Novel Mutations of mtDNA m.14568G>A/m.14568C>T in MT-ND6 and m.7299A>G in MT-CO1: Evidence of Pathogenicity in Leber Hereditary Optic Neuropathy.

Leber hereditary optic neuropathy (LHON) is a mitochondrial genetic disease characterized by bilateral, painless, subacute visual failure that develops during young adult life (1). It is a mitochondrial genetic disease with a maternal pattern of inheritance. Identifying the mutations is important not only in respect to the diagnosis but also as different LHON mutations result in variations of severity and recovery rate in the affected individuals. Ninety-five percent of LHON cases are associated with 1 of 3 primary mutations: m.3460G>A, m.11778 G>A, and m.14484T>C with defect in the mtDNA (mitochondrial DNA) respiratory chain complex I subunit genes MT-ND1, MT-ND4, and MT-ND6 (mitochondrially encoded NADH:ubiquinone oxidoreductase core subunit 1,4, and 6). However, there are many reports about new primary and potential mtDNA mutations associated with LHON from different parts of the world. A genotype–phenotype correlation depending on the mutated gene and the type of pathogenic variant has been discussed in different articles (2). In this report, we discuss the case of 2 young Lebanese patients with LHON who were found to have 2 novel mutations: m.14568G>A/C>T and m.7299A>G (Table 1). TABLE 1. - Ophthalmologic examination of Patient #1 and Patient #2 at presentation and on follow-up Patient #1, 24-Year-Old At Presentation (1 Month After Onset of Symptoms) 10 Months Later Visual acuity OD Counting fingers at 2 feet - pinhole 20/100 20/30 OS 20/40 - pinhole 20/30 20/40 Pupils Equal, reactive, no relative afferent pupillary defect (RAPD) Equal, reactive, no RAPD Intraocular pressure (IOP) OD 13 15 OS 14 17 Color vision OD 4/14 8/14 OS 11/14 9/14 Fundus examination Full and hyperemic discs Less full and hyperemic discs Visual field Central and centrocecal scotomas in OD > OS Improved Extraocular movements/slit lamp examination Normal Normal Patient #2, 16-Year-Old At Presentation (8 Months After Onset of Symptoms) 3 Months Later Visual acuity OD Counting fingers 20/400 OS 20/400 20/150 Pupils Equal, reactive, no RAPD Equal, reactive, no RAPD IOP OD 14 14 OS 15 15 Color vision OD 4/14 5/14 OS 5/14 6/14 Fundus examination Pale optic nerve head with normal macula and vessels OU Pale optic nerve head with normal macula and vessels OU Visual field Cecocentral scotomas OU Cecocentral scotomas OU Extraocular movements/slit lamp examination Normal Normal CASE PRESENTATION At presentation, a workup was performed looking for infectious, inflammatory, demyelinating, and toxic causes, which was nonrevealing in both patients. Based on history and physical examination, these 2 patients were thought to have LHON, which was confirmed with genetic testing. Genetic testing detected a homoplasmic variant in the MT-ND6 gene m.14568G>A (Patient #1) and m.14568C>T (Patient #2) and a homoplasmic variant in the MT-CO1 gene m.7299A>G in both patients. In addition, the mother of Patient #1 had the same mitochondrial mutation. Patient #1 was compliant with treatment and lifestyle modification. He was started on Idebenone. He was advised to avoid smoking and alcohol. He showed significant improvement on subsequent follow-up, last being 10 months after his initial presentation. Patient #2 unfortunately refused treatment and lifestyle changes and was lost to follow up. DISCUSSION Here, we report on 2 cases with LHON pathogenic variants. The MT-ND6 gene encodes 1 of the 7 mitochondrial subunits of the respiratory Complex I. The 14568 mutation in ND6 gene lies in close vicinity of other LHON-related mutations within the evolutionarily most conserved region of the ND6 gene forming a hydrophobic pocket in the gene, making it a hotspot for LHON (3,4). Mutations involving the specific position 14568 in the mitochondrial ND6 gene are proven to be as causative in LHON (4). It was previously described in the literature in 2 cases only as of pathogenic significance in LHON by Besch et al (3) in 1999. The second mutation found is m.7299A>G in MT-CO1. It involves the cytochrome c oxidase subunit I gene, which is 1 of the 3 mitochondrial DNA encoded subunits of respiratory Complex IV. Mutations involving the MT-CO1 gene are proven to be involved in LHON, and this genotype–phenotype correlation is previously described in the literature (5). However, the specific variant found in our 2 patients involving the position 7,299 has not been previously described and there are no data in the literature concerning this mutation and its pathogenicity in LHON (5). It is important to note that secondary mutations are previously described in LHON, specifically in the ND6 gene. Altogether, 7 mutations in ND6 gene are described in the same hydrophobic transmembrane helix of the ND6 gene: 14459, 14482, 14484, 14495, 14498, 14568, and 14596, making it a mutational hotspot for the disease (4). This adds further evidence that the 14568 mutation is of pathogenic significance and that this region may be useful to sequence in patients where no known LHON mutations are found. Also, secondary mutations were previously suggested and suspected to increase the penetrance of a primary LHON mutation (4). This is another strong argument in favor of the significance of the 2 secondary mutations found in this report. Whether the phenotype of LHON in these patients is due to the synergistic presence of the 2 mutations involving both MT-CO1 and MT-ND6 genes or only due to the presence of the mutation involving the MT-ND6 gene is yet to be determined. Even if the m.7299A>G in MT-CO-1 is not the primary cause of LHON in this patient, it still could be an exacerbating risk factor. To note that vision of Patient #1 improved with time. The main difference between the 2 patients was the time of presentation after initial vision loss and lifestyle changes including treatment with Idebenone, which Patient #1 followed and was the one with significantly better outcomes. We realize that this could be due to chance alone; however, this observation raises the possibility of different mutations having different phenotypic outcomes in LHON and stresses the importance of avoiding mitochondrial stressors such as smoking and the possible benefit of starting treatment with Idebenone as early as possible, as proven in several studies such as the Rescue of Hereditary Optic Disease Outpatient Study (6). In addition to that, we raise the possibility of different mutations existing in different parts of the world and affecting the phenotypic outcome of the patients' disease. Both of our patients are Lebanese and of Lebanese descent. Therefore, a better knowledge of the mtDNA mutations related to LHON and their interaction is crucial and will further improve the sensitivity and reliability of mtDNA based diagnostics (2). Also, establishing geographic distribution of the mutations might help diagnose and understand the prognosis of LHON patients and will lead to better counseling of these patients. However, these findings have some limitations. In fact, although these results can help confirm the diversity of LHON clinical phenotypes and increase the understanding of the clinical features of LHON patients with rare mtDNA mutations, it does not provide data to further explain potential mechanisms of the mutations. In addition, there are no previous data concerning our patients' population, which makes it harder to prove the pathogenicity of the 2 uncommon secondary mutations found. CONCLUSION We suggest that more advanced genetic testing of patients must be performed to establish more accurate references for mtDNA based diagnosis and prognosis of LHON. We also suggest that different populations might have different genetic and phenotypic presentation of LHON. Further studies can help establish geographic distribution of the mutations as well as aid in understanding the prognosis and treatment for those patients.