New Gene Therapy Approach Able to Repair Mutations Causing SMA, Other Inherited Diseases, Mouse Study Suggests

A new gene therapy approach using RNA molecules called transfer RNAs (tRNAs) was able to repair a subset of mutations causing spinal muscular atrophy (SMA) and other inherited diseases in living muscle tissue of mice, a study shows.

These genetic alterations, called nonsense mutations, alter the DNA sequence and introduce so-called stop codons — triplets of nucleotides, which are the building blocks for DNA — that prematurely stop gene expression and impair protein production. Besides SMA, these mutations also underlie diseases such as Duchenne muscular dystrophy, cystic fibrosis, and polycystic kidney disease.

The study, “Engineered transfer RNAs for suppression of premature termination codons,” was published in the journal Nature Communications.

Several lines of research have attempted to find compounds that are able to repair nonsense mutations. However, previous research has shown that certain small molecules can generate a different type of mutation that is able to disrupt protein function, but they can be toxic to the ears and kidneys. In addition, using the CRISPR/Cas9 gene editing technique — a potential treatment for diseases caused by nonsense mutations — has presented other challenges, such as off-target effects.

Aiming to find a suitable approach with the ability to repair nonsense mutations precisely, a team from the University of Iowa Carver College of Medicine, The Wistar Institute, the Cystic Fibrosis Foundation Therapeutics Lab and Integrated DNA Technologies investigated the potential of using tRNAs.

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All genetic information contained within genes (DNA) is ultimately translated into proteins. However, the process is complex, with several steps: DNA is transformed into messenger RNA (mRNA), then a process called translation begins, which results in the production of proteins.

tRNA is a type of RNA molecule that helps decode a messenger mRNA sequence into a protein. Through specific sequences called anticodons, tRNA molecules match up with the corresponding mRNA and deliver the correct amino acid to build a protein in a cellular structure called ribosome.

Researchers engineered tRNAs to recognize and suppress three different stop codons — triplets of nucleotides that prematurely stop gene expression and impair protein production. (Gene expression is the process by which information in a gene is synthesized to create a working product, such as a protein).

To be effective, however, the edited tRNAs should still be recognized by the cells’ machinery that’s involved in translation.

The results showed that the engineered tRNA molecules were able to repair the most common disease-causing nonsense mutations with any amino acid. Also, the cell-based approach identified multiple engineered tRNAs for each amino acid and type of stop codon.

The researchers then observed that the tRNAs, when encoded and formulated for efficient delivery, were produced at high levels and effectively corrected nonsense mutations in mammalian cells, including in mouse muscle tissue. The repair was found in diverse disease-related genes such as CFTR, where 1,000 mutations have been linked with cystic fibrosis.

Also, the results revealed that the tRNA activity was sustained for weeks and did not affect stop codons that signal the normal end of the protein sequence.

“In total, the data support the possibility that such engineered tRNA satisfy the broad requirement for coverage of disease-causing (stop codons) and thus represent a promising new class of RNA therapeutic agent,” the researchers wrote.

“What I like about this study is that a number of different labs with different expertise all verified our engineered tRNA technology in a variety of contexts,” Christopher Ahern, PhD, the study’s senior author, said in a press release. “That suggests the approach is robust.”

Ahern added that the high-throughput technology “turns ‘stops’ into ‘gos’ and hopefully one day may be used to correct defective genetic sequences in people.”

However, he cautioned that more information is still required, such as the ideal delivery strategies, and that there are technical obstacles to overcome before this gene therapy can be used in human therapies.

“For many diseases caused by nonsense mutations, even correcting a small percent of the mutated protein could be enough to be therapeutic to the patient,” Ahern said. “If this were to work as a human therapy, we would have a way to target every known stop codon disease.”