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Neuron
Overview
Neurogenetics: Advancing the ‘‘Next-Generation’’
of Brain Research
Huda Y. Zoghbi1,* and Stephen T. Warren2
1Departments of Molecular and Human Genetics and Pediatrics, Program in Developmental Biology, Howard Hughes Medical Institute,
Baylor College of Medicine, and Jan and Dan Duncan Neurological Research Institute at Texas Children’s Hospital, Houston, TX 77030, USA
2Departments of Human Genetics, Pediatrics and Biochemistry, Emory University School of Medicine, Atlanta, GA 30322, USA
*Correspondence: [email protected]
DOI 10.1016/j.neuron.2010.10.015
There can be little doubt that genetics has transformed our understanding of mechanisms mediating brain
disorders. The last two decades have brought tremendous progress in terms of accurate molecular diagnoses and knowledge of the genes and pathways that are involved in a large number of neurological and
psychiatric disorders. Likewise, new methods and analytical approaches, including genome array studies
and ‘‘next-generation’’ sequencing technologies, are bringing us deeper insights into the subtle complexities
of the genetic architecture that determines our risks for these disorders. As we now seek to translate these
discoveries back to clinical applications, a major challenge for the field will be in bridging the gap between
genes and biology. In this Overview of Neuron’s special review issue on neurogenetics, we reflect on progress made over the last two decades and highlight the challenges as well as the exciting opportunities for the
future.
2011 marks ten years since the release of
the first draft of the human genome, and
as often happens with anniversaries,
there has been much recent discussion,
within both the scientific community and
the general public, about what has often
been called ‘‘the genetics revolution’’
and its impact on science and medicine.
In this essay, we will outline the gains and
the challenges of neurogenetic diseaseoriented research. Considering the past
two decades of advances in neurogenetics, there has been a wealth of exciting
discoveries, but there are also opportunities to learn some lessons from failed
experiments and a chance to reflect on
challenges that might have prevented or
delayed the development of successful
interventions for some disorders.
There is no doubt that the sequencing
of the human genome has been a critical
scientific milestone that has revolutionized biology and medicine. Yet, it is
noteworthy that in the decade before the
completion of the human genome project,
neurogenetics was already on the rise.
Looking back, it is clear that many
exciting discoveries would not have
been possible without some key collaborations between astute clinicians and
technically innovative basic scientists. It
is also astounding how much these gene
discoveries have taught us not only about
particular diseases but also about basic
neurobiology. The discovery of the gene
for Duchenne muscular dystrophy (DMD)
(Koenig et al., 1987) in 1986–1987 highlights the critical role of clinical genetics,
cytogenetics, and linkage in delineating
the location of a gene (Francke et al.,
1985; Lindenbaum et al., 1979; Murray
et al., 1982). DMD was one of the first
gene discoveries for an inherited disorder,
and over the last two decades, DMD
has been a model disorder for the development of new diagnostics and therapeutics for a genetic disorder. In 1983,
mapping the gene of Huntington disease
(HD) to the short arm of chromosome 4
by using restriction fragment length polymorphisms and linkage in a large family
marked a new era, wherein a disease
gene can be mapped without any prior
knowledge from cytogenetic abnormalities (Gusella et al., 1983). Likewise, the
discovery of dinucleotide polymorphic
repeats (Gyapay et al., 1994) and the
ease of genotyping such repeats with
poylmerase chain reaction (PCR) facilitated genetic mapping and was a key
factor in uncovering duplications and
deletions of the PMP22 locus as the
cause for Charcot-Marie tooth disease
(CMT1A) and hereditary neuropathy with
liability to pressure palsy (HNPP), respectively (Chance et al., 1994; Lupski
et al., 1991). These landmark discoveries
opened up the field of genomic disorders
in neurobiology and beyond (Lupski,
2009).
Similar combinations of advanced
cytogenetics, somatic hybrid techniques,
and molecular genotyping played a
critical role in refining the maps of several
neurodevelopmental disorders including
fragile X syndrome, Miller-Dieker lissencephaly, and Prader-Willi-syndrome
(Ledbetter et al., 1981; Reiner et al.,
1993; Verkerk et al., 1991). The discovery
of polymorphic tri- and tetranucleotide
repeats (Edwards et al., 1991) was a critical advance for defining dynamic mutations as a new mutational mechanism in
several neurological disorders (see
below). Thanks to this discovery, clinical
enigmas such as the Sherman paradox
in fragile X syndrome and the clinical
phenomenon of anticipation, involving an
earlier onset and more severe disease
in successive generations, such as seen
in disorders like myotonic dystrophy,
HD, and the ataxias, were resolved. The
development of large insert cloning and
other physical mapping techniques
(Burke et al., 1987; Schwartz and Cantor,
1984), as part of the framework for
sequencing the human genome, played
a crucial role in facilitating the discovery
of many disease genes during the
nineties. Certainly, cloning the gene for
Rett syndrome would not have been
possible in 1999 had it not been for the
Neuron 68, October 21, 2010 ª2010 Elsevier Inc. 165
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Overview
intense mapping and sequencing efforts
on the X chromosome (Amir et al., 1999).
