Are genome-wide association studies all that we need to dissect the genetic component of complex human diseases?

Abstract

With the availability of dense maps of anonymous and frequent SNPs spanning the whole human genome, genome-wide association studies are now becoming a reality. In this paper, we discuss the utility of these approaches to detect genetic risk variants involved in complex disease susceptibility and, in the best case scenario where a signal is detected, how helpful it will be to the understanding of the pathological process.

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Are genome-wide association studies all that we need to dissect the genetic component of
complex human diseases?


Catherine Bourgain1, Emmanuelle Génin1, Nancy Cox2 and Françoise Clerget-Darpoux1

1: INSERM U535 and Univ. Paris Sud, 94817 Villejuif. France
2: Department of Medicine and Department of Human Genetics, University of Chicago. USA



Given the disappointing results obtained by the Human Genetics community through
systematic linkage screenings of the genome, Risch and Merinkangas argued that “the future
of the genetics of complex diseases is likely to require large scale testing by association
studies” 1. If linkage studies have low power to detect common variants with small odds ratios
(OR), they are also doing a poor job at detecting very frequent variants with high genotypic or
allelic ORs, a situation where association tests might perform better. A good illustration of
this point is provided by the VNTR flanking the insulin gene. The association between the
locus and type I diabetes is easily detected, the risk allele is frequent (about 70% in the
general population) with an allelic OR around 3.92. The distribution of alleles shared Identical
By Descent (IBD) in sib pairs, however, is very close to its null expectation and linkage is
very difficult to detect. Even on large sib pair samples, discordant results have been
described3,4. An additional interesting feature of association studies is to allow a much more
precise location of risk factors. Whereas the precision of location provided by linkage studies
is sized in cM, it is sized in kb or even bp for association studies.
Technological developments have rapidly made large scale association studies a reality. The
HapMap project (www.hapmap.org) launched by an international consortium by the end of
2002, publicly released the first data in 2005. The first genome-wide association studies have
been published recently5-7. Large scale association studies using maps of anonymous and
frequent SNPs are presented as the new tool that “will accelerate the discovery of genes
related to common diseases” (HapMap Press Release, 7th February 2005). In the present
paper, we would like to debate around two questions that are central to the discussion on the
power of genome-wide association studies: will most genetic variants involved in complex
diseases be detectable by systematic association testing and, in the best case scenario where a
signal is observed, what will be its contribution to the understanding of the pathological
process?
inserm-00119120, version 1 - 15 Jun 2007
Author manuscript, published in "Eur J Hum Genet 2007;15(3):260-263"
DOI : 10.1038/sj.ejhg.5201753

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Will most genetic variants involved in complex diseases be detectable by systematic
association testing?

The standard strategy for systematic association testing is to perform allelic tests using SNPs
tagging common variants and/or haplotypes to limit both the multiple testing correction and
the genotyping cost.
By construction, this approach has been recognized to have low power to detect effects of rare
variants, which are poorly tagged by common SNPs. Relatively rare variants are known to
contribute to a variety of common, complex diseases. In Crohn’s disease, the three risk alleles
in the NOD2 gene have frequencies smaller than 5% in populations of European descent 8,9.
Population genetics modelling of the expected genetic variation at disease susceptibility loci
has also suggested that rare variants may play an important role in complex disease
susceptibility 10,11. When selection is weak, as seems likely for many complex disease
mutations, genetic drift becomes important, and the total frequency of susceptibility mutations
is expected to vary widely among loci.
In contrast, almost all common SNP-based variation as well as other SNP-correlated common
variants (including common deletions12,13) can be interrogated either directly or by surrogate
SNPs or haplotypes. The issue is whether we will be able to design studies powerful enough
to detect them. A number of variants with both high frequency and sizable effects have been
identified, including ApoE4 in Alzheimer’s disease and cardio-vascular diseases or HLA
factors in autoimmune diseases. Variants with similar sizeable effects may still have to be
discovered. Some may have escape identification by linkage studies because of a very high
susceptibility allele frequency. Others may be located in regions already broadly detected by
linkage as in the recent whole genome association successes 6,7.
Aside from these large effect variants, common susceptibility alleles or haplotypes with quite
modest ORs – in the range of 1.15 – 1. 5, with the majority in the range of 1.15-1.2514- have
been shown to have reproducible effects (via meta-analysis) on various phenotypes. These
variants with low marginal risks may, in fact, have strong effects on disease, but restricted to
particular genetic background or environment. Indeed, complex models of interaction
between genes, sex and environment have been extensively described in animal models for
various complex traits15. Identifications of these variants – that may be far from exceptions in
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humans – may thus constitute important steps in the understanding of the physiopathology of
the diseases, but very challenging ones.
Finally, to control for the multiple testing in genome-wide context, effects of SNPs may only
be tested at the allelic6,7 or simple haplotypic level (haplotypes made of 2 or 3 markers). But
interaction between alleles or between distant SNPs within a susceptibility gene may be
poorly detected at the allele level even in the presence of non negligible effects at the
genotype/haplotype level.
Whether we will be able to power studies to detect common risk alleles with OR<1.25 for all
diseases, is a critical issue. Apart from the challenge to collect huge samples without
compromising on the quality of phenotypes, the crucial issue is whether human samples of
sufficient sizes with homogeneous allele frequencies and pattern of linkage disequilibrium
(LD) do exist. For some diseases, the inter-population variability may be too large to allow
the recruitment of samples not subject to population stratification and large enough to detect
variants with small effects. Similarly, once the variant is detected in a first study, it might be
difficult to obtain samples to perform replication studies. First, these samples will need to be
as large or even larger16 and second, the difference in both variant frequencies and LD pattern
among populations may make the signal undetectable in other populations. The difficulty of
replication is well illustrated in the recent work of Reich et al17 on Multiple Sclerosis where
an association on chromosome 1 in African-Americans is not replicated in another sample of
Afro-Caribbeans. Consequently, it is not possible to decide whether the association described
is driven by a causal variant or whether it is just another false positive result.

