Synthetic molecules targeting the hallmark gene mutation in Friedreich’s ataxia (FA) were able to restore frataxin protein levels to normal in cells taken from patients, a study has found.
Specifically, the FXN gene in FA patients contains an expansion of a GAA trinucleotide repeat. Nucleotides, which are the building blocks of genes, include adenine (A), cytosine (C), guanine (G), and thymine (T).
Previous studies have suggested that the expanded GAA repeat leads to the formation of complexes that damage FXN gene expression, or protein production, and lower protein levels.
Scientists hypothesize that targeting the GAA repeat expansion may prevent the formation of the complexes and boost production of frataxin.
In prior work using patient-derived cells, they found that two types of synthetic molecules, known as anti-GAA duplex RNAs and antisense oligonucleotides (ASOs), increased frataxin protein levels.
In this study, “Activating frataxin expression by single-stranded siRNAs targeting the GAA repeat expansion,” which appeared in the journal Bioorganic & Medicinal Chemistry Letters, they explored two additional types of compounds for their potential to boost protein production.
One is ASOs with chemical alterations called butane linkers and 2-F-ANA to enable a more flexible conformation, while the other is single-stranded silencing RNAs (ss-siRNAs), which are RNA molecules modified to inhibit, or silence, gene expression, These RNAs have shown positive results in mouse and human disease cell models.
Results showed that the modified ASOs had low potency, suggesting that a more flexible conformation impairs activity.
But ss-siRNAs showed high potency. These molecules were then tested in cells derived from FA patients, as well as in cells that were either nonmutant or had one mutant and one normal FXN gene copy (heterozygous), to explore whether ss-siRNAs are able to increase gene expression.
Data revealed that messenger RNA — which contains the information to make frataxin — and protein levels were lowest in patient-derived cells, two to three times higher in nonmutant cells, and intermediate in heterozygous cells.
Investigators then found that ss-siRNAs were able to boost FXN expression to normal levels in patient-derived cells, but did not did not increase mRNA and frataxin protein levels in non-FA cell lines.
These findings indicate that ss-siRNAs do not induce production of frataxin beyond normal levels in healthy cells, but they do restore protein levels in FA patients’ cells. Because previous studies showed that making frataxin in excess in cell models may cause oxidative stress and cell death, the careful regulation of gene expression with ss-siRNAs may be important, the researchers noted.
According to the study, “ss-siRNAs provide an additional option for developing nucleic acid therapeutics to treat [FA].”