Laboratory Diagnosis in Neurology, 1 Ed.

Monogenic Hereditary Neurological Disorders

In recent years, a growing number of neurological disorders have been recognized as monogenic diseases, i. e., they are caused directly by mutations of a single gene. These include the classic hereditary neurological disorders such as Huntington's disease or Wilson's disease, but also monogenic subforms of more complex syndromes such as spinocerebellar ataxia, spastic spinal paralysis, or various neuropathies.

Pathogenic Mutations

Expansions of Trinucleotide Repeats

Diseases. A number of hereditary neurological disorders, particularly those belonging to the neurodegenerative diseases, are caused by pathological expansion of unstable trinucleotide repeats in or near the coding region of a gene. These disorders include Huntington's disease (Fig. 8.2), several forms of spinocerebellar ataxia, myotonic dystrophy (Steinert's disease), and also Friedreich's ataxia (Table 8.1).

Trinucleotide Repeats. Units of three bases (trinucleotides, or triplets) represent the words of the genetic code, in which each triplet codes for one amino acid. DNA sequences consisting of multiple tandem copies of the same triplet are called trinucleotide repeats. For example, the first exon of the HD gene (or huntingtin gene) normally contains 15–25 tandem repeats of the CAG triplet coding for the amino acid glutamine. Through errors made by the DNA polymerase during cell division, such sequences are expanded (a shortening of the sequence may occur as well, but this happens less often).

Dysfunctions. Expansion of the CAG repeats to 39 or more tandem triplets in the huntingtin gene on one of the two chromosomes will cause the development of Huntington's disease in the person affected. According to the currently prevailing view, such trinucleotide repeat expansions result in dysfunctions that are not directly related to the primary function of the gene product. This genetic change is called gain-of-toxic-function mutation; it is probably caused by the pathological aggregation of extremely long polyglutamine chains in the nucleus (“nuclear inclusions”).

Special features. Diseases caused by trinucleotide repeats have several characteristics:

• Extent of expansion: The age at onset of the disease, and to a certain extent also the severity of the disease, depend on the extent of the trinucleotide repeat expansion. The larger the number of triplet elements, the earlier the onset of symptoms and the more severe the course of the disease. However, the range of expression is so large that one cannot predict the age at onset of symptoms in any given case.

• Anticipation: Trinucleotide repeat sequences are “unstable”—that means, the number of triplets often increases when the gene is passed on from one generation to the next (“dynamic mutation”). This instability, which is responsible for the formation of expanded repeats, also explains the anticipation phenomenon described in relation to Huntington's disease and myotonic dystrophy long before the discovery of mutations. “Anticipation” refers to the fact that gene carriers of subsequent generations develop the disease earlier and exhibit a more severe course than their predecessors.

Diagnosis. Trinucleotide expansions are relatively easy to detect by laboratory techniques (Fig. 8.3). This enables routine application of molecular diagnostic testing to confirm the clinical diagnosis and to determine the risk status of family members.

Point Mutations, Deletions, or Insertions

Diseases. Most hereditary neurological disorders can be caused by a number of different mutations (base exchanges, deletions, insertions, and other small changes in the DNA sequence) within a single gene (“allelic heterogeneity”). Examples are neurofibromatosis 1 and 2, Wilson's disease, and spastic spinal paralysis (Table 8.1).

Diagnosis. In these cases, direct DNA diagnosis can be carried out by complete sequencing of the gene in question, either to confirm the clinical diagnosis or establish a presymptomatic or prenatal diagnosis. Depending on the size of the individual gene, however, this search for mutations can be time-consuming and costly as genes with 40 or more exons are not uncommon. Hence, the most likely pathogenic mutations are analyzed first:

• Predilection sites: In some cases there are predilection sites for mutations, which can be examined first. For example, the microangiopathy called CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy) is caused by mutations in the large Notch3 gene which contains 29 exons. However, 70% of the mutations are found in exons 3 and 4; thus, molecular diagnostic testing is initially restricted to these two sequences.

• Type of mutation: In other cases, such as muscular dystrophy, certain common mutations may be identified by targeted use of molecular diagnosis. For example, muscular dystrophies of the Duchenne and Becker types are caused by different mutations of the dystrophin gene located on the short arm of the X chromosome (Xp21.1). With more than 70 exons, this is the largest gene of the human genome, and a complete analysis of its sequence is therefore extremely expensive. However, about 60% of all cases of these diseases are caused by rearrangements (large deletions, duplications) that are easier to detect. If no deletions can be found, a muscle biopsy remains the diagnostic method of choice to confirm the absence of dystrophin by immunohistochemistry or its abnormal size by western blot analysis.

Genetic Heterogeneity

Many hereditary neurological disorders are genetically heterogeneous, i. e., a defined clinical picture can be caused by mutations in several different genes.

Charcot-Marie-Tooth disease. One example is hereditary motor and sensory neuropathy type 1 (HMSN 1), now once again called Charcot-Marie-Tooth disease (CMT). The CMT1 form, which is associated with greatly slowed nerve conduction velocity, is in most cases caused by duplication of a 1.5-million base pair fragment on chromosome 17 (CMT1A). This fragment contains the gene for a myelin sheath constituent called peripheral myelin protein 22 (PMP-22). It is assumed that duplication of this chromosomal segment leads to overexpression of the otherwise intact gene and thus to disturbed myelination. In a small percentage of cases, however, point mutations in this gene are also responsible for the disease (allelic heterogeneity, see above), and in other cases mutations in entirely different genes, namely, the gene for the P0 protein on chromosome 1 or the gene for connexin 32 on the X chromosome, lead to the same clinical picture (phenotype): CMT1B and CMTX.

Limb-girdle muscular dystrophy. Even more variable is the genetic base of limb-girdle muscular dystrophy. Mutations in a multitude of genes largely coding for structural proteins of the muscle cell membrane (e. g., sarcoglycans α, β, γ, and σ, and other components of the dystroglycan–sarcoglycan complex) are responsible for diseases of this group that clinically cannot be clearly differentiated. In these cases, immunocytochemical analysis of a muscle biopsy continues to be the method of choice. A search for mutations usually follows only when the pathologically altered protein has already been identified. If the mutation is known, the genetic status of family members (including presymptomatic or prenatal diagnosis) can be determined without the invasive method of muscle biopsy.

This genetic heterogeneity is rather the rule than the exception: it is seen in many hereditary neurological disorders. Currently, any molecular subclassification that might be developed on this basis would have no consequences for therapy. This will change, however, when causal treatment strategies are developed on the basis of the molecular defects.