With the release of the first drafts of the
human genome sequence in 2001, the
landscape of gene discovery changed
tremendously, so that the once tedious
physical mapping and cloning experiments slowly gave way to candidate
gene analysis by sequencing. Computational analysis of mapping intervals and
careful selection of candidate genes for
sequencing replaced months and years
of bench work. Having the sequence of
genomes from several other species, like
mouse, Drosophila, and, more recently,
nonhuman primates, has certainly advanced neurobiological disease research,
permitting in vivo functional studies in
a whole range of model organisms.
Without doubt, during the last ten years,
thanks to the integration of phenotypic
mapping and sequencing data with
tremendous analytical and computational
resources, the neurobiology community
has witnessed disease gene discoveries
at an amazing pace, sometimes on a
monthly or even weekly basis. Equally
inspiring and impressive has been the
fact that these gene discoveries have revealed new insights into basic biological
mechanisms that extend far beyond
the specific disease in question. Yet in
the face of such tremendous progress, it
has been surprising to see that there has
been a fair amount of pessimism within
the popular press about whether ‘‘the
genetics revolution’’ has fulfilled its
promise. In June of this year, the title of
a prominent NY Times article stated, ‘‘A
decade later, genetic map yields few
new cures’’ (Nicholas Wade, NY Times,
June 12, 2010). The first sentence read,
‘‘Ten years after President Bill Clinton
announced the first draft of the human
genome was complete, medicine has yet
to see any large part of the promised
benefits.’’ Questions have been raised
about whether the investment in genetics
and genomics has delivered.
Yet, when one considers some of the
transformative changes in the realm of
human neurogenetic disorders, there
can be little doubt that, in fact, genetic
discoveries over the past two decades
have already changed not only the practice of clinical medicine in neurology and
psychiatry, but also the outlook for many
families afflicted by these devastating
disorders. A mere 20 years ago, patients
with hereditary ataxia had to undergo
a number of expensive and sometimes
invasive investigations including brain
scans, spinal taps, numerous blood
tests, possibly electromyograms, nerve
conduction studies, and sometimes
peripheral nerve biopsies. Similarly, children with neurodegenerative disorders
or cognitive disabilities had to endure
a large number of tests, scans, and sometimes skin or conjunctival biopsies; worst
yet, a definitive diagnosis could not be
reached and the family was typically left
with an uncertain 50% or 25% chance
of recurrence in subsequent offspring.
Today, a large number of childhood and
adult neurological disorders can be diagnosed by a simple DNA test on peripheral
blood, saving patients the pain and cost
of many additional tests and providing
them with a definitive diagnosis. This
advancement affords families a better
understanding of the disorder afflicting
their relatives while providing them with
the option of genetic counseling and
allowing for preimplantation or prenatal
genetic testing, which could not have
been done previously. There are several
hundred neurological disorders that
can now be diagnosed molecularly,
including hundreds of cognitive and
developmental disabilities such as autism
spectrum disorders, dozens of inherited
ataxias, inherited neuropathies, dystonias, muscular dystrophies, epilepsies,
familial degenerative disorders, and a
handful of psychiatric disorders (http://
www.ncbi.nlm.nih.gov/sites/GeneTests/
?db=GeneTests). While arguably the
pace of development of potential therapies has been relatively slow compared
to the speed of disease gene discovery,
we should not underestimate the great
benefits to families of disease prevention
through prenatal diagnosis, and the gains
in fundamental neurobiology from pathogenesis studies of neurological disorders.
