Antioxidants Increase Frataxin Levels in Friedreich’s Ataxia Cells, Study Reports

Specific antioxidant compounds and derivatives were able to boost levels of the protein frataxin in cells from Friedreich’s ataxia (FA) patients, according to researchers from Arizona State University in Tempe.

Their study, “Phenothiazine antioxidants increase mitochondrial biogenesis and frataxin levels in Friedreich’s ataxia cells,” appeared in the journal MedChemComm.

FA is caused by a mutation in the FXN gene, which leads to reduced production of frataxin, a key protein for proper functioning of mitochondria — energy-producing powerhouses of cells. A deficiency of frataxin is associated with iron accumulation in mitochondria, increased sensitivity to oxidative stress, and impaired energy production.

Phenothiazines are a class of compounds characterized by three ring-like structures of carbon and hydrogen, and with sulfur and nitrogen atoms. This overall structure has been used in the development of medications with a broad range of biological properties.

Prior studies revealed that methylene blue (MB) — a phenothiazine derivative — increased frataxin levels in cells from FA patients.

Both MB and its analog compound methylene violet (MV) are able to alter energy production in mitochondria under disease conditions, and could potentially suppress the associated oxidative stress, the scientists said. Earlier work showed that MV derivatives also protected cells against harmful events.

The researchers hypothesized that modifying methylene violet could generate compounds with optimal properties. They compared previously described methylene blue analogs to newly synthesized MV-derived products.

In lymphoblasts — immature cells that can develop into mature white blood cells — from Friedreich’s ataxia patients, MB analogs with side chains (attached to the core part of the molecule) comprising 5-13 carbon atoms (named 3a-5a) showed increased cell toxicity.

In turn, MV and one of its analogs also demonstrated cell toxicity after 48 hours, but less pronounced than MB and related compounds.

The data also showed that in FA cells with oxidative stress, methylene blue and analogs with shorter carbon side chains were more protective at a lower rather than higher concentration. Compounds with longer chains showed improved cell protection. MV derivatives’ cell protective effects increased with both higher concentrations and longer chains.

Subsequent experiments explored the ability to improve production of adenosine triphosphate (ATP), the energy currency of the cell.

Increased ATP generation without producing cellular toxicity is necessary for therapeutic use of any coenzyme Q10-related antioxidant, the investigators said.

These compounds should be able to bind to mitochondrial complexes — part of the energy production process — but not tightly, so as not to inhibit them, the authors noted.

MB and derivatives with shorter carbon side chains, as well as MV and its 3b (5-carbon side chain) derivative, strongly reduced ATP levels at higher concentrations. The MB analog 6a and the MV derivatives 4b and 6b (11-16 carbon side chains) boosted ATP levels at lower concentrations, but suppressed it at the highest concentration.

The investigators then found that both MB and MV increased frataxin levels and the cellular production of mitochondria in cells derived from Friedreich’s ataxia patients.

Their derivatives showed mostly similar effects, but with some exceptions. Methylene violet analog 4b was ineffective at both doses tested, which, according to the authors, “underscores the ability of small structural differences to support real changes in potential therapeutic benefit.”

Also, a five-day exposure revealed that the increased levels of frataxin were associated with improved generation of iron-sulfur clusters — proteins that participate in mitochondrial energy production.

Although future studies are needed to understand how MB/MV and their analogs increase frataxin levels, the scientists hypothesized that it may involve altered FXN gene levels (protein production) or changes in frataxin metabolism.