gimeno-Blanes JR, Marin F, et al. Prevalence of Fabry disease in a cohort of
variants in human disease. Nature 2014;508:469–76.
15. Marian AJ. Causality in genetics: the gradient of genetic effects and back to Koch’s postulates of causality. Circ Res 2014;114:e18–21.
508 unrelated patients with hypertrophic cardiomyopathy. J Am Coll Cardiol 2007;50: 2399–403.
10. Oder D, Uceyler N, Liu D, et al. Organ manifestations and long-term outcome of Fabry disease in patients with the GLA haplotype D313Y. BMJ Open 2016;6:e010422.
4. Thomas AS, Hughes DA. Fabry disease. Pediatr Endocrinol Rev 2014;12 Suppl 1:88–101.
11. Andrade J, Waters PJ, Singh RS, et al. Screening for Fabry disease in patients with chronic kidney disease: limitations of plasma alpha-galactosidase
5. Elstein D, Altarescu G, Beck M. Fabry Disease. New York, NY: Springer, 2010:181.
assay as a screening test. Clin J Am Soc Nephrol 2008;3:139–45.
6. Romeo G, Migeon BR. Genetic inactivation of the alpha-galactosidase locus in carriers of Fabry’s disease. Science 1970;170:180–1.
12. Berletch JB, Yang F, Disteche CM. Escape from X inactivation in mice and humans. Genome Biol 2010;11:213.
16. Marian AJ, Belmont J. Strategic approaches to unraveling genetic causes of cardiovascular diseases. Circ Res 2011;108:1252–69. 17. Lukas J, Scalia S, Eichler S, et al. Functional and clinical consequences of novel alphagalactosidase A mutations in Fabry disease. Hum Mutat 2016;37:43–51.
KEY WORDS alpha-galactosidase A, causality, genetic testing, hypertrophic cardiomyopathy, loss of function
1053
VOL. 68, NO. 10, 2016
ª 2016 BY THE AMERICAN COLLEGE OF CARDIOLOGY FOUNDATION
ISSN 0735-1097/$36.00
PUBLISHED BY ELSEVIER
http://dx.doi.org/10.1016/j.jacc.2016.06.026
EDITORIAL COMMENT
Challenges in the Diagnosis of Anderson-Fabry Disease A Deceptively Simple and Yet Complicated Genetic Disease* Ali J. Marian, MD
A
nderson-Fabry disease (AFD), a pleiotropic
into the diagnostic work-up and clinical management
lysosomal storage disease, is a masquerader
of patients with AFD.
wearing
multiple
cloaks.
It
commonly
In accord with the aforementioned pleiotropic
mimics hypertrophic cardiomyopathy, expressing it-
manifestations, Favalli et al. (7) report in this issue of
self as left ventricular hypertrophy without outflow
the Journal the results of a 10-year multidisciplinary,
tract obstruction (1–3). It often manifests with angio-
multicenter evaluation of 2,034 patients for AFD.
keratoma, corneal deposits, renal insufficiency with
Phenotypic characterization included organ-specific
proteinuria, and less commonly with peripheral neu-
clinical testing and sequencing of the GLA gene to
ropathy and cerebrovascular events (4,5). The pleio-
detect pathogenic variants. Additionally, a -Gal enzy-
tropic manifestations often delay the diagnosis,
matic activity was measured in the majority of
emphasizing the need for a comprehensive multidis-
participants and organ histology examined in some.
ciplinary approach to evaluate and manage patients
In all, 37 probands (1.8%) and 64 family members
with or at high risk of AFD.
carried pathogenic variants in the GLA gene. Clini-
AFD is caused by loss-of-function (LoF) mutations
cally, cardiac involvement, manifesting as left ven-
in the GLA gene, which encodes alpha-galactosidase
tricular hypertrophy with a wall thickness of >13 mm
A ( a -Gal), an enzyme responsible for hydrolysis of
(hypertrophic cardiomyopathy phenocopy), was the
a-D-galactose residues in glycosphingolipids (6).