Humans Provide the Largest Series
of Phenotyped Alleles Revealing
New Mutational Mechanisms
We should also not underestimate the
impact that genetics has had on our
understanding of brain development and
function. It is a truism that humans are
the best model system. It has been said
that every nucleotide change in the
166 Neuron 68, October 21, 2010 ª2010 Elsevier Inc.
human genome compatible with life is
present at least once in someone on the
globe. Likewise, no other species on earth
is scrutinized so closely at a phenotypic
level. Advances in clinical medicine and
diagnostic technologies have radically
expanded the range of diagnostic tests
that physicians can use to unravel a
patient’s symptoms and history. With
respect to understanding brain function,
diagnostic imaging techniques such as
high-resolution MRI in 2D or with 3D
reconstruction allow for visual characterization of the human central nervous
system at an unprecedented level, and
functional MRI and MR spectroscopy
are starting to bridge the gap between
sheer structural phenotyping and physiological consequences with functional
relevance. Moreover, electronic communication spreads the word of unusual
phenotypes globally, whereas in the
past, knowledge of an extraordinarily
rare disorder may have been confined to
one country or even village. Thus, humans
possess a genome rich in variation being
examined daily by physicians for phenotypic variation—a geneticist’s dream
come true.
Our ability to appreciate full phenotypic
variation is directly related to the concept
of intraspecies examination. Human
variation is so rich because we, as examiners, and humans ourselves, can discern
remarkably subtle differences among
each other. This precision of deducing
subtle variation and the degree by which
such variation is cataloged in humans
has uncovered completely new mechanisms of disease. Several novel and now
well-appreciated mutational mechanisms
of neurological disease owe their original
description to human studies. Uniparental
disomy (Spence et al., 1988), which
uncovered the importance of genetic
imprinting in neurobiology, has its origins
in human genetics. Prions, which cause
various transmitted spongiform encephalopathies, were discovered in humans
and led to the then startling notion of the
propagation of protein misfolding by
misfolded proteins (Pruisner, 2001;
Mastrianni, 2010). Dynamic mutations
involving trinucleotide repeat expansions,
originally discovered in fragile X syndrome
and X-linked spinal and bulbar muscular
atrophy, now account for well over a
dozen other disorders and remain a largely
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Overview
human-specific mutational mechanism
(Orr and Zoghbi, 2007).
Further, as we have learned more about
our own human genetic make-up, we
have found insights into common biologies shared with other species as well.
Consider the FOXP2 gene, identified in a
family segregating developmental verbal
dyspraxia or the inability of sequencing
muscle movements required for articulating speech (Lai et al., 2001). While a clearcut phenotype observed by humans in
humans, the subtlety of the phenotype
would be unrecognizable by us in another
species. Recognizing the human phenotype, however, has now allowed the
demonstration that the FOXP2 transcription factor is important in neuronal
circuitry in mice and songbirds (Kelley
and Bass, 2010; Schulz et al., 2010).
Same Phenotype, Many Genes—
One Gene, Many Phenotypes
With better diagnostics and greater levels
of clinical scrutiny over the past century,
we have been able to more thoroughly
document the clinical and pathological
features of various disorders and classify
disorders into different categories. While
these clinical characterizations have
been extremely valuable in making some
sense of various phenotypically overlapping neurological and psychiatric disorders, they are also limiting in other
respects. For one, these classifications
change every few years depending on
the experts evaluating the patients, better
documentation of signs and symptoms,
and the availability of new diagnostic
tests, rendering it a challenge to keep up
with the changing classifications. Likewise, at times what seem to be discrepancies between clinical criteria and pathological measures can blur the lines
between categories. Now with genetics,
this is beginning to change and in the
process, some of the apparent confusion
about classifications and categories is
starting to make sense. A number of inherited ataxias have clinically similar,
if not identical, features, yet genetic
studies proved to us that they are indeed
caused by mutations in distinct genes.
Similarly, many neuropathies, dystonias,
myopathies, and cognitive disorders are
clinically indistinguishable yet genetically
distinct. In many cases where there is
apparent clinical overlap, but different
genes, the similarity may be reflective of
a convergent biology and common pathways. Basic research has revealed that
there is indeed a biological reason for the
clinical confusion and that understanding
the biological basis of these disorders
may shed light on why they overlap clinically. The protein products of many genes
that cause overlapping phenotypes do
indeed interact either directly or indirectly,
and several of these proteins function
in coincident pathways. This is true
for proteins causing ataxias, muscular
dystrophy, tuberous sclerosis, autism
spectrum disorders, Parkinson disease,
and Alzheimer disease (Brouwers et al.,
2008; Cookson and Bandmann, 2010;
Ess, 2010; Lim et al., 2006; Toro et al.,
2010; see also the Perspective by Hardy
[2010] in this issue). Ultimately, the
discovery of the causative genes for these
disorders may make the clinical classifications obsolete whereby new generations
of neurologists could simply focus on
the causative DNA mutation to classify
a disorder. Likewise, from the standpoint
of basic science, given the wealth of documented phenotypes in humans and the
rapidly increasing knowledge of the causative genes, revisiting clinical data and
reconsidering the implications of distinct
genetic causes could lead to new hypotheses about functionally related pathways
just as Drosophila geneticists have
beautifully elucidated the Notch signaling
pathway by starting with mutants that
share similar phenotypes (Fortini, 2009).
Indeed, this is a lesson learned. The
brain represents a huge mutational target
where many loci encode interacting
proteins and a dysfunction of any one
may produce a similar consequence to
the entire pathway. With this in mind,
perhaps it is not surprising that very few
strong genome-wide association signals
have been uncovered for psychiatric
disorders. If we lump phenotypes into
a single clinical category (i.e., schizophrenia), the degree of genetic heterogeneity present renders association studies
extremely underpowered. It may be wise
to consider stratification of behavioral
phenotypes into very specific subcategories, as was done for the many genetic
ataxias, when pursuing gene identification.
A more surprising discovery is the
finding that mutations of one gene might
cause clinically distinct phenotypes in
different patients. Examples include
the Aristaless-related homeobox gene
(ARX), which causes X-linked lissencephaly, agenesis of corpus callosum with
abnormal genitalia, cognitive deficits
with or without seizures, or cognitive deficits, dystonia, and seizures (Partington
syndrome) (Shoubridge et al., 2010), as
well as the SHANK3 gene, whose
mutations can cause Phelan McDermid
syndrome, Asperger syndrome, autism,
and rare cases of schizophrenia (Durand
et al., 2007; Gauthier et al., 2010; Phelan,
2008). Similarly, mutations in NRXN1 can
cause rare forms of autism and schizophrenia (Kim et al., 2008; Rujescu et al.,
2009), whereas mutations in LMNA,
encoding for lamin A and lamin C (Stoskopf and Horn, 1992), can cause a variety
of disorders including Emery-Dreifuss
muscular dystrophy Type 2 (Bonne
et al., 1999), Charcot-Marie-Tooth axonal
neuropathy (CMT2B1) (De Sandre-Giovannoli et al., 2002), limb girdle muscular
dystrophy Type 1B, Hutchinson-Gilford
progeria syndrome (Eriksson et al.,
2003), and various other distinct clinical
phenotypes. Altogether, these findings
suggest that the neuroanatomical and
physiological disturbances resulting from
dysfunction of the respective genes may
be influenced by the genetic background
and environmental experiences of the
affected individuals, leading to different
clinical outcomes in different patients.
Related to this, some of the most interesting discoveries pertain to the finding
that the nervous system is exquisitely
sensitive to the dosage of many proteins.
Haploinsufficiency or loss as well as
doubling of several genes seems to
cause overlapping neurological phenotypes including Parkinson disease, Alzheimer disease, the case of peripheral
myelin protein 22 in neuropathies,
MeCP2 in Rett syndrome, and MeCP2
duplication disorders, and the example
of gain-of-function and loss-of-function
mutations in neuronal ion channels
causing epilepsy and other neurological
deficits (Amir et al., 1999; Catterall et al.,
2008; Van Esch et al., 2005; Zhang et al.,
2010). Although it is still not clear how
either loss or gain of the same protein or
proteins causes similar cognitive and
social behavior phenotypes, it is conceivable that the phenotype is a manifestation
Neuron 68, October 21, 2010 ª2010 Elsevier Inc. 167
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of a failure in neuronal homeostatic
response due to downstream effects of
various molecular changes (Ramocki
and Zoghbi, 2008; Toro et al., 2010).
Understanding neuronal homeostatic
responses and their modulation may facilitate development of potential therapies
for a broad class of disorders, irrespective
of the underlying primary genetic defect.
CNVs and Variable Penetrance
Gene dosage alteration has emerged as
a widespread phenomenon in neuropsychiatric disease, largely in the form of
copy-number variation (CNV) (Pollack
et al., 1999). The delineation of the human
genome sequence allowed the widespread survey of deletions, duplications,
and inversions. Surprisingly, the normal
human genome is littered with such
variation (Redon et al., 2006; Sebat
et al., 2004). Most are considered common benign polymorphisms (although it
remains unclear how such normal variation might contribute to the mutational
load predisposing to disease), but the
rare, large, or, even more important, de
novo CNVs seem to contribute substantially to several complex genetic disorders. Sebat et al. (2007) reported in 2007
a ten-fold elevation in large, de novo
CNVs in autism with similar observations
reported for schizophrenia, epilepsy, and
idiopathic intellectual disability. Such
recurrent CNVs may finally provide a
definitive genomic location for genes
that may predispose to these disorders.
