A study in Japan finds that unidentified genes for hereditary diseases and that molecular etiologies for the majority of patients with sporadic neurological diseases remain to be elucidated.
Emerging new therapies for neurological diseases herald brilliant future directions for cures of devastating neurological diseases.These new therapies directly target disease mechanisms brought by mutations in causative genes for hereditary diseases. Recent accomplishments of new efficacious treatments employ broad disciplines, including gene transfer, gene silencing and activation of alternative genes.Given these remarkable therapeutic efficacies, identification of molecular basis of neurological diseases provides solid starting point for developing therapies.
Diseases are generally classified into hereditary and sporadic diseases. Sporadic diseases are also called complex trait diseases, since multiple genes and environmental factors are considered to underlie the disease development. In neurological diseases, quite similar disease presentations arise not only as hereditary diseases but also as sporadic diseases. For example, in amyotrophic lateral sclerosis (ALS) 5-10% of cases are hereditary forms, while the rest are sporadic diseases.
In our recent study, pathogenic mutations in ALS causative genes have been identified in 54% of families with hereditary ALS, but in 3.9% of patients with sporadic ALS in the Japanese ALS case series.6 These findings indicate that there still remain as yet unidentified genes for hereditary diseases, and, moreover, that molecular etiologies for the majority of patients with sporadic neurological diseases remain to be elucidated.
Exploring molecular basis of hereditary neurological diseases
The research paradigm identifying causative genes for hereditary diseases have already been well established, where many families needed to be collected to conduct linkage analysis based on genome-wide analysis of polymorphic markers to determine the locus of causative genes on the human genome, followed by laborious sequence analysis of genes located in the candidate regions. Availability of next generation sequencers (NGS) has dramatically changed the landscape of ‘gene hunting’. Now, candidate mutations can be directly identified by comprehensive genome sequencing of affected individuals without performing linkage analysis. NGS is now applied for clinical sequencing to establish the molecular diagnosis of hereditary diseases. The recent study has demonstrated that the diagnostic yield employing NGS has been increased up to 36.7-46.8%.
The results also raise various issues. Firstly, a substantial number of causative mutations may not be detected by currently popular NGS that generate short reads of 100-150bp. In addition to this, a substantial number of causative genes are as yet to be identified. According to the current statistics of OMIM there remain 1,566 phenotypes with unknown molecular basis, indicating that we should put further effort to elucidate molecular basis for these hereditary diseases.
Most of the currently available NGS generates billions of short reads of 100-150bp. This is the limitation in exploring structural variations, large insertions/deletions and tandem repeat expansions. Recently, single-molecule sequencing technologies that enables acquisition of far larger sequences have been introduced.These technologies are expected to facilitate identification of such variations in the human genome that are difficult to be identified by short read sequencers.
Of note, an increasing number of neurological diseases have recently been identified to be caused by non-coding repeat expansions.We have recently discovered non-coding repeat expansions in six diseases, including TTTCA repeat expansions in benign adult familial myoclonic epilepsy (BAFME) and CGG repeat expansions in neuronal intranuclear inclusion disease (NIID) and related disorders.To quickly identify expanded repeats employing short read sequencers, we developed a new program called TRhist, which generates a histogram of short reads filled with repeat motifs by analysing the whole genome short read sequence data from affected individuals. Employing TRhist, we identified short reads filled with (TTTCA)n and those with (CGG)n exclusively in the affected individuals, which enabled us to quickly discover the six causative genes. Intriguingly, expansions of the same repeat motifs have been shown to lead to identical or overlapping clinical phenotypes, indicating that expanded repeat motifs, irrespective of genes where expanded repeats are located, are directly involved in development of phenotypes of the diseases. Since families with hereditary diseases are rare, collaboration and data sharing among the researchers and clinicians on the globe will become essential.
Exploring molecular basis of sporadic neurological diseases
Currently, the molecular basis of sporadic diseases is not fully understood. Previous observations suggested involvement of genetic factors for sporadic neurological diseases including Alzheimer disease, Parkinson disease and amyotrophic lateral sclerosis. Over the past couple of decades, genome-wide association studies (GWAS) employing common single-nucleotide polymorphisms (SNPs) have been intensively conducted to identify risk alleles. The theoretical framework of GWAS is the ‘common disease-common variants’ hypothesis. Although GWAS have successfully revealed numerous susceptibility genes, the odds ratios of the risk alleles are generally low and account for only a small proportion of estimated heritability.20The current experience with GWAS suggests that rare variants that are difficult to be detected by GWAS may account for the missing heritability. Such rare variants may have large effect sizes for complex trait diseases. Thus, the research paradigm should be shifted to the ‘common disease-multiple rare variants’ hypothesis to identify disease-relevant alleles with large effect sizes.
An excellent example of rare variants with large effect sizes is best illustrated by the recent discovery of the glucocerebrosidase gene (GBA) as a robust genetic risk factor for Parkinson disease (PD). Recent clinical observations have suggested the association of sporadic PD with heterozygous mutations in GBA encoding glucocerebrosidase, the enzyme that is deficient in patients with Gaucher disease. The carrier frequency of the GBA variants was as high as 9.4% in PD patients and significantly higher than that in controls (0.37%) with a markedly high odds ratio of 28.0 in the Japanese population.In multiple system atrophy (MSA), we identified bi-allelic mutations of COQ2 in multiplex MSA families, and furthermore revealed that a heterozygous COQ2 variant, V393A, is associated with sporadic MSA.26 Association of V393A with MSA has been confirmed by meta-analysis in East Asian populations.Of note, the V393A variant is present in East Asian populations, but not in populations of European descent. Thus, we should pay attention to regional difference in allele frequencies, since rapid recent population growth has increased the load of rare variants in various regions and is likely to play a role in the individual genetic burden of complex disease risk.
We can draw the following lessons from the recent discoveries of disease-susceptibility gene with a large effect size, firstly in accordance with the large effect size, there is a tendency of familial clustering. Secondly, disease-relevant alleles could be missed by GWAS employing common SNPs and may be identified only by nucleotide sequence analysis of the human genome. These lessons strongly encourage the search for disease-susceptibility genes with large effect sizes based on comprehensive genome sequence analysis.
A well-powered rare variant association studies (RVAS) should involve a discovery set with at least 25,000 cases, together with a substantial replication set. To accomplish this goal, harmonised collaborative studies will be needed. In particular, given the diversities in regional distribution of rare variants in the human genome, we should emphasise the necessity of international collaboration to evaluate pan-ethnic as well as ethnicity-specific variants associated with complex trait diseases.
We expect identification of molecular basis of hereditary and sporadic neurological diseases should accelerate the research for developing new efficacious treatments directly targeting the disease mechanisms, which is highly expected to bring cures for currently devastating neurological diseases.
Department of Molecular Neurology, Graduate School
The University of Tokyo
Institute of Medical Genomics
International University of Health and Welfare, Japan
Please note, this article will appear in issue 32 of SciTech Europa Quarterly, which is available to read now.