most common finding, occurring in about one-half of
Mutations
the affected individuals, followed by acroparesthesia
affect
synthesis,
trafficking,
folding,
degradation, and enzymatic activity of a -Gal, result-
and renal insufficiency. The findings emphasized the
ing in accumulation of globotriaosylceramide (GB3),
need for a systemic approach to increase the diag-
the main a -Gal substrate, in the lysosomes of multiple
nostic yield of screening tests in AFD.
organs. Consequently, genetic, biochemical, and his-
SEE PAGE 1037
tological testing (whenever possible) are incorporated
AFD is the diagnostic epitome of single-gene disorders, as various diagnostic tools—including genetic testing,
biochemical
assays,
and
histological
*Editorials published in the Journal of the American College of Cardiology
examination—are available to validate the clinical
reflect the views of the authors and do not necessarily represent the
diagnosis. Yet, an accurate pre-clinical diagnosis of
views of JACC or the American College of Cardiology.
this seemingly straightforward genetic disease, orig-
From the Center for Cardiovascular Genetics, Brown Foundation Institute
inally described more than a century ago, has
of Molecular Medicine, The University of Texas Health Science Center
remained challenging (8). Genetic screening by
and Texas Heart Institute, Houston, Texas. Dr. Marian is supported in part by grants from National Institutes of Health; National Heart, Lung,
deoxyribonucleic acid sequencing to identify patho-
and Blood Institute (R01 HL088498, 1R01HL132401, and R34 HL105563);
genic variants offers the most pragmatic and desir-
Leducq Foundation (14 CVD 03); Roderick MacDonald Foundation
able first approach. Yet, one has to recognize that
(13RDM005); TexGen Fund from Greater Houston Community Founda-
unambiguous identification of the causal variant in a
tion; Texas Heart Institute Foundation; and George and Mary Josephine Hamman Foundation. P.K. Shah, MD, served as Guest Editor for this
proband or a small family is exceedingly challenging,
paper.
if not impossible (9). Of course, no gene is perfect and
1052
Marian
JACC VOL. 68, NO. 10, 2016 SEPTEMBER 6, 2016:1051–3
Diagnosis of Fabry Disease
the GLA gene is no exception, even though it is highly
activity overlaps with normal values, diminishing the
intolerant to the LoF variant. Considering this intol-
diagnostic utility of a -Gal activity assays. Genetic
erance, identification of such a variant—defined as
testing, performed by whole exome sequencing and
nonsense, splice acceptor, and splice donor variants
targeted analysis of the GLA gene, could increase
caused by single nucleotide changes—in the GLA gene
confidence in the biochemical and clinical diagnosis if
should isolate the genetic cause of AFD.
it leads to identification of a pathogenic variant.
In contrast, GLA also carries several missense and
Nevertheless, the findings of a missense variant
typically rare variants that are present in the general
should not be considered an unequivocal validation
population and, hence, should not be considered
of the diagnosis.
causal
variants.
