Motor Neurone Disease

Question 1

Motor Neurone Disease

Answer 1

• Age related condition with selective irreversible loss of a specific group of neurones. Late onset and relentless progression ALS linked with amyotrophy (muscle wasting) and lateral sclerosis
• No clear understanding or aetiology but likely linked with complex interaction between genetically determined predisposition and environmental triggers
  
  • 95% sporadic
  • 5% inherited (AD) mendelian inheritance - most common 9p chromosome C9ORF72 mutation
    
    • other mutations: SOD1, TLS/FUS, ANG, TDP43
    • Sometimes there is incomplete penetrance and therefore sporadic disease may have genetic link

Question 2

Clinical features of MND

Answer 2

• Progressive degeneration of the motor system with mixed UMN and LMN signs with fasciculations
  
  • UMN: increased tone decrease power and increased reflexes
  • LMN: Decreased bulk, tone, power and reflexes

Sparing of intellect (15% get FTL dementia, 35% get cognitive dysfunction), sensation, sphincter function and eye movement. This results in progressive, painless paralysis and wasting 80% limb onset 20% bulbar onset (speech and swallowing problems)

Question 3

4 main phenotypes of MND

Answer 3

4 main phenotypes

• (75%) Amyotrophic lateral sclerosis (ALS): mixed UMN and LMN degeneration. Presents with progressive weakness and wasting in any distribution. Retained weak reflex and wasted muscles. Fasciculations and muscle cramps prominent. Foot drop, proximal Upper and lower limb weakness, flail arms
• (20%) Progressive bulbar palsy (PBP): speech and swallowing difficulties progression to dysarthria over 6-24 months and dysphagia (1-3 years)complicated by pneumonia, weight loss and anorexia
  
  • In about 12 months, you will get the other presentations as well. However, PBP gets shorter life expectancy
• (3%) Progressive myotrophic atrophy (PMA): LMN degeneration
• (2%) Primary Lateral sclerosis (PLS): UMN degeneration dominated by weakness and spasticity

Question 4

Genetic causes:

Answer 4

FUS

SOD1

C9ORF72

TDP-43

Question 5

FUS

Answer 5

Tsubota et al

FUS (fused in sarcoma) normally involved in gene expression and RNA processing.

· FUS contains a C-terminal NLS, this is recognised by transportin a nuclear transport receptor that localises FUS to the cell nucleus. In mutant FUS this terminal leads to relocation into the cytoplasm, cell stress for example heat shock oxidative stress leads to the formation of large FUS inclusions. These inclusions are thought to cause motor neuron cell death and lead to ALS pathogenesis. Toxicity is likely to be caused by loss of function in the nucleus and gain of toxic function through aggregates in the cytoplasm.

· FUS inclusion are also seen in FTLD -but in absence of FUS mutations

Question 6

SOD1

Answer 6

Tsubota et al

SOD 1 – Three main pathways to toxicity:

· Neurotoxicity occurs trough the mis-localisation of SOD1 to the extracellular space, mitochondria and ER.

· Microgriosis: This occurs through the secretory pathway where chromogranin a neuronal secretory protein chaperones mutant SOD1 to the extracellular space. The extracellularly secreted SOD1 mutants trigger microgliosis and neurotoxicity

· Mitochondrial dysfunction: Macrophage migration inhibitory factor (MIF) is a chaperone that erroneously places mutated SOD1 to the outer membrane of mitochondria. Binding of SOD1 to the mitochondrial membrane causes conformational changes in Bcl-2, this inhibits the interaction between Bcl-2 and Voltage-dependent anion channels, resulting in cytotoxity.

· ER stress: SOD1 mutants interact with BiP, an ER chaperone or Derlin-1 a component protein of the ERAD complex. These interactions cause ER stress by disrupting protein folding in the ER and endoplasmic reticulum- associated degradation (ERAD), the role of which is to degrade unfolded proteins in the ER.

Question 7

TDP43

Answer 7

TDP-43

• TDP-43 was identified in the abnormal protein aggregates that occur in ALS patients and mutations of TARDBP, which encodes TDP-43, have been shown to cause ALS. This was found in both familial and sporadic ALS. TDP-43 is also a component of the ubiquitin-positive inclusion found in other neurodegenerative diseases, especially fronto-temporal lobar degeneration.
  