An emerging feature of disease-associated CNVs is the variable expressivity
exhibited by many of these DNA rearrangements (Girirajan and Eichler,
2010). Deletions and duplications at
16p13.11 were initially reported in children with intellectual disability (Ullmann
et al., 2007), then autism (Hannes et al.,
2009), then epilepsy (Heinzen et al.,
2010), and finally schizophrenia (Ingason
et al., 2009). Similarly, 1q21.1, 15q11.2,
15q13.3, and 16p11.2 CNVs, to name
just a few, have all been associated with
diverse neuropsychiatric phenotypes
(Lee and Scherer, 2010). Another example
of this potential clinical continuum is seen
at 3q29 where similar deletions have all
been associated with either intellectual
disability and mild dysmorphic feature
(Willatt et al., 2005), autism (QuinteroRivera et al., 2010), bipolar disorder
(Clayton-Smith et al., 2010), or schizophrenia (Mulle et al., 2010). Within the
3q29 interval lie two genes, PAK2 and
DGL1, both of which have paralogs elsewhere in the genome that lead to intellectual disability and are strong candidates
for further study.
While disease-related CNVs must operate, at a very fundamental level, by
dosage sensitivity, it remains unclear
how specific CNVs result in such disparate phenotypes. There are several
possibilities. First, the precise CNV breakpoints, in many cases, are not mapped
to the nucleotide level, so overlapping
variants could exhibit different phenotypes. A deletion may unmask a recessive
variant on the second allele or there could
be cis effects where the CNV influences
the expression of nearby loci. Alternatively, as has been well documented in
imprinting disorders, the loci within the
CNV may exhibit parent-of-origin effects
with the consequence of differing levels
of expression of the now hemizygous
genes on the second allele.
Another possibility and perhaps a
general concept for rare disease-associated CNVs is that such variants lower
susceptibility thresholds and based upon
genomic background and/or environmental influences result in phenotypes
on a clinical continuum. Eichler proposed
a more specific two-hit model (Girirajan
et al., 2010) where a second CNV, elsewhere in the genome, influences penetrance and expressivity of the first CNV.
Evidence for this model is best illustrated
by a 600 kb microdeletion at 16p12.1 that
has been found in children with intellectual disability and/or autism. Unlike most
disease-associated CNVs, the 16p12.1
deletion is rarely de novo, being inherited
from one parent in !95% of the cases.
The carrier parent, while cognitively
intact, often exhibits milder phenotypes
such as depression, mild learning
disability or seizures. In 25% of the cases,
the child with the more severe phenotype
has a second large CNV elsewhere in the
genome, a 40-fold increase over the predicted rate. A corollary of this model is
that rather than a second CNV, a second
conventional mutation elsewhere in the
genome tips the balance to a severe
phenotype. Low-cost deep-sequencing
technologies should uncover such genegene interactions. Clearly, many more
168 Neuron 68, October 21, 2010 ª2010 Elsevier Inc.
genetic studies on CNVs influencing
neuropsychiatric disease are required,
but from our perspective, CNV research
is well worth following for the neuroscientist interested in disease mechanisms.
Going Back to the Basics
With the excitement and rapid pace of
disease gene discovery came the expectation that therapies for these disorders
are also readily within reach, and a
major frustration that is often expressed
by patients, their families, members of
Congress, and disease-based foundations is the lack of effective therapies
in spite of the formidable and exciting
rate of disease gene discovery. This
may be in part due to the false expectations or the hype around what a gene
discovery can deliver in the short term.
Gene discovery is a critical but only
a first step in the path to therapeutics
development. A variety of intricate indepth investigations must take place in
order to reveal pathways that lend themselves to therapeutic intervention in any
given disease.
For instance, if one considers the path
to the discovery of the tyrosine kinase
inhibitor, Gleevec, one realizes that it
took approximately 40 years of dedicated
and intense basic research from the time
of the identification of the Philadelphia
chromosome until the FDA approved the
drug. Perhaps what is more important is
to identify the potential obstacles that
might have contributed to such a delay
and the steps in the process that are
most challenging, and then one can
implement solutions to accelerate the
path to drug discovery. One thing is
certain: to increase the likelihood of
success in developing targeted therapies,
one needs to understand the function of
the protein or RNA mediating a specific
disease process.
