Scientists at the University of Massachusetts Medical School have developed a strategy for editing and repairing a particular type of genetic mutation associated with microduplications using CRISPR/Cas9 and a rarely-used DNA repair pathway. Published in Nature, this approach to programmable gene editing overcomes prior issues in gene correction.

"It's like hitting the reset button," said Scot A. Wolfe, professor of molecular, cell and cancer biology. "We don't have to add any corrective genetic material, instead the cell stitches the DNA back together minus the duplication. It's a shortcut for gene correction with potential therapeutic appeal."

Microduplications are changes in chromosomes where small segments of DNA are copied. In certain genes, these duplications can lead to ‘frameshift mutations,’ when the number of added nucleotides is not divisible by three. This alters the translation of gene into protein, resulting in a loss of function. Frameshift mutations occuring from microduplications cause over 140 different diseases, including limb-girdle muscular dystrophy, Hermansky-Pudlak syndrome, and Tay-Sachs.

Dr. Wolfe, a co-investigator of the Nature study, is an expert in CRISPR/Cas9 and other programmable nuclease-based methods of gene editing. Most current methods of gene editing require both generating a break of the DNA strands at the defective gene and the introduction of corrective genetic material. The new sequence is inserted into the break and repaired by an innate DNA repair mechanism found in cells, called the homology-directed repair pathway. Although this has therapeutic potential, it is often inefficient and has other technical challenges.

Researchers hypothesised that if the microhomology-mediated end joining (MMEJ) pathway could be effectively harnessed, instead of the homology-directed repair pathway, it would remove the duplicated sequence and restore the gene's functional sequence. As they predicted, the MMEJ repair mechanism deleted one copy of the microduplication, in effect stitching the DNA back together again, leaving out the mutated genetic material and restoring the gene to enable normal TCAP protein to be produced.

"The simplicity and efficiency of microduplication gene editing of the TCAP gene was a very exciting discovery moment, and presented a unique opportunity to develop a therapy for LGMD 2G, which currently is untreatable, and this has become our immediate goal," said Charles P. Emerson Jr, professor of neurology.