Genomic and RNA base editing are potential therapeutic approaches for treating human disease. Genomic editing has been used to successfully repair a gene responsible for hearing loss in mice. In a recent study, scientists have successfully edited RNA in living animals so that the repaired RNA can correct mutations in proteins, thus improving the neurasthenia caused by Rett syndrome. The results were published in the recent journal Cell Reports.
Rett syndrome (RTT) is a severe neurological disorder that is caused by mutations in the methyl-CpG-binding protein 2 (MECP2) gene. Classical RTT affects around 1 in 10,000 live female births and is characterized by an about 6‑month period of overtly normal development. This is followed by the onset of symptoms, which include deceleration of head growth, breathing disturbances, gait abnormalities, loss of speech and the replacement of purposeful hand movements with repetitive stereotypes. The disease is rarely observed in males, as mutations in MECP2 are generally paternally rather than maternally derived, and inactivation of the sole X‑linked copy of MECP2 results in severe neonatal encephalopathy and early lethality.
Mandel and co-authors, led by postdoctoral fellow John Sinnamon, Ph.D., have been working on a method to repair the mutated MeCP2 protein at the RNA or RNA level, which acts as a messenger for the synthesis of proteins controlled by DNA. They focused on RNA base editing because it does not have the same sequence constraints as DNA base editing. Earlier work has proved the potential of site-directed RNA editing for treating human disease. Recently, a demonstration of successful in vivo RNA editing was published using G>A mutant mouse models of Duchenne muscular dystrophy and ornithine in vivo transcarbamylase deficiency. Their study suggests that in vivo programmable RNA editing can also efficiently edit target RNA in heterogeneous nervous tissue, leading to functional repair of a patient mutation in mice representing a human neurological disease.
Figure 1. Editing repairs the Mecp2 RNA mutation and protein function across neuronal types
This new study targets and repairs MeCP2 protein in a variety of cell types. "We repaired MeCP2 protein in three different neuronal populations," Mandel said. "So, assuming that we can provide a wide range of editing components, it's possible that it would work throughout the brain."
In this study, using a Rett syndrome mouse model, they determine whether mutant Mecp2 RNA, in distinct neuronal subpopulations in the postnatal mouse brain, is accessible to programmable RNA repair and whether MeCP2 protein function is also restored. They also determine the on- and off-target editing landscape in one of these populations of neurons using whole-transcriptome analysis.
Their results showed a uniformity of 50% editing and a comparable association of MeCP2 protein with heterochromatin across several hippocampal neuronal subtypes. This result indicates that, using peripheral injections, neuronal populations across the brain should share a similar repair rate. However, how much repaired MeCP2 per cell is necessary and how many neuronal and glial cells need to be repaired to reverse Rett syndrome phenotypes in mice is unclear. Previous reports suggest that the mouse and human nervous systems are very sensitive to the levels of MeCP2 expression, and even a 2-fold increase in MeCP2 levels leads to a neurological phenotype. Although it is unlikely that 50% repair per cell will result in a wