Suspected Diagnosis of Leigh Syndrome

Question :

You are presented below with a case study, including a pedigree, of a specific genetic condition. You will need to Research and critically evaluate the literature on the clinical and molecular basis of the condition.

Analyse and interpret the pedigree, then discuss the possible modes of inheritance for the disease.

Case

A nine-month-old male child, born from a consanguineous marriage, presented with seizures, delayed developmental milestones, and regression of achieved milestones. He was the 5th child to his parents and had two brothers that died within the first 12 months of life due to respiratory failure and an uncle who died at 3 years with similar symptoms.

In view of the above history, a genetic disease was considered, and he was investigated accordingly. A family history and pedigree analysis are shown below.

Magnetic resonance imaging (MRI) demonstrated characteristic symmetrical necrotic lesion in the basal ganglia and brainstem leading to a suspected diagnosis of Leigh syndrome.

Answer :

Analysis and interpretation of the pedigree, the possible mode of inheritance for the Leigh Disease

• Punnett Squares helps to determine the inheritance pattern in given pedigree (Annetta et al, 2010). According to Punnett Square, for example, Xc indicates recessive gene that results into Leigh syndrome (LS) and X+ indicates the normal dominant gene. Females with X+X+ or X+Xc show no LS, alongside XcXc females affect by LS. X+Y containing males have no LS, males with XcY affect by LS.

• The given pedigree discloses X-linked recessive inheritance. These defective traits frequently transferred from a mother who is carrying and son gets the defects. The transfer of X-linked genes between father and son never happened. And moreover, the probability of getting affected by LS in males is more over females comparatively.

• The following observations were made from the given pedigree.

• The II generation female 4 is a carrier of recessive gene X+ Xc is on consanguineous mating with male 3 in the same generation.

• The off springs of 3 & 4 of II generation are one female (4) and two male (5&6) in III generation.

• The off springs of this couples are in III generation.

• In this generation III the male child (6) was died at the age of three years with Leigh Syndrome.

• Female 4 in III generation becomes a carrier of recessive gene X+ Xc, while male 5 is not affected may be because of X+Y dominant gene but male 6 is affected may be because of Xc Y recessive gene inherited from mother.

• The IV generation off springs are from mating 3&4 of III generation, which is again consanguineous mating.

• Here, in this generation III, female carries recessive gene X+ Xc , so all three male off springs (1,4 &5) are affected by the syndrome because they inherited Xc Y. females are not affected in generation IV.

• Based on these findings, it was concluded that it might be due to X –linked recessive inheritance.

• X –linked recessive inheritance pattern was observed in least number of LS cases as per the literature.

There are 2 possible inheritance patterns have been identified, one is X-Linked inheritance pattern and Mitochondrial Inheritance pattern.

The first one, that is X-Linked inheritance pattern is already explained in the above description and the latter i.e. Mitochondrial inheritance pattern was apparent in around twenty percentage of humans with LS. This kind of inheritance pattern is also known as Maternal Inheritance pattern.

Developing embryo needs mitochondrial contribution, egg cells mitochondria involves in development of embryo but not of the sperm cells (Dumollard et al, 2007). In such cases the off-springs inherit genetic defects occurring in mtDNA mutations can only be from their mother. The disorders arising from these defective genes become apparent through all the generations of that particular family and both the male and female genders are at risk, and also these mutated traits are not going to be transferred by the fathers to his children. Spontaneous mutations may also cause LS in the family without history of LS. This particular maternal pattern of inheritance implies to genes of mitochondrial DNA (mtDNA).

Research and critically evaluation of the literature on the clinical and molecular basis of the condition.

Leigh syndrome is also known as Leigh disease or sub-acute necrotizing encephalo-myelopathy (Rahman et al, 1996).  This is a rare genetically inherited neuronal dependent metabolic disorder which cause the Central Nervous System (CNS) functions to be impaired.  

Signs and symptoms:

Appears in infancy and would lead to death within a window of years. However so far it was understood that symptoms can be apparent at any age including adolescence and in most of the cases, patients survived for years after diagnosis (Finsterer, 2008). Generally, symptoms were observed often initially after a triggering episode that attacks the body’s ATP producing pathways like surgery or an infection (Lake et al, 2016).

The normal course of LS includes central nervous system anatomical regression in periods of difficulty in metabolic pathways for energy production. Some individuals show no disease progression for many years while the other patients may develop progressive worsening (Baertling et al, 2014). 

Symptoms in infants usually are vomiting, dysphagia, difficulty in swallowing and diarrhea leading to failure of development. Seizures are often seen and excess lactate seen in CSF, blood and urine of LS patients, hypotonia and dystonia are also seen (Sofou et al, 2014).

