Mestroni, Luisa

Taylor, Matthew R.G.

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AJournal of the American College of Cardiology Vol. 57, No. 23, 2011
© 2011 by the American College of Cardiology Foundation ISSN 0735-1097/$36.00
Published by Elsevier Inc. doi:10.1016/j.jacc.2011.02.026EDITORIAL COMMENT
Hearing the Noise
The Challenges of Human
Genome Variation in Genetic Testing*
Luisa Mestroni, MD,
Matthew R. G. Taylor, MD, PHD
urora, Colorado
The completion of the Human Genome Project was a
landmark achievement that revealed the reference deoxyri-
bonucleic acid (DNA) sequence for our own genome.
Almost immediately, it became clear that there was no
single “reference” DNA sequence, as even the approximately
one-half dozen human DNA samples used by the Human
Genome Project contained tens of thousands of variations
(1). As clinical genetic testing becomes more mainstream
and various projects under way perform full DNA genome
sequencing in hundreds of subjects, the extent of this
genetic variation is increasingly being appreciated. It is
widely recognized that most of this variation is probably not
relevant for determining health or risk for disease and
See page 2317
collectively has been referred to as “genetic noise.” As in
much of biology, separation of the “signal” from the “noise”
can be challenging, and as molecular genetic sequencing
expands in use and in the amount of DNA that can be
sequenced in a single assay, distinguishing a diagnostic
genetic change from background genetic variation will
remain a difficult task for researchers and clinicians. Newer
DNA sequencing technology can now complete the se-
quencing of an entire human genome several times over in
a matter of days, orders of magnitude faster than the nearly
13 years required for the initial first pass done by the
Human Genome Project consortium (2). This technology,
which will shortly be widely used in clinical genetic testing,
will undoubtedly add to the difficulty of distinguishing
signal from noise.
*Editorials published in the Journal of the American College of Cardiology reflect the
views of the authors and do not necessarily represent the views of JACC or the
American College of Cardiology.
From the Cardiovascular Institute and the Adult Medical Genetics Program,
University of Colorado Denver, Aurora, Colorado. This work was supported by grant
MO1 #RR00051-1575 from the National Institutes of Health. The authors have
reported that they have no relationships to disclose.In this issue of the Journal, Kapplinger et al. (3) elo-
quently illustrate the breadth of genetic variation (the
“noise”) in an important and life-threatening genetic dis-
ease, arrhythmogenic right ventricular cardiomyopathy
(ARVC). Various investigators had previously studied
ARVC families to identify genes and screen patient cohorts
to determine the contribution of various ARVC genes to the
overall population of ARVC families. Although the screen-
ing of control populations for discovered disease-causing
mutations is fairly standard in genetic studies, comprehen-
sive DNA sequencing of controls to measure background
genetic variation usually is not undertaken. This report and
a modest number of others provide compelling data that this
evaluation in control populations should become standard
for many future genetic studies.
Kapplinger et al. (3) studied 175 subjects from the
Netherlands and the United States with confirmed diagno-
ses of ARVC. In this group of subjects, mutations were
found in 58% of cases after sequencing 5 ARVC genes
(PKP2, DSP, DSG2, DSC2, and TMEM43). Although
these mutations were not labeled by the investigators as
being definitely “pathogenic,” the evaluation criteria used
would have led to these mutations being classified as
“probably” or “presumptively” pathogenic by clinical labo-
ratories and, more importantly, by many clinicians reading
the laboratory reports. Interestingly, using the same evalu-
ation criteria for mutations, 16% of 427 healthy controls
without ARVC also had mutations, illustrating the level of
genetic “noise” and an overall frequency well beyond that
predicted assuming the low prevalence of ARVC. In con-
trols, the majority of detected mutations were missense
mutations, suggesting that many of this class of mutations
were likely benign. When the investigators turned to mu-
tations predicted to cause more substantial consequences to
the predicted ARVC protein structures (so-called radical
mutations), the prevalence of ARVC mutations dropped to
0.5% in controls yet remained as high as 43% in probands.
The criteria for “radical mutations” included in-frame and
frame-shift insertions and deletions, splice junction, and
nonsense mutations, providing some guidance in how to
interpret the likelihood of a given variant’s contributing to
disease risk. However, as missense mutations may also cause
disease, the interpretation of a novel ARVC gene missense
mutation likely requires more than just reading the muta-
tion report from the laboratory. The investigators found
that in subjects with confirmed ARVC, missense mutations
were grouped in “hot spots” in DSP and DSG2 in regions of
protein binding domains and that mutations in affected
patients occurred in highly conserved residues across spe-
cies, whereas controls’ missense mutations localized in