For
example,
the
p.Asp313Tyr
Alpha-Gal activity in female subjects, who carry a
variant, which has been reported as a causal variant
heterozygous pathogenic variant in the GLA gene, is
for AFD, has a frequency of 0.4% in the non-Finnish
subject to X chromosome inactivation, which is
European population and is several orders of magni-
typically random, cell type–dependent, and often
tude greater than prevalence of AFD in that popula-
nonuniform across the silenced chromosome (12). The
tion. Therefore, the p.Asp313Tyr variant is not
process could lead to mosaic inactivation and escape
considered a disease-causing variant, even though
of the GLA gene from inactivation in the germ or
bioinformatics tools predict it to be “probably
somatic cells. The complexity of X chromosome
damaging” and “deleterious” (10). Further compli-
inactivation further compounds interpretation of
cating the genetic landscape is the potential influence
GLA-variant functional data, relevance of a -Gal ac-
of genetic backgrounds on functional and phenotypic
tivity measured in the blood or white blood cells to
consequences of a specific variant, which might
other organs, and, hence, clinical significance of the
render the variant pathogenic in some but not other
findings. Likewise, it complicates correlations among
genetic backgrounds. Whereas detection of a LoF
the genetic variants, functional data, and organ
variant strongly indicates causality, detection of a
involvement. Nevertheless, as a group, a -Gal activity
rare missense variant is insufficient to conclude cau-
is higher in female subjects with the pathogenic GLA
sality or evidence of genetic validation for the clinical
variants than the corresponding values in male sub-
diagnosis of AFD. Consequently, overreliance on ge-
jects. Consequently, a -Gal activity in female subjects
netic screening alone, even when focused on patho-
who carry GLA variants often overlaps with normal
genic variants, risks overdiagnosing AFD. Commonly used with genetic testing, biochemical
values in about one-third of cases, posing significant diagnostic challenges (5).
assays test the activity of the a -Gal enzyme in the
Accumulation of a -Gal substrates in various cells
blood or white blood cells and measure plasma levels
and organs in a heterogeneous manner produces the
of the storage products GB3 and globotriaosyl-
clinical phenotype in AFD. Consequently, measure-
sphingosine (Lyso-GB3), the latter a GB3 degradation
ment of GB3 and Lyso-GB3 levels in biological speci-
product. Using genetic testing with biochemical
mens could facilitate AFD diagnosis (13). Plasma (and
assays also helps discern functional significance of
urinary) levels of GB3 and Lyso-GB3 are sensitive
genetic variants, increasing each assay’s overall
biomarkers, albeit not without limitations, which are
diagnostic utility. Biochemical assays, however, have
more pronounced in female subjects and in the early
their own limits (11); factors such as pH, temperature,
stages of AFD (8). Likewise, the assays may be
endogenous substances including free hemoglobin,
insufficiently specific (8). Nevertheless, these bio-
and additional galactosidases (5) can affect the
markers are emerging as useful diagnostic tools as
in vitro assay readouts of a -Gal activity. Thus, perti-
well
nent laboratories must establish and report assay
replacement therapy in AFD.
as
for
assessing
effectiveness
of
enzyme
performance metrics and consider various factors
Consistent with effects of genetic variants on
that might influence the readouts, including biolog-
downstream phenotypes (14–16), GLA variant effects
ical sample quality.
range from negligible to severe reduction of a -Gal
As GLA is located on Xq22.1 chromosome locus, sex
activity (17). Typically, a -Gal activity of less than one-
chromosomes compound the effect of the genetic
third of the mean normal value is considered patho-
variants on a -Gal activity. Male individuals with a LoF
genic in AFD (8). However, a true biologic threshold
variant typically exhibit very low or undetectable
may not exist and each functional variant might
a-Gal activity and, thus, a phenotype well in accord
impose phenotypic effects, whether biological, his-
with classic AFD. Consequently, diagnostic accuracy
tological, or clinical. Detecting the phenotypic effects
of exceedingly low and undetectable a -Gal activity is
depends on the resolution of the screening tools.
quite high. In some male subjects, however, a -Gal
Similarly, one might speculate that a -Gal activity
Marian
JACC VOL. 68, NO. 10, 2016 SEPTEMBER 6, 2016:1051–3
Diagnosis of Fabry Disease
measured in vitro correlates closely with its substrate
seemingly simple yet complicated storage disease
utilization in vivo and in various organs. Whether
requires a multifaceted approach integrating various
there is a critical threshold for reduced a -Gal activity
available diagnostic modalities. Large datasets, such
in a given organ beyond which substrate utilization is
as the one reported by Favalli et al. (7), offer glimpses
impaired, producing substrate accumulation, remains
into the power of an integrated approach in detecting
to be determined. A linear correlation between
and evaluating patients at risk of AFD.
reduced a -Gal activity and substrate accumulation in vivo in a specific organ would pose the intriguing
REPRINT REQUESTS AND CORRESPONDENCE: Dr.
hypothesis that phenotypic expression of AFD is a
A.J. Marian, Center for Cardiovascular Genetics, The
continuum, wherein clinically manifested pheno-
Brown Foundation Institute of Molecular Medicine,
types reflect the spectrum’s extreme end.
