Conference Agenda

Mitochondrial diseases are a family of genetically inherited conditions impairing the oxidative phosphorylation system. They are characterized by extreme clinical and genetic heterogeneity and poorly understood tissues specificity, with some of them effecting multiple organs and somea single organ/tissue. No cure is currently available for most mitochondrial diseases. AAV-based gene therapy is one of the most promising approaches for several genetic determined conditions. However, its application to mitochondrial diseases is so far limited to a very few conditions, one of which is Leber’s hereditary optic neuropathy. We tested AAV-based gene therapy in mouse models of mitochondrial encephalopathies and myopathies with encouraging results.

Leigh disease is a genetically heterogeneous condition characterized by defective mitochondrial bioenergetics and is the most common oxidative phosphorylation disease in infancy. A knockout mouse for complex I subunit Ndufs4 recapitulates the main features of Leigh syndrome. We exploited double-stranded self-complementary AAV9 (scAAV9) vectors, to deliver the wild-type form of human NDUFS4 to Ndufs4-/- mice by double intra-vascular and intra-cerebro-ventricular injections at post-natal Day 1. This strategy prolonged healthy lifespan up to 9 months of age. Robust expression of hNDUFS4 was detected in different cerebral areas preserving normal morphology and restoring Complex I activity and assembly. Future work is warranted to explore translatability of scAAV9-NDUFS4 in the prodromal phase of the disease in mice and eventually humans.

We also tested a new myotropic serotype in a muscle-specific knockout mouse for Cox15, a gene encoding the last enzyme of the biosynthesis of heme A. We observed substantial recovery of the motor performance in AAV-treated vs untreated Cox15 KO mice, and marked improvement of the myopathy.

The general relevance and implications for mitochondrial diseases and optic neuropathies will be discussed.

Congenital sideroblastic anemia (CSA) is a heterogeneous group of inherited disorders of erythropoiesis characterized by iron overload in the mitochondria of developing erythroblasts. Mutations in the mitochondrial glycine carrier, encoded by SLC25A38, cause the most common recessive form of CSA. Unfortunately, our understanding of its pathogenic mechanisms is limited, and pharmacological studies have been hindered by the lack of suitable biological models.

To overcome these limitations, we generated two different cell models: K562 erythroleukemia cells with reduced SLC25A38 expression, and a lymphoblastoid cell line derived from a CSA patient with a nonsense mutation in the SLC25A38 gene. Both cell lines recapitulated the main features associated with this rare anemia, including reduced heme content and respiratory defect. However, other features were exclusively observed in the K562 mutant cells, such as an increase in mitochondrial iron and high ROS levels indicating that altered iron homeostasis and oxidative stress are secondary to heme reduction and occur erratically in different cells.

Furthermore, we found that externally added pyridoxal 5'-phosphate, but not the B6 vitamers pyridoxine, pyridoxal, and pyridoxamine, rescues the altered parameters of both CSA models via a mechanism that is independent of the intracellular PLP content.

Beside shedding light on the interrelation between heme biosynthesis and mitochondrial iron homeostasis, our results pave the way to new potential therapeutic strategies.

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