It is becoming abundantly clear that
most proteins serve many diverse functions rather than only one or two isolated
functions. Yet, traditionally, investigators
have focused on one aspect of a disease
protein based on one or more observations reproduced in several outstanding
labs. While this research strategy is
productive because it is focused and will
likely yield insight into some function
of the culprit protein, it is often not sufficient when one is tackling a complex
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Overview
neurological disorder. Perhaps the case
of amyloid precursor proteins (APP) and
presenilin1 (PS1) is illustrative of this
point. There is no doubt that dosage of
APP is critical to the generation of Ab,
and that Ab accumulates in the aging
brain and in Alzheimer disease (AD). The
vast majority of scientists and pharmaceutical companies have focused on this
aspect of AD for the development of therapeutics. In recent years, however,
research into other potential functions of
APP and PS1 is beginning to yield additional potential mechanisms by which
excess APP or mutant PS1 might
compromise neuronal function. For
instance, the study by Nikolaev and
colleagues demonstrating that APP binds
death receptor 6 (DR6) and activates caspase 6, which in turn triggers axonal
pruning and neuronal death, is interesting
and important. The finding highlights
a role for a portion of APP distinct from
Ab, (N-terminal cleaved fragment that is
also generated in a b-secretase dependent manner) as a potential contributor
to AD (Nikolaev et al., 2009). More
recently, Lee and colleagues demonstrated that presenilin1 is critical for targeting the v-ATPase V0a1 subunit to the
lysosome, and thus for the proper acidification of the autolysosome (Lee et al.,
2010). Interestingly, several AD-causing
mutations in PS1 compromised this function, raising the possibility that defective
lysosomal proteolysis is a key contributor
to AD pathogenesis. The odds are that
there are many mechanisms that
contribute to AD and that it may be necessary to target more than one of these
mechanisms and at multiple stages of
the disease (perhaps including initiation
and progression, rather than just the end
stage) to rescue and treat the disease
effectively. Taking a fresh and unbiased
look at disease-causing proteins to
pursue their characterization in depth
might remove some of the barriers to
accelerating research. As clues about
potential new functions of these proteins
emerge, validating such functions in vivo
and in various disease models will be critical. Cross-species studies have been
crucial for revealing in vivo protein functions, but ultimately the studies have to
be done in rodent or nonhuman primates
in preparation for translational studies
and therapeutic interventions.
On Animal Models
One of the clear lessons learned over
the past two decades is the importance
of animal models in delving deep into
the pathophysiology of neurogenetic
diseases. Without question, mice,
Drosophila, zebrafish, C. elegans, and
even yeast models have provided an
experimentally tractable model to test
mechanistic hypotheses that might be
impossible to evaluate in humans. The
mouse has been the mainstay in animal
models for neurogenetic disorders due
to robust gene manipulation techniques,
similar neuroanatomical structures, and
the ability to perform electrophysiology
to directly measure neuronal function
and circuit activity. The toolbox for the
genetic manipulation of the mouse is full
of clever and highly useful approaches.
In addition to knockout and knockin
approaches, spatial or cell-specific regulation, as well as temporal and/or reversible regulation, have been widely utilized.
A spectacular example is Adrian Bird’s
demonstration that activation of a silenced
Mecp2 gene reverses the Rett syndromelike phenotype in adult mice (Guy et al.,
2007). This work fueled the notion that
seemly hopeless developmental disorders, such as Rett syndrome or fragile X
syndrome, may be therapeutically
approachable.
Less robust but still of strong utility are
mouse behavioral models of neurogenetic
disorders. Interlab variation has sometimes led to discordance in the literature.
Some of this variation was due to less
appreciated but substantial strain differences and the need for true isogenic
backgrounds for behavioral studies.
Ideally, neurobehavioral studies should
be performed on F1 hybrids derived
from two different pure isogenic backgrounds to control for strain-specific
deficits that might confound the phenotype of the engineered mutation. We
also learned that investigator-specific
differences sometimes lead to failure in
replication, often due to subtle differences
in procedures or housing environment.
Alternatively, we may be simply asking
too much of the humble rodent (Sousa
et al., 2006). For example, a number of
human loci, when mutated, lead to severe
intellectual disability. The same mutations
in the mouse do cause learning differences, but sometimes only by phenotyp-
ing large numbers of mutant and control
mice can such differences be discerned
statistically (Bouwknecht and Paylor,
2008). This is not the ideal situation for
preclinical testing of drugs, since the
effect size in the rodent is so small.
What can be done to improve this?
Better testing paradigms may be of
some help but, for many cognitive tests,
laboratory mice, descendents of ‘‘fancy’’
or pet mice at the turn of the last century,
were directly or inadvertently selected for
a docile manner, which perhaps dulled
their abilities to perform in many paradigms of learning and memory. Use of
more outbred mice, such as M. spretus
(Dejager et al., 2009), is one possibility
although difficult to breed and house.
Another possibility is to return to the rat,
for decades the animal of choice for
behavior studies that fell out of favor to
the mouse, which was genetically modifiable. Recent advances in making targeted
mutations in the rat may mark a return of
this species (Geurts et al., 2009; Tong
et al., 2010).