highly variable residues. The complexity involved in the
interpretation of these mutations suggests that although
ARVC clinical testing is accessible to many patients and
cardiologists, significant skill may be required to properly
2329JACC Vol. 57, No. 23, 2011 Mestroni and Taylor
June 7, 2011:2328–9 The Challenge of Genetic Testinginterpret mutation test results and separate pathogenic
genetic mutations from background noise.
A previous study of long-QT syndrome by Kapa et al. (4)
investigated the pathogenicity of long-QT genes (SCN5A,
KCNQ1, and KCNH2). The investigators screened a large
cohort (n  388) of definitive long-QT syndrome cases and
also studied more than 1,300 normal controls. In that case,
the “background noise” of mutations was again significant,
although at 6% less than in the current study in ARVC.
One explanation for the difference in background noise
levels is that the total number of patients and controls
sequenced for long-QT syndrome was substantially greater
than for ARVC; thus, we may expect that as more subjects
are sequenced for the ARVC genes, some of the background
mutation noise may be reclassified as rare genetic variation,
reducing the ARVC mutation noise level somewhat.
An important element missing from the study of Kap-
plinger et al. (3) was the genetic evaluation of the patients’
families. An analysis of cosegregation of a mutation within
a family can be critical to help assess the causal role of a
putative mutation. Unfortunately, this is difficult in research
studies, in which the ascertainment of large families is not
always possible. In clinical circumstances, efforts to evaluate,
recruit, and test multiple patients in a given family are even
less likely to be undertaken. In some circumstances, inves-
tigators have taken additional steps to assess mutation
pathogenicity using in vitro cellular or in vivo animal assays,
but this approach is difficult when large numbers of muta-
tions are identified and is not possible in clinical situations
when working with clinical laboratories.
Conclusions. Genetic testing continues to evolve, reveal-
ing it to be an imperfect tool that requires careful interpre-
tation before and after testing is done. Criteria for patho-
genic mutations are not convincingly settled upon and are
liable to undergo some changes as more knowledge is
gained. Indeed, the pace of clinical testing seems at times to
have moved faster and without circumspect consideration
than perhaps research efforts would dictate. Stringent crite-ria (“radical” mutations, or missense mutations located in
highly conserved and functionally important domains) and,
whenever possible, cosegregation analysis can be used when
applicable to help with genetic test result interpretation.
Furthermore, as underlined by Kapplinger et al. (3) and
other investigators (5–7), genetic tests must be integrated
in the context of an expert clinical evaluation, together
with a good family history and accurate clinical informa-
tion, as with any other diagnostic test. Until the speci-
ficity of these types of molecular genetic tests is robust
and understood, the clinical application of such tests is
probably still better performed at referral centers with
expertise in cardiovascular genetics (5–7).
Reprint requests and correspondence: Dr. Luisa Mestroni,
Cardiovascular Institute, University of Colorado, 12700 East 19th
Avenue, Mail Stop F442, Aurora, Colorado 80045-6511. E-mail:
luisa.mestroni@ucdenver.edu.
REFERENCES
1. The 1000 Genomes Project Consortium. A map of human genome
variation from population-scale sequencing. Nature 2010;467:1061–73.
2. Metzker ML. Sequencing technologies—the next generation. Nature
Rev Genet 2010;11:31–46.
3. Kapplinger JD, Landstrom AP, Salisbury BA, et al. Distinguishing
arrhythmogenic right ventricular cardiomyopathy/dysplasia–associated
mutations from background genetic noise. J Am Coll Cardiol 2011;57:
2317–27.
4. Kapa S, Tester DJ, Salisbury BA, et al. Genetic testing for long-QT
syndrome: distinguishing pathogenic mutations from benign variants.
Circulation 2009;120:1752–60.
5. MacRae CA. Closer look at genetic testing in long-QT syndrome: will
DNA diagnostics ever be enough? Circulation 2009;120;1745–8.
6. Hershberger RE, Lindenfeld J, Mestroni L, Seidman CE, Taylor MR,
Tobwin JR. Genetic evaluation of cardiomyopathy: a HFSA compre-
hensive heart failure practice guideline. J Card Fail 2009;15:83–97.
7. National Center for Biotechnology Information. GeneTests. Available
at: http://www.genetests.org. Accessed December 19, 2010.
Key Words: arrhythmogenic right ventricular cardiomyopathy y
arrhythmogenic right ventricular dysplasia y diagnosis y genetic testing
y mutation.


Hearing the Noise The Challenges of Human Genome Variation in Genetic Testing⁎⁎Editorials published in the Journal of the American College of Cardiology reflect the views of the authors and do not necessarily represent the views of JACC or the American College of Cardiology.

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Hearing the Noise The Challenges of Human Genome Variation in Genetic Testing⁎⁎Editorials published in the Journal of the American College of Cardiology reflect the views of the authors and do not necessarily represent the views of JACC or the American College of Cardiology.

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