Altogether, we believe that these different elements invite to cautiousness regarding the
ability of genome-wide association studies to detect most genetic variants involved in
complex diseases.

In the best case scenario where a signal is observed, what will be its contribution to the
understanding of the pathological process?

The main motivation behind looking for genetic risk factors for complex diseases is to get a
better insight into the pathological process of the diseases. However, to go from an
association signal to a deeper knowledge on the causal variants and their effect measurements
is not that simple. The study of the HLA component in auto-immune diseases well illustrates
the caveats encountered.
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If the association between type 1 diabetes (IDDM) and the HLA class II regions has been
known for more than twenty years, the causal variants of the region have still to be identified.
A particular amino-acid (non-“Asp57”) of the DQ chain was suspected for a while18 as it is
strongly associated with the disease: patients with an aspartic acid at position 57 in the DQ
chain are very rare. However, this hypothesis was rejected19, because it could neither explain
the overall HLA genotype distribution, nor the HLA haplotype sharing of affected sibs19. This
example illustrates the importance of not only considering allelic association but also
genotypic distributions and the importance of identifying models that can explain both the
association and linkage information and not only one of them.
In celiac disease, genetic susceptibility in the HLA region is mainly explained by one DQ
heterodimer which two components (DQA1 and DQB1 alleles) can either be on the same
HLA haplotype (cis-position) or one on each complementary haplotype (trans-position)20.
This functionally relevant model was found using both biological elements on the structure
and function of HLA molecules21 and the observed genotype distribution in patients.
However, because it cannot clearly explain the HLA sharing among affected sibs and because
there are geographical differences in the risks conferred by the DQ heterodimers 22, this model
cannot account for the whole HLA component of the disease, which suggests that other HLA
factors must be involved.

In these two examples, the HLA component acts differently, through complex interactions of
numerous variants. Deeper understanding could be obtained only through more detailed
knowledge of gene diversity, patient genotype distributions, IBD sharing in affected siblings
conditional on patient genotype, and biological understanding of the function of genes in the
HLA region. Detection of an association was only the very first step.
In a recent study, Lincoln et al 23 used a large-scale SNP association strategy for studying the
involvement of the MHC in Multiple Sclerosis (MS). The strongest association is observed
with haplotype blocks in the HLA class II region and conditional on this association, there is
no evidence for other HLA-region risk factors for MS. These results are an additional
illustration of the limits of strategies relying on only association information. Despite the use
of a large numbers of simplex and multiplex families with dense SNP typing, the study just
ends at a result which has been known for many years.
HLA is certainly a unique region of the genome for its genetic complexity (level of
polymorphism, pattern of LD and clustering of many functionally related genes). Therefore,
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the challenges encountered in this region are not fully representatives of the ones that we face
elsewhere on the genome. Still, the HLA region is also one of the most extensively studied
and one exhibiting the strongest genetic factors for many complex diseases described so far.
This, together with the accumulation of experiences in other regions of the genome, makes us
believe that, though different, these non-HLA challenges may not be less complex.

The availability of tools (e.g. high-throughput genotyping platforms) and information (e.g. a
haplotype map of the human genome) are a valued and welcome addition to the
armamentarium geneticists can use to identify and characterize the genetic component to
common diseases with complex transmission. But we should resist the temptation to assume
that the models most favorable for the utility of these new tools must be the right models for
all the variants involved in complex diseases just because our studies will work better if these
most favorable models are correct. We can ill afford to go from a generation of underpowered
and ultimately disappointing linkage and family studies directly to a generation of
underpowered and disappointing association studies.
At the same time, the impressive diversity of causal variant models already identified favors
continued diversification of strategies. Identification of biologically relevant novel pathways
of disease susceptibility followed by candidate gene strategies allowing for the simultaneous
consideration of these connected genes may be quite effective at identifying new variants
affecting susceptibility. Linkage data contain considerable additional information capable of
aiding the discrimination of causal variation from the nearby variation that is merely in LD.
Consequently, linkage studies may still pay dividends even if the newer tools are as successful
as we all hope and we believe efforts to recruit family data should not be given up too quickly
in favor of only large samples of cases and controls. Linkage and association studies should
be considered as complementary strategies and not as concurrent.

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