Another model system that has proven
itself very useful in neurogenetic research
is Drosophila. It was somewhat surprising
initially that Drosophila models have replicated many of the pathogenic processes
of human neurological disorders, despite
gross anatomical and genomic differences between humans and fruit flies. Of
an estimated 392 human genes that lead
to cognitive deficiency when mutated,
over 300 have Drosophila orthologs
(A. Schenk, personal communication).
The power of Drosophila as a genetic
model to identify genetic interactions
and epistatic loci is unmatched and
permits assignment of loci to established
pathways given the rich gene ontology
and phenotypic data in this model
organism. Studies of the consequences
of loss of parkin function illustrate this
point most clearly. Parkin null mice did
not manifest overt neurodegeneration or
behavioral phenotypes. Drosophila parkin
mutants on the other hand revealed the
importance of parkin for mitochondrial
functions. This discovery eventually
opened up the field on the importance of
several Parkinson disease-causing loci
for mitochondrial integrity and function
(Clark et al., 2006; Greene et al., 2003;
Jones, 2010; Park et al., 2006; Pesah
et al., 2004).
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In recent years, with the ability to develop transgenic monkeys, the nonhuman
primate Rhesus macaque has emerged as
a potential model system. Chan and
colleagues (Yang et al., 2008) developed
a Huntington disease transgenic Rhesus
model that displayed a number of clinical
features, seen in human that are absent in
the mouse model. Although currently
limited to transgenic manipulation only,
the monkey, even with the expense, may
prove valuable for preclinical testing,
particularly when the mouse phenotype
lacks certain human phenotypes, such
as neuronal loss in Huntington disease
(HD). In a similar vein, other large animal
models, such as transgenic pigs that
have recently been shown to better mimic
HD than the mouse model (Yang et al.,
2010), may also prove to be useful. It
should, however, be pointed out that
while incredibly valuable, there are potential limitations to animal models and it is
critical to assess these limitations for
better understanding of human disease
pathogenesis. For instance, a great deal
has been learned using these and other
animal models of HD. However, a large
number of HD pathogenesis studies rely
on models that do not accurately recapitulate the HD mutation. Rather, many use
a truncated short peptide containing the
polyQ tract for experimental convenience
and because animals expressing this
short peptide manifest neurodegeneration in a short time frame. Since therapeutic strategies must target pathways
affected by the entire protein, such
shortcuts might, in the end prove unfortunate and counterproductive. Mouse and
Drosophila animal models for spinocerebellar ataxia type 1 and spinobulbar
muscular atrophy, two polyQ disorders,
have clearly demonstrated the importance of domains beyond the polyQ tract
in disease pathogenesis (Duvick et al.,
2010; Fernandez-Funez et al., 2000;
Kratter and Finkbeiner, 2010; Nedelsky
et al., 2010).
Regardless of the model system, it will
be important to conduct preclinical
testing over an extended period of time.
Too often, such studies are short term
and leave the long-term consequences
unanswered. It will be important to determine during preclinical testing which
phenotypic improvements are sustained
and which are not. While expensive, it is
infinitely less expensive and devastating
than a failed human trial. With the revelation of new insights into the molecular
causes of Mendelian disorders and the
wide availability of chemical library
screening, it is likely that lead compounds
for therapeutic development will arise
for numerous human genetic disorders.
The challenge will be to maintain and
translate the quality and comprehensiveness of the preclinical testing into model
systems. Scientists must bear the responsibility of conducting and replicating
preclinical trials and insuring that meaningful outcome measures are improved.
Rigorous design of preclinical trials will
save hundreds of millions of dollars in
failed clinical trials and will liberate
patients to be enrolled in clinical trials
with higher promise.
The Future
The field of neurobiology is well positioned to take advantage of two decades
of gene discoveries, excellent animal
models, and lessons learned from failed
experiences. Key to the success of
disease-oriented research are a few but
important ingredients: in depth studies
to gain knowledge about the function
of the culprit proteins, excellent animal
models of disease, and an understanding
of the physiological and pathological
consequences of disease protein dysfunction. As much as possible, investigators must use models that express the
human mutation in the context of the full
proteins, and in the correct spatial and
temporal expression patterns. Conclusions must be based on in vivo studies
that use interdisciplinary approaches
including cell biological, molecular,
behavioral, and physiological methods.
Investigators with diverse expertise must
work together for better characterization
of animal models and for identification
of quantifiable and reliable outcome
measures.