Genomics:

The main causes of Leigh Syndrome are mitochondrial DNA (mtDNA) mutations, Nuclear DNA mutations, like gene SURF1 and cytochrome c oxidase assembly factors (Martinelli et al, 2012). Disorders of oxidative phosphorylation, which is the process where cell produce the source of energy like ATP, ADP, will develop due to mutations in the genes. Mutations in mtDNA and Nuclear DNA encoded genes for oxidative phosphorylation account for the disease (Thorburn et al, 2017). It is always may not be possible to detect the mutation in particular gene in particular individual. Four protein complexes that are involved in the oxidative phosphorylation has shown impaired functions in the Leigh Syndrome. Five protein complexes such as complex I to V which during oxidative phosphorylation, drive the production of adenosine triphosphate (ATP), through pathway by transfer of electrons (Ruhoy & Saneto, 2014).  So far the findings of LS reveals that the mutations in the genes encoded for these complexes resulted in altered sequence of amino acids and their assembly. 

Either mal formed protein or misplacement in the sequence of these complexes which are made of amino acids, irrespective of the genetic bases, it resulted the inability of the complexes affected by the mutations to affects the oxidative phosphorylation process. In the LS, crux regions that are affected more are the brain stem and basal ganglia than other regions. This causes the sever drop down of energy in the cells which lead to necrosis of cells and in turn impairs the nervous and motor functions (Quintana et al, 2010). The heart and muscle cells which demand more energy are directly impacted by cell death caused by sever energy deficiency in LS.

One third of the cases of LS accounts for the complex I disruption. There are around 25 genes that translate the complex I from Nuclear or mitochondrial DNA.

Mitochondrial DNA mutation:

The function of mitochondria is to metabolize carbohydrates, amino acids and fatty acids to generate ATP (Zhang et al, 2016). The self mtDNA expresses the enzymes required for ATP synthesis. Between 20 to 25% of LS are due to mutations in mtDNA. 10 to 20 percentage of mutations occurs in mtATP6 gene that codes for protein in the complex V. ATP synthase is required by the electron transport chain to produce ATP which is primary source of energy, and the lack of energy is the primary reason for psycho-motor dysfunction in LS.

The most common mtATP6 mutation found with LS is point mutation at nucleotide 8993 that change a thiamine to guanine (Fruhman et al, 20111). Alongside of this other kinds of gene alterations or mutations in LS destabilize the protein complexes and keep down the energy production in the affected cells.

In a case of LS, mutations in genes mtND2, mtND3, mtND5 and mtND6 and several other genes which contribute for complex I were identified. Mitochondrial DNA is passed down matrilineal in parentis know as maternal inheritance where a mother can transfer the mitochondrial genes to either sex children but where fathers cannot transfer mitochondrial genes.

LS caused by Nuclear DNA mutations were generally inherited by autosomal recessive pattern (Wong, 2012). 75 to 80 % of LS caused by the mutations in Nuclear DNA. Mutations affecting the assembly or functions of complex IV involved in the oxidative phosphorylation is the main cause of LS. Mutations in a SURF1 gene is the most common cause of sub type of LS. The protein that has SURF1 codes for, terminated early therefore it cannot produce its function. That leads to deficiency of COX protein and reduction of energy production.

Another enzyme Pyruvate Dehydrogenase plays a role in glycolysis gets affected by mutations in genes (Koga et al, 2012). The following table lists the genes linked with LS.

List of nuclear genes that are involved in Oxidative phosphorylation and causes the LS

1. Complex I:  NDUFA1, NDUFA2, NDUFA9, NDUFA10, NDUFA12, NDUFS1, NDUFS2, NDUFS3, NDUFB8, NDUFS4, NDUFS7, NDUFS8, NDUFV1, NDUFV2

2. Complex II: SDHA

3. Complex III: UQCRQ

4. Complex IV: NDUFA4, COX8A

5. Complex I Assembly factors deficiency: NDUFAF2, NDUFAF4, NDUFAF5, NDUFAF6, C17ORF89, FOXRED1

6. Complex II Assembly factors deficiency: SDHAF1

7. Complex III Assembly factors deficiency: BCS1L, TTC19

8. Complex IV Assembly factors deficiency: SURF1, COX10, COX15, SCO2

9. Pyruvate Dehydrogenase Complex: PDHA1, PDHX, PDHB, DLAT, DLD

10. Biotinidase Deficiency: BTD

11. Thiamine Deficiency: TPK1, SLC19A3

12. Lipoic acid: LIPT1, LIAS, BOLA3

13. Amino acid: HIBCH, ECHS1

14. Coenzyme Q10 Deficiency: PDSS2, COQ9

One study demonstrated that, NDUFAF4 missense variant c.194 T>C (p.Leu65Pro) exhibiting an early onset of neuronal development related regression, hypotonia and irritability (Schubert & Vilarinho, 2020). Biochemical results suggest that elevated lactic acid in blood and cerebral spinal fluid. One study confirmed that late-onset LS presented with progressive ataxia and dysarthria alongside a lactate surge was seen in putamen and right caudate topography. One study discussed a case where a 7-month-old boy presenting with torticollis, MRI of brain with LS  was evolved to crisis of metabolism and gradual progression of lesions in the regions of basal ganglia. MTND1 m.3697G > A previously reported in MELAS is suggested to be implied as well.

Pathophysiology:

Some of the characteristic LS symptoms are caused by bilateral focal lesions in the brain stem, basal ganglia, cerebellum and other sections of the brain. There would be deferent forms of lesions in areas of demyelination, spongiosis, cell death and angiogenesis. Brainstem is involved in maintaining basic life functions such as breathing, swallowing and circulation. Cerebellem and basal ganglia regulate body movement and balance. Any impairment in these areas leads to major symptoms of LS. Lactic acidosis is due to defects in OXPHOS which implies to deficiency of pyruvate dehydrogenase.

Clinical findings:

The symptoms like nystagmus, dystonia, and defects with ANS (autonomic nervous system) suggest the impairment of the basal ganglia and brainstem strongly caused by LS. Neurologically caused deafness is an indication of neurological damage in LS. Increased levels of alanine in blood. Organic acids in urine indicate the dysfunction in metabolic pathway.

Treatment: 

LS is a rarest of the genetic disorders, and certainly there has not been a quality and effective treatment available. A more fat, less sugar containing diet would be helpful LS. In-case of pyruvate dehydrogenase deficiency, vitamin B1 and thiamine will be supplied. The symptoms of Lactic acidosis will be treated by sodium bicarbonate or sodium citrated.

In 2016, scientists performed maternal spindle transfer of mitochondria donation technique in a pregnant lady at Mexico who is likely to deliver a baby with LS. She was blessed with baby boy on April 2016 without LS. However, it is not clear about the safety and reliability of technique. 

Still there is a lot of research to be done to identify the potential drug candidates and other effective gene therapies and there is need to educate the society on to develop awareness that lead to genetic disorders on consanguineous marriages.

References

Annetta, L. A., Cheng, M. T., & Holmes, S. (2010). Assessing twenty‐first century skills through a teacher created video game for high school biology students. Research in Science & Technological Education, 28(2), 101-114.

Baertling, F., Rodenburg, R. J., Schaper, J., Smeitink, J. A., Koopman, W. J., Mayatepek, E., ... & Distelmaier, F. (2014). A guide to diagnosis and treatment of Leigh syndrome. J Neurol Neurosurg Psychiatry, 85(3), 257-265.

Dumollard, R., Duchen, M., & Carroll, J. (2007). The role of mitochondrial function in the oocyte and embryo. Current topics in developmental biology, 77, 21-49.

Finsterer, J. (2008). Leigh and Leigh-like syndrome in children and adults. Pediatric neurology, 39(4), 223-235.

Fruhman, G., Landsverk, M. L., Lotze, T. E., Hunter, J. V., Wangler, M. F., Adesina, A. M., ... & Scaglia, F. (2011). Atypical presentation of Leigh syndrome associated with a Leber hereditary optic neuropathy primary mitochondrial DNA mutation. Molecular genetics and metabolism, 103(2), 153-160.

Koga, Y., Povalko, N., Katayama, K., Kakimoto, N., Matsuishi, T., Naito, E., & Tanaka, M. (2012). Beneficial effect of pyruvate therapy on Leigh syndrome due to a novel mutation in PDH E1α gene. Brain and Development, 34(2), 87-91.

Lake, N. J., Compton, A. G., Rahman, S., & Thorburn, D. R. (2016). Leigh syndrome: one disorder, more than 75 monogenic causes. Annals of neurology, 79(2), 190-203.

Martinelli, D., Catteruccia, M., Piemonte, F., Pastore, A., Tozzi, G., Dionisi-Vici, C., ... & Hinman, A. (2012). EPI-743 reverses the progression of the pediatric mitochondrial disease—genetically defined Leigh Syndrome. Molecular genetics and metabolism, 107(3), 383-388.

Quintana, A., Kruse, S. E., Kapur, R. P., Sanz, E., & Palmiter, R. D. (2010). Complex I deficiency due to loss of Ndufs4 in the brain results in progressive encephalopathy resembling Leigh syndrome. Proceedings of the National Academy of Sciences, 107(24), 10996-11001.

Rahman, S., Blok, R. B., Dahl, H. H., Danks, D. M., Kirby, D. M., Chow, C. W., ... & Thorburn, D. R. (1996). Leigh syndrome: clinical features and biochemical and DNA abnormalities. Annals of Neurology: Official Journal of the American Neurological Association and the Child Neurology Society, 39(3), 343-351.