  • Normal function of tDP-43:
    
    • RNA processing.
    • Associated with heterogeneous nuclear ribonucleoproteins (hnRNPs) e.g. transcription, splicing, transport and localisation.
    • Simulate dendritic growth and synapse formation as seen in Drosophila
  • It is believed that several burdens e.g. ER stress, genetic impairment of TDP-43, disruption of protein degradation in cells facilitate the aggregation and stabilisation of TDP-43. Because TDP43 aggregations have been reported to include non-specific RNAs and RNA binding proteins, the formation of aggregates could exhibit cell toxicity by isolating the essential factors for RNA metabolism. A reduction in functional TCP-43 through mutations could also suppress the original neuroprotective function of TDP43
  • Mutation: Over 30 mutations of TARDBP reported in ALS are clustered in the glycine-rich domains, indicating the importance of the C-terminal region in ALS pathogenesis
    
    • The c-terminal glycine-rich domain is essential for the interaction with some RNA binding proteins and the slicing activity of TDP-43
    • This plays an important role in mediating protein-protein interactions – a prion like propagation ability.
    • Insoluble TDP-43 isolated from patients with TDP-43 proteinopathy serves as the seed of the aggregation, accumulating the endogenous intact TDP-42 in abnormal aggregations.
  • ER stress is also suggested to be involved in the formation of TDP-43 aggregates.
    
    • Weak ER stress in the early stage of neurodegenerative disease is reported to induce TDP-43 dysfunction and modulate its distribution. These impairments lead to greater ER stress through prion-like propagation and subsequent stabilization of aggregates.
    • Thus ER stress could facilitate the accumulation of TDP-43 and cell toxicity.
  • Post translational modifications of TDP-43 were found only in inclusions.
    
    • A biochemical analysis found that phosphorylation of S409/410 was identified as the major modification of TDP-43 aggregates.
    • It was also revealed that TDP-43 is cleaved abnormally into a 17- to 25-kDa C-terminal fragment (CTF) mediated by caspase-3 (this is activated in apoptotic conditions and in the late stage of disease)
    • Most inclusions including pathological 25-kDa CTF are recognised by the phosphorylated TDP-43 specific antibody.
    • Although the relationship between toxicity of phosphorylated CTF aggregation and its effect on disease progression is still unclear, the pathological characteristics of TDP-43 proteinopathy (e.g. cytoplasmic accumulation, insolubility, over-phosphorylation, poly-ubiquination, and abnormal toxicity) are present in cells expressing the 25-kDa CTF
  • Wild type TDP43 mainly localises to the nucleus, however ALS-related TDP-43 mutants typically translocate to the cytosol and form aggregates in the early stages of ALS
• Summary: TDP43 shuttles between cytoplasm and nuclear, and mutation or dysregulation of the amount causes the aggregation formation of TDP-43 in cytoplasm. TDP43 positive aggregations contain an abnormally cleaved and phosphorylated CTF

Question 8

C9ORF72

Answer 8

• Mutation: C9ORF72 mutation: The GGGGCC (G4C2) hexa-nucleotide repeating expansion in the intron region of chromosomal 9 open reading frame 72 was reported as a cause of ALS and also found in FTLD. In normal subjects, the G4C2 hexa-nucleotide is repeated 2 to 3 times, and this is never repeated more than 20 times, however, abnormal expansions is repeated up to hundreds or thousand times in patients with C9ORF72-associated ALS
• Loss of function toxicity: C9ORF72 is a homologue of “differentially expressed in normal and neoplasia (DENN)”, a GDP/GTP exchange factor that activates Rab-GTPase, and it is involved in endosomal trafficking. Therefore mutations are considered to disrupt vascular trafficking. However, it is still unclear whether loss of function of C9ORF72 causes motor neuron degeneration and further studies are required
• Gain of function toxicity: The toxicity of abnormal RNA repeat expansions can be observed in several diseases. In the case of C9ORF72 mediated ALS, RNA foci are observed in neurons derived from iPSC as well as in spinal and frontal cortex neurons of ALS patients. It has been proposed that C9ORF72 RNA containing a G4C2 repeat expansion also sequesters essential proteins and disrupts RNA metabolism to cause nucleolar stress.
  
