Targeted high throughput sequencing in hereditary ataxia and spastic paraplegia

Targeted high throughput sequencing in hereditary ataxia and spastic paraplegia


Hereditary ataxia and spastic paraplegia are heterogeneous monogenic neurodegenerative disorders. To date, a large number of individuals with such disorders remain undiagnosed. Here, we have assessed molecular diagnosis by gene panel sequencing in 105 early and late-onset hereditary ataxia and spastic paraplegia probands, in whom extensive previous investigations had failed to identify the genetic cause of disease. Pathogenic and likely-pathogenic variants were identified in 20 probands (19%) and variants of uncertain significance in ten probands (10%). Together these accounted for 30 probands (29%) and involved 18 different genes. Among several interesting findings, dominantly inherited KIF1A variants, p.(Val8Met) and p.(Ile27Thr) segregated in two independent families, both presenting with a pure spastic paraplegia phenotype. Two homozygous missense variants, p.(Gly4230Ser) and p.(Leu4221Val) were found in SACS in one consanguineous family, presenting with spastic ataxia and isolated cerebellar atrophy. The average disease duration in probands with pathogenic and likely-pathogenic variants was 31 years, ranging from 4 to 51 years. In conclusion, this study confirmed and expanded the clinical phenotypes associated with known disease genes. The results demonstrate that gene panel sequencing and similar sequencing approaches can serve as efficient diagnostic tools for different heterogeneous disorders. Early use of such strategies may help to reduce both costs and time of the diagnostic process.


Fig 3Fig 1Table 1Table 2Table 3Table 4Fig 2Fig 3Fig 1Table 1Table 2

Citation: Iqbal Z, Rydning SL, Wedding IM, Koht J, Pihlstrøm L, Rengmark AH, et al. (2017) Targeted high throughput sequencing in hereditary ataxia and spastic paraplegia. PLoS ONE 12(3): e0174667. doi:10.1371/journal.pone.0174667
Editor: Klaus Brusgaard, Odense University Hospital, DENMARK
Received: December 23, 2016; Accepted: March 13, 2017; Published: March 31, 2017
Copyright: © 2017 Iqbal et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Data Availability: All relevant data are within the paper and its Supporting Information files.
Funding: Z.I., I.M.W., and L.P. have been supported by grants from the South-Eastern Norway Regional Health Authority. M.T. and A.H.R. have received funding from the Research Council of Norway. M.T. is also supported by a career fellowship from the South-Eastern Norway Regional Health Authority. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.

The spinocerebellar degenerative disorders; hereditary ataxias (HA) and hereditary spastic paraplegias (HSP) are heterogeneous disorders causing progressive gait difficulties due to degeneration of the cerebellum, corticospinal tracts, brainstem, and/or spinal cord [1]. These disorders are relatively rare with an estimated total prevalence of 13.9/100,000 in southeast Norway [2]. HA is characterized by progressive limb and gait ataxia, loss of coordination and disturbances of speech and oculomotor control. HSP is characterized by progressive spasticity and weakness of the lower limbs, the weakness often being mild relative to the spasticity [1, 2]. Onset is reported at all ages, and all monogenic modes of inheritances—autosomal dominant, autosomal recessive, and X-linked—have been identified [3]. To date, pathogenic variants in more than 100 genes have been identified in spinocerebellar degenerative disorders [4–7]. Identifying molecular diagnoses in such genetically heterogeneous disorders is challenging. Usually multitier, expensive and time-consuming investigations are performed. Nevertheless, a large number of affected individuals remain without a molecular diagnosis.

With the progress in sequencing technologies, there are several methods available to screen hundreds or thousands of genes at once and possibly identify a molecular diagnosis in a shorter time period at lower costs. Gene panel sequencing (GPS) or targeted high throughput sequencing, whole-exome sequencing (WES), and whole-genome sequencing (WGS) methods are currently being used by researchers and diagnostic laboratories. These methods have different advantages related to quality and interpretation of data, management of ethical issues, and economic effectiveness. Besides other high throughput sequencing methods, GPS has been proven successful in several heterogeneous neurological disorders [8–10].

In the present study, we have evaluated the use of GPS in 105 clinically well-characterized probands affected with HA or HSP in whom previous extensive investigations had failed to identify a genetic cause. The study provides insights into the value of this diagnostic strategy and illustrates the diversity of genetic causes of spinocerebellar degenerative disorders.



In 2002, a research study was initiated at the Department of Neurology, Oslo University Hospital, carefully registering patients with HA and HSP in Norway. In 2014 the database consisted of 683 individuals with a diagnosis of HA and HSP, of whom 446 were probands [2]. The database has been designed to comprehensively cover the South-Eastern Norway health region where 55.8% of the Norwegian population lives. In addition, patients have been referred from the rest of the country since 2002. Main inclusion criteria for HA were cerebellar gait and/or limb ataxia, and for HSP, spasticity in the lower limbs, brisk reflexes and positive Babinski sign [11, 12]. In addition, most of the included probands had a known family history of disease. A minority had sporadic disease, which after thorough investigation was considered compatible with a hereditary type of spinocerebellar degenerative disorder. 17% of the HA probands and 37% of HSP probands had an exact genetic diagnosis (Fig 1) at start of the present study. Molecular investigations were carried out according to what was diagnostically available at the time of examinations. All HA probands were previously screened for SCA1, SCA2, SCA3, SCA6, SCA7, and for Friedreich ataxia in recessive and sporadic cases. HSP probands were screened for variants in the genes linked to SPG4, SPG3A, and most also for SPG31. To detect gene-dosage defects, multiple ligation-dependent probe amplification (MLPA) was performed in all HSP probands for SPG4 and SPG3A. Additional molecular tests were performed depending on the phenotype and the pedigree structure, including variants in the genes linked to; SPG7, SPG1, SPG2, FXTAS, POLG, SCA8, SPG11, AOA1, AOA2, Ataxia Telangiectasia, ARSACS, SPG8, DRPLA, and SPG42. Array comparative genomic hybridization (aCGH) was performed in all probands with cognitive impairment. Also, biochemical tests for metabolic disorders such as adrenoleukodystrophy and gangliosidosis, as well as biomarkers as carbohydrate-deficient transferrin, albumin, cholesterol, gamma globulins, alpha-fetoprotein and vitamin E were tested when relevant. Brain magnetic resonance imaging (MRI) was performed in most of the probands.

Full article at