As important as good preclinical
science is to the ultimate success
of translating scientific discoveries to
clinical application, we also desperately
need appropriate infrastructure. This
requires successful partnerships between
academic research, governments, private institutions, and foundations and
pharmaceutical industries. Academic
researchers are best at performing funda-
170 Neuron 68, October 21, 2010 ª2010 Elsevier Inc.
mental research studies. They are motivated to do these studies irrespective
of how esoteric or rare the problem
they are tackling. Performing multidisciplinary in vivo and interdisciplinary pathogenesis studies is certainly best done in
academia, and such studies are likely to
reveal pathways that can be targeted therapeutically. On the other hand, pharmaceutical companies have unparalleled
expertise in medicinal chemistry and
a rich resource of drugs and compounds
that can be repurposed for specific neuropsychiatric disorders based on relevant
discoveries. Somehow, we need to better
bridge the interests and expertise of
academic science and industry. The
challenges that might keep the pharmaceutical companies from sharing their
compounds for repurposing must be identified and overcome. Barriers such as
a fear of the U.S. Food and Drug Administration’s response to side effects that
might result from a new use must be
managed by educating the FDA that
drug/genotype effects need not be generalized. The case of the mGluR5 antagonist
is an example where an existing drug
was put to good use. When it was discovered that the loss of FMRP in fragile X
syndrome leads to apparent overstimulation of mGluR (Huber et al., 2002), it
became apparent that lowering mGluR5
signaling might suppress several phenotypes in the fragile X syndrome mouse
and fly models (Bear et al., 2004). Merck,
having mGluR5 antagonists in its collection, licensed a compound for development as therapeutic in fragile X syndrome,
despite the fact that fragile X is a relatively
rare disorder and perhaps not likely to be
as profitable a market as would be
expected for a development of a new
drug. Pharmaceutical companies often
have to weigh the risks and benefits of
repurposing drugs or forging collaborations to treat rare disorders, but this is
the right thing to do and a risk worth taking.
Most disorders are indeed rare by
themselves but cumulatively account for
a large number of cases of a ‘‘common’’
disorder. Also, predisposition to common
disorders may reflect accumulation of
several rare genetic variants. Thus, if we
can develop effective therapies through
a successful collaboration between
academia and pharmaceutical companies, the odds that we can gradually tackle
Neuron
Overview
rare neuropsychiatric disorders effectively
will also be enhanced.
A few principles are worth considering
as we ponder solutions for the growing
list of neuropsychiatric disorders. We
need to invest in studies aimed at a better
understanding of normal brain development. While we often think of development as being critical for understanding
the integrity of the brain in infants and
in children, data are gradually pointing
to the fact that early developmental
processes might impact the integrity and
health of the adult and the aging brain as
well. A better understanding of both the
genetic and environmental factors that
affect synapse development and maintenance is likely to provide information on
how such factors might be co-opted to
protect the aging brain or a brain dealing
with mutations causing late-onset neurodegenerative disorders. The role of epigenetics in modulating disease processes
must also be investigated. The reality is
that many genes will be very hard to target
or manipulate to suppress a disease
phenotype. However, one can epigenetically modulate the function of genes
without altering their sequence or activity.
The findings that environmental factors
such as diet (Waterland et al., 2006;
Weaver et al., 2005; Wolff et al., 1998) or
experiences (McGowan et al., 2009) can
alter the epigenome open up the possibility that modulating gene function
through epigenomic modifications might
be a productive way to treat some disorders. Of course, it is equally important
to consider the flip side of the coin, that
damage to the epigenome from changing environmental factors could also
contribute to or aggravate disease. The
growing list of genetic defects that may
lead to neuropsychiatric disorders point
to the many ways any molecular perturbation can lead to devastating neurological
phenotypes. However, we must not forget
what we can learn from individuals who
have similar mutations but milder phenotypes. Exploring environmental factors
and experiences that might have protected such individuals, or identifying
suppressor mutations through the new
deep-sequencing technologies, might
reveal factors that modify a disease
process.
Lastly, and perhaps a most exciting
note with which to conclude, is the fact
that genetic research has revealed the
plasticity and resilience of the developing
and adult brain. Gene discovery and creative genetic engineering in mouse models
have taught us that many neuropsychiatric disorders are reversible (at least
genetically) (Guy et al., 2007; Santacruz
et al., 2005; Yamamoto et al., 2000; Zu
et al., 2004). The findings that several
disorders, including some of the most
devastating developmental and degenerative diseases are reversible in mouse
models, provides hope that discovering
ways to counteract or suppress disease
processes might halt or even reverse
some of the most serious neurological
and psychiatric disorders.
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