Elsevier

The Lancet Neurology

Volume 11, Issue 1, January 2012, Pages 54-65
The Lancet Neurology

Articles
A C9orf72 promoter repeat expansion in a Flanders-Belgian cohort with disorders of the frontotemporal lobar degeneration-amyotrophic lateral sclerosis spectrum: a gene identification study

https://doi.org/10.1016/S1474-4422(11)70261-7Get rights and content

Summary

Background

Amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) are extremes of a clinically, pathologically, and genetically overlapping disease spectrum. A locus on chromosome 9p21 has been associated with both disorders, and we aimed to identify the causal gene within this region.

Methods

We studied 305 patients with FTLD, 137 with ALS, and 23 with concomitant FTLD and ALS (FTLD-ALS) and 856 controls from Flanders (Belgium); patients were identified from a hospital-based cohort and were negative for mutations in known FTLD and ALS genes. We also examined the family of one patient with FTLD-ALS previously linked to 9p21 (family DR14). We analysed 130 kbp at 9p21 in association and segregation studies, genomic sequencing, repeat genotyping, and expression studies to identify the causal mutation. We compared genotype-phenotype correlations between mutation carriers and non-carriers.

Findings

In the patient-control cohort, the single-nucleotide polymorphism rs28140707 within the 130 kbp region of 9p21 was associated with disease (odds ratio [OR] 2·6, 95% CI 1·5–4·7; p=0·001). A GGGGCC repeat expansion in C9orf72 completely co-segregated with disease in family DR14. The association of rs28140707 with disease in the patient-control cohort was abolished when we excluded GGGGCC repeat expansion carriers. In patients with familial disease, six (86%) of seven with FTLD-ALS, seven (47%) of 15 with ALS, and 12 (16%) of 75 with FTLD had the repeat expansion. In patients without known familial disease, one (6%) of 16 with FTLD-ALS, six (5%) of 122 with ALS, and nine (4%) of 230 with FTLD had the repeat expansion. Mutation carriers primarily presented with classic ALS (10 of 11 individuals) or behavioural variant FTLD (14 of 15 individuals). Mean age at onset of FTLD was 55·3 years (SD 8·4) in 21 mutation carriers and 63·2 years (9·6) in 284 non-carriers (p=0·001); mean age at onset of ALS was 54·5 years (9·9) in 13 carriers and 60·4 years (11·4) in 124 non-carriers. Postmortem neuropathological analysis of the brains of three mutation carriers with FTLD showed a notably low TDP-43 load. In brain at postmortem, C9orf72 expression was reduced by nearly 50% in two carriers compared with nine controls (p=0·034). In familial patients, 14% of FTLD-ALS, 50% of ALS, and 62% of FTLD was not accounted for by known disease genes.

Interpretation

We identified a pathogenic GGGGCC repeat expansion in C9orf72 on chromosome 9p21, as recently also reported in two other studies. The GGGGCC repeat expansion is highly penetrant, explaining all of the contribution of chromosome 9p21 to FTLD and ALS in the Flanders-Belgian cohort. Decreased expression of C9orf72 in brain suggests haploinsufficiency as an underlying disease mechanism. Unidentified genes probably also contribute to the FTLD-ALS disease spectrum.

Funding

Full funding sources listed at end of paper (see Acknowledgments).

Introduction

Amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) are two fatal neurodegenerative diseases without effective therapies. ALS is the most common neurodegenerative motor neuron disorder1 and FTLD has a similar prevalence to Alzheimer's disease in people younger than 65 years.2 These diseases are two extremes of a spectrum of clinically, pathologically, and genetically overlapping disorders,3 which suggests an overlap in disease mechanisms. Patients with ALS have reduced control of voluntary muscle movement due to progressive motor neuron degeneration in the motor cortex, brainstem, and spinal cord, resulting in muscle weakness and disturbances of speech, swallowing, or breathing. Up to 50% of patients with ALS have mild disturbances in executive function and some develop overt FTLD.4, 5 Symptoms of FTLD include behavioural, personality, and language disturbances and cognitive dysfunction caused by degeneration of frontal and temporal cortical neurons. Patients with advanced FTLD can also present with clinical signs of ALS.6 Although the diseases affect different neurons, TAR DNA-binding protein-43 (TDP-43) is a major constituent of neuronal deposits in both ALS and TDP-43-positive FTLD (FTLD-TDP), the most common pathological FTLD subtype.7, 8

Family-based linkage and population-based association studies have identified common genetic factors underlying ALS and FTLD. For example, mutations in the ALS genes TARDBP and FUS are occasionally noted in patients with FTLD8, 9, 10 and mutations in VCP (which is associated with an FTLD-related disorder) have been detected in ALS.11 Most striking is the finding that ALS and FTLD can occur within the same family or even the same patient. In more than 15 families worldwide, autosomal-dominant ALS and FTLD are causally linked with a major disease locus on chromosome 9 (ALSFTD2 locus).12, 13, 14, 15, 16, 17, 18 The minimal region linked in all these families is about 3·6 Mbp long and contains five known protein-coding genes (C9orf11, MOBKL2B, IFNK, C9orf72, and LINGO2; figure 1). Moreover, recent genome-wide association studies in ALS cohorts of different European origins have provided evidence for a major genetic risk factor in the same chromosome 9p region.19, 20, 21 We aimed to identify the disease-associated mutation from genetic studies of a family with FTLD-ALS conclusively linked to the 9p21 region and in an extended well characterised patient-control cohort from Flanders (Belgium).13 After initiation of our study, a Finnish study narrowed the associated region to a 232 kbp linkage disequilibrium block containing three known genes (MOBKL2B, IFNK, and C9orf72).19 Another such study implicated the same chromosomal region in an FTLD-TDP cohort,22 which was confirmed in other FTLD and FTLD-ALS cohorts.23 Two recent studies reported a GGGGCC repeat expansion in C9orf72 as the underlying genetic cause in chromosome 9 linked FLTD and ALS.24, 25

Section snippets

Participants and study design

The Flanders-Belgian cohort consisted of 337 patients with FTLD, 141 with ALS, and 23 with concomitant FTLD and ALS (FTLD-ALS).26, 27 Patients with FTLD were enrolled from 1998 onwards through an ongoing multicentre collaboration of neurology departments and memory clinics in Belgium (Hospital Network Antwerp Middelheim and Hoge Beuken, University Hospital Antwerp, University Hospitals Leuven, and University Hospital Ghent). Patients with ALS were recruited through the neuromuscular reference

Results

We excluded mutations in all five protein-coding genes in the 3·6 Mbp long minimal region (figure 1) through exon-based sequencing in family DR14. Subsequently, we sequenced 61 kbp of the most conserved non-coding sequences in the minimal region without finding patient-specific variants segregating with disease in family DR14. Whole-genome sequencing identified 120 new sequence variations in the minimal candidate region that segregated with disease, 37 of which were absent in controls.

We

Discussion

Previous genetic linkage and association studies have established a major locus for the spectrum of ALS and FTLD disorders on chromosome 9p21. In two reports,24, 25 a pathological GGGGCC repeat expansion in C9orf72 was shown to be the underlying genetic defect. Here, we report the independent finding of the same GGGGCC repeat expansion in patients with FTLD, FTLD-ALS, and ALS in a Flanders-Belgian cohort (panel). We identified several lines of evidence for the causative relation between the

References (44)

  • SM Rosso et al.

    Frontotemporal dementia in The Netherlands: patient characteristics and prevalence estimates from a population-based study

    Brain

    (2003)
  • C Lomen-Hoerth et al.

    Are amyotrophic lateral sclerosis patients cognitively normal?

    Neurology

    (2003)
  • GM Ringholz et al.

    Prevalence and patterns of cognitive impairment in sporadic ALS

    Neurology

    (2005)
  • D Neary et al.

    Frontotemporal lobar degeneration: a consensus on clinical diagnostic criteria

    Neurology

    (1998)
  • M Neumann et al.

    Ubiquitinated TDP-43 in frontotemporal lobar degeneration and amyotrophic lateral sclerosis

    Science

    (2006)
  • GG Kovacs et al.

    TARDBP variation associated with frontotemporal dementia, supranuclear gaze palsy, and chorea

    Mov Disord

    (2009)
  • T Van Langenhove et al.

    Genetic contribution of FUS to frontotemporal lobar degeneration

    Neurology

    (2010)
  • AL Boxer et al.

    Clinical, neuroimaging and neuropathological features of a new chromosome 9p-linked FTD-ALS family

    J Neurol Neurosurg Psychiatry

    (2010)
  • I Gijselinck et al.

    Identification of 2 loci at chromosomes 9 and 14 in a multiplex family with frontotemporal lobar degeneration and amyotrophic lateral sclerosis

    Arch Neurol

    (2010)
  • I Le Ber et al.

    Chromosome 9p-linked families with frontotemporal dementia associated with motor neuron disease

    Neurology

    (2009)
  • AA Luty et al.

    Pedigree with frontotemporal lobar degeneration—motor neuron disease and Tar DNA binding protein-43 positive neuropathology: genetic linkage to chromosome 9

    BMC Neurol

    (2008)
  • M Morita et al.

    A locus on chromosome 9p confers susceptibility to ALS and frontotemporal dementia

    Neurology

    (2006)
  • Cited by (0)

    View full text