The University of Texas Health Sciences Center, 6770
Considering each specific diagnostic test’s shortcomings, an accurate and early diagnosis of this
Bertner Street, Suite C900A, Houston, Texas 77030. E-mail: [email protected].
REFERENCES 1. Kozor R, Grieve SM, Tchan MC, et al. Cardiac involvement in genotype-positive Fabry
7. Favalli V, Disabella E, Molinaro M, et al. Genetic screening of Anderson-Fabry disease in probands
13. Aerts JM, Groener JE, Kuiper S, et al. Elevated globotriaosylsphingosine is a hallmark of Fabry
disease patients assessed by cardiovascular MR. Heart 2016 [Epub ahead of print].
referred from multispecialty clinics. J Am Coll Cardiol 2016;68:1037–50.
disease. Proc Natl Acad Sci U S A 2008;105: 2812–7.
2. Linhart A, Kampmann C, Zamorano JL, et al., for the European FOS Investigators. Cardiac manifestations of Anderson-Fabry disease: results from the international Fabry outcome survey. Eur Heart J 2007;28:1228–35.
8. Schiffmann R, Fuller M, Clarke LA, Aerts JM. Is it Fabry disease? Genet Med 2016 May 19 [E-pub ahead of print].
14. Marian AJ. Nature’s genetic gradients and the clinical phenotype. Circ Cardiovasc Genet 2009;2: 537–9.
9. MacArthur DG, Manolio TA, Dimmock DP, et al. Guidelines for investigating causality of sequence
3. Monserrat L, Gimeno-Blanes JR, Marin F, et al. Prevalence of Fabry disease in a cohort of
variants in human disease. Nature 2014;508:469–76.
15. Marian AJ. Causality in genetics: the gradient of genetic effects and back to Koch’s postulates of causality. Circ Res 2014;114:e18–21.
508 unrelated patients with hypertrophic cardiomyopathy. J Am Coll Cardiol 2007;50: 2399–403.
10. Oder D, Uceyler N, Liu D, et al. Organ manifestations and long-term outcome of Fabry disease in patients with the GLA haplotype D313Y. BMJ Open 2016;6:e010422.
4. Thomas AS, Hughes DA. Fabry disease. Pediatr Endocrinol Rev 2014;12 Suppl 1:88–101.
11. Andrade J, Waters PJ, Singh RS, et al. Screening for Fabry disease in patients with chronic kidney disease: limitations of plasma alpha-galactosidase
5. Elstein D, Altarescu G, Beck M. Fabry Disease. New York, NY: Springer, 2010:181.
assay as a screening test. Clin J Am Soc Nephrol 2008;3:139–45.
6. Romeo G, Migeon BR. Genetic inactivation of the alpha-galactosidase locus in carriers of Fabry’s disease. Science 1970;170:180–1.
12. Berletch JB, Yang F, Disteche CM. Escape from X inactivation in mice and humans. Genome Biol 2010;11:213.
16. Marian AJ, Belmont J. Strategic approaches to unraveling genetic causes of cardiovascular diseases. Circ Res 2011;108:1252–69. 17. Lukas J, Scalia S, Eichler S, et al. Functional and clinical consequences of novel alphagalactosidase A mutations in Fabry disease. Hum Mutat 2016;37:43–51.
KEY WORDS alpha-galactosidase A, causality, genetic testing, hypertrophic cardiomyopathy, loss of function
1053