Three Families With Amyotrophic Lateral Sclerosis and Frontotemporal Dementia With Evidence of Linkage to Chromosome 9p FREE

Amyotrophic lateral sclerosis (ALS) is the most frequently observed motoneuron disorder worldwide, with an estimated prevalence of 8 or 9 per 100 000 in the general population. In ALS, both upper and lower motoneurons selectively degenerate in the brainstem, spinal cord, and motor cortex. This incurable disease is progressive and eventually leads to death, typically from respiratory failure 3 to 5 years after the onset of symptoms. Although ALS is most frequently sporadic, approximately 15% of cases are familial. Of these, the most frequently identified cause is mutations in the copper-zinc superoxide dismutase 1 (SOD1) gene¹; additionally, a mutation in the vesicle-associated membrane protein-associated protein (VAPB) gene has also been identified in familial ALS.²

Frontotemporal dementia (FTD) is increasingly recognized as a common type of dementia, after Alzheimer disease and vascular dementia. The clinical manifestations of FTD are highly variable and can include impairments in executive function and language, as well as behavioral dysfunction (eg, impaired personal conduct or interpersonal social behavior). In most cases, the course of the disease is approximately 8 to 10 years and involves the degeneration of neurons in the frontal or anterior temporal lobes. Survival is typically about 3 years after onset of symptoms.³ Mutations in 3 different genes were primarily reported in FTD. The valosin-containing protein (VCP) gene, on chromosome 9p13.3, leads to FTD with Paget disease of the bone⁴; the chromatin-modifying protein 2B (CHMP2B) gene, on chromosome 3p11.2, leads to classic FTD,⁵ as does the progranulin (PGRN) gene, on chromosome 17q21.31.^6- 7 In the CHMP2B gene, mutations were also observed in patients with ALS-like motoneuron degeneration.⁵ About one fourth of FTD probands screened for PRGN had a family history of motoneuron disease, but none within this subset of probands exhibited PGRN mutations.⁷ Mutations in the microtubule-associated protein tau (MAPT) gene, also at chromosome 17q21.31, caused either ALS or FTD,^8- 9 as well as Parkinson disease, Pick disease, and progressive supranuclear palsy.

In 2 unrelated families, ALS and FTD were reported to show evidence of linkage to chromosome 9p. A Scandinavian family had 5 members with ALS and 9 with FTD, and no family members had both ALS and FTD.¹⁰ A genomewide scan performed in this family enabled the identification of a 21.8-centimorgan [cM] (14-megabase [Mb] pairs) locus on chromosome 9p21.3-p13.3. The same locus was identified in another family from the Netherlands.¹¹ This second family has 12 affected members, including 7 with ALS only, 2 with FTD only, and 3 with both ALS and FTD. By comparing the 2 haplotypes, a candidate interval of 12 cM (11 Mb pairs) between markers D9S2154 and D9S1874 was established.¹¹

Informed consent was obtained from all participating family members in each family. These forms were approved by the ethics committee of the Centre Hospitalier de l’Université de Montréal, Montreal, Quebec. DNA was extracted from blood samples using standard methods. The diagnosis of ALS in members of each family was considered definite or probable per El Escorial World Federation of Neurology criteria.¹² Individuals with FTD exhibited cerebral atrophy at neuroimaging, cognitive defects and verbal deficits, and mutism at the end stages of disease.

The Que-23 family is from Canada and consists of 34 collected individuals (Figure 1). Five family members have ALS only and 3 others have FTD only. Mean age at onset of ALS in affected individuals was 55.8 years (age range, 46-58 years), with a mean disease duration of 2.4 years (range, 1.5-3 years). The Fr-104 family is of Spanish origin, though the information was collected in France (Figure 2). The pedigree contains 4 affected siblings and 1 affected cousin. Patients had either bulbar or spinal onset of disease. One patient with ALS (patient III-5 in Figure 2) had shown early signs of FTD, namely, disinhibition and impairment in executive function. The Que-1 family consists of 43 collected members (Figure 3). Genotypes for 8 affected individuals have been collected or can be confidently inferred based on the genetic information of their spouses and children. Among them, 5 had ALS only and 3 had ALS and FTD. The family is French-Canadian and resided in Quebec for several generations. The mean age at onset of disease was 54.3 years (age range, 45-63 years), with a mean disease duration of 4.8 years (range 2-9 years). Both bulbar and spinal onset was observed in individuals with ALS.

Linkage analysis was performed using 3 liability classes based on previously reported values¹¹: class 1, age 0 to 46 years, 5% penetrance; class 2, older than 46 years, 40% penetrance; and class 3, spouses, who were given the background population risk. GeneHunter software (version 2.1; Ward Systems Group Inc, Frederick, Md) was used for multipoint analysis, but, because of the constraints in pedigree size that can be used in GeneHunter, only affected individuals and their spouses and children of an unsampled affected group were included.¹³

Before inclusion in this study, all families were screened to exclude the presence of mutations in the following genes: SOD1 at chromosome 21q22.1, VAPB at chromosome 20q13.3, MAPT at chromosome 17q21.1, and VCP at chromosome 9p13-12. No mutation was identified in any of the 4 genes. Furthermore, linkage analysis excluded the ALS6 locus on chromosome 16 using markers D16S757, D16S771, and D16S3057 and the ALS3 locus on chromosome 18 using markers D18S1103, D18S858, and D18S1144 (Table 1).

Family Que-23 is composed of 2 collected affected individuals and 6 reconstructed affected individuals and showed a maximum logarithm of odds (LOD) score of 2.81 at D9S1121 (θ = 0). Recombination events in affected individuals placed this disease haplotype between markers D9S157 and D9S1805. Multipoint analysis in the affected members of this family increased the LOD score to 3.01 spanning markers D9S1679 to D9S1791.

The Que-1 family contained 2 collected affected individuals, 6 reconstructed affected individuals, and 33 unaffected family members. All affected individuals shared 12 markers across the ALS-FTD locus on chromosome 9. The upper recombinant occurred in individual V:1 (Figure 3) at D9S1869. This extended beyond the current upper limit of the region as previously reported at marker D9S2154.¹¹ The lower recombinant was between markers D9S1791 and D9S1862. The maximum 2-point LOD score identified for the family was 2.51 at θ = 0 for marker D9S1118.

Family Fr-104 had an upper recombinant at D9S171; the lower boundary in this family extended into the q arm far beyond the current lower limit at this locus. The 5 affected members yielded a maximum LOD score of 1.55 at marker D9S1791. Table 2 compares haplotypes observed for these families with those previously reported.

In combining the 3 families, the maximum GeneHunter multipoint LOD score obtained was 7.22, ranging from D9S1121 to D9S1791 for a 15.1-cM interval (Figure 4). The same region is defined when considering the recombination breakpoints of D9S171 for family Fr-104 (Figure 2) and D9S1805 for family Que-23 (Figure 1). This corresponds to a 9.7-Mb locus containing 35 genes (UCSC Genome Browser, May 2004 freeze; University of California at Santa Cruz). If marker D9S2154 is considered as a flanking marker, based on a recent publication,¹¹ the number of known genes is decreased to 33. If nonparametric linkage is considered for the locus, the result is a z value of 14.2 centered on D9S1121.

Among the genes in this newly defined region, aprataxin (APTX), BCL2-associated athanogene 1(BAG1), and ciliary neurotrophic factor receptor (CNTFR), which were previously screened in other families, were closely examined for the presence of mutations in our families. Given that one of its functions was pertinent in another ALS gene, the tyrosine kinase endothelial (TEK) gene was also selected for mutation screening. None of these 4 genes contained mutations in exons or 50 base pairs past intron or exon junctions that could be causative.

The 3 families studied are relatively large from an ALS standpoint. Considering that 2 separate teams recently reported families also mapping to chromosome 9p, this locus may have a significant effect on ALS and FTD research. Bearing in mind that a rapidly growing number of families are reported to have linkage to the chromosome 9p locus, it is increasingly plausible that a single gene is responsible for the joint occurrence of the 2 diseases rather than 2 separate genes with independent mutations. It is likely that additional medium-sized families with ALS or FTD will also map to this locus and aid in the reduction of what is still a sizeable region. Similar to what was observed in the other families with ALS or FTD linked to chromosome 9p, 2 of the new families reported here exhibit an overlap of ALS and FTD in several patients, while in the third family, patients either develop ALS or FTD but not both.

A promising gene lies within this reduced candidate interval, namely, the TEK gene. This is based on its similarity to the vascular endothelial growth factor (VEGF) gene. The VEGF gene is ALS-related inasmuch as a haplotype of single nucleotide polymorphism (SNP) in its 5′ region was significantly associated with the disease.¹⁴ Furthermore, deletion of the hypoxic response element of VEGF resulted in an ALS-like motoneuron disease phenotype in mice.¹⁵ Not only do TEK and VEGF physically interact, but TEK also contains a hypoxic response element in its promoter region, enabling its translation to remain unaffected in hypoxic conditions.¹⁶ The association and related regulation of VEGF and TEK raises the intriguing possibility that TEK may have a role in ALS and FTD; however, no mutations were observed within TEK or in its upstream promoter region in our families.

Other genes in the newly defined 9p locus will be further prioritized for mutation screening by sequencing in our 3 new families with ALS, FTD, or both, particularly those genes expressed in the brain. Finding the mutation and the responsible gene would have a substantial effect on understanding the etiology of these 2 diseases.

Correspondence: Guy A. Rouleau, MD, PhD, Center for the Study of Brain Diseases, University of Montréal, Centre Hospitalier de l’Université de Montréal (CHUM) Research Center, Notre-Dame Hospital, J. A. de Sève Pavilion, Room Y-3633, 1560 Sherbrooke St E, Montreal, Quebec, Canada H2L 4M1 (guy.rouleau@umontreal.ca).

Accepted for Publication: October 2, 2006.

Author Contributions:Study concept and design: Valdmanis and Rouleau. Acquisition of data: Valdmanis, Bouchard, Camu, Salachas, Meininger, Strong, and Rouleau. Analysis and interpretation of data: Valdmanis, Dupre, and Rouleau. Drafting of the manuscript: Valdmanis. Critical revision of the manuscript for important intellectual content: Dupre, Bouchard, Camu, Salachas, Meininger, Strong, and Rouleau. Statistical analysis: Valdmanis. Obtained funding: Rouleau. Administrative, technical, and material support: Valdmanis, Dupre, Meininger, and Rouleau. Study supervision: Bouchard, Salachas, and Rouleau. Family collection: Camu and Strong.

Financial Disclosure: None reported.

Funding/Support: This study was supported by the ALS Association, the Muscular Dystrophy Association, the Canadian Institutes of Health Research (Mr Valdmanis and Dr Rouleau), the Association Pour la Recherche sur la Sclerose Laterale Amyotrophique, the Association Francaise Contre les Myopathies, and the French Group on MND.

Acknowledgment: We thank Patrick Dion, PhD, for helpful comments about the manuscript; Judith St-Onge for technical assistance; and Melanie Benard, BA, for extensive sample collection. The DNA extraction and cell lines were provided by Généthon.

This article was corrected for error in data on 4/24/2007, prior to publication of the correction in print.