  • However, toxicity of abnormally expanded dipeptide repeats was also reported. Gly-Ala DPR positive inclusion were one of the few positive inclusion found in the brains of ALS and FTLD patients (along with TDP-43 negative/p62 positive inclusions). Different DPRs promote different kinds of toxicities and the expression of Gly-Ala DPR has been reported to evoke ER stress through activating PKR-like endoplasmic reticulum kinase pathway (PERK pathway).
• Summary: The intronic hexanucleotide repeat expansion in C9ORF72 causes the formation of RNA foci in the nuclear. RNA foci might sequester RNA binding proteins that are essential for cell survival. Dipeptide repeat proteins (DPRs) are generated from aberrant repeat expansion through Ran translation, evoking ER stress. DPR-positive aggregation is colocalised with p62 in the cytosol. The interaction of abnormal RNA repeat expansion with ribonucleoproteins is considered to cause nucleolar stress. A defect in nucleocytoplasmic transport has also been reported as the toxicity if the C9ORF72 mutation.

Question 9

Investigation in MND

Answer 9

• Diagnosis of MND is based on thorough clinical evaluation, but may be a diagnosis of exclusion.
• Bloods, scans, LP, imaging negative
• Electrophysiology remains an important tool in the evaluation of patients presenting with signs and symptoms of motor neuron disease (MND).
• The electro diagnostic study should include peripheral nerve conduction studies (NCS) and needle electromyography (EMG) to both exclude treatable disease and gather evidence toward a diagnosis of amyotrophic lateral sclerosis (ALS).
• Nerve conduction studies:
  
  • Motor conduction studies are an essential part of the electro diagnostic evaluation of a patient suspected of having a MND. These studies allow the exclusion of treatable neuropathies, such as multifocal motor neuropathy with conduction block, from the differential diagnosis. Common findings observed from motor NCS in patients with MND include asymmetric side-to-side Compound Muscle Action Potential (CMAP) differences, or CMAPs with decreased amplitude, prolonged distal motor latency, and slowed conduction velocity consistent with axon loss.
  • Sensory conduction studies should include the ulnar and sural nerves; it is generally accepted that sensory nerves are normal in ALS – although this may not always be the case.
• EMG:
  
  • Needle electromyography is the most important component of the electro diagnostic evaluation in MND. It allows identification of LMN involvement often before it becomes clinically evident, extending the physical examination and enabling earlier diagnosis.
  • However, two EMG features are required for confirmation of neurogenic change consistent with a diagnosis of ALS:
    
    • Evidence of chronic neurogenic change.
    • Evidence of acute denervation.

Question 10

Management of MND

Answer 10

• Riluzole
  
  • Riluzole is the only drug identified to have a beneficial effect on survival, following a double blind, randomized placebo controlled trial in patients with the common amyotrophic lateral sclerosis variant of motor neuron disease.
  • The effect is modest, with a prolongation of life of approximately 3-4 months on average. Lacomblez L. et al
  • It is generally well tolerated and it is now standard practice in the UK to commence Riluzole therapy in all amyotrophic lateral sclerosis patients following diagnosis.
  • These conclusions were confirmed by a Cochrane review examining evidence from four randomized clinical trials involving 1477 patients and are supported by NICE guidelines in the UK.
• Respiratory care
  
  • Use of non-invasive ventilation, in which the patient uses a mask ventilator system (usually bilateral positive airway pressure) overnight during sleep.
  • This prolongs life
    
    • A randomized controlled trial found a median survival benefit of about seven months in patients with good bulbar function using non-invasive ventilation.
    • The extended survival was associated with an improvement in multiple quality of life measures—an important observation as one does not wish to extend life merely to prolong suffering. This effect on survival is much greater than that currently provided by neuroprotective treatments such as Riluzole.
• Other supportive measures: Nutrition and feeding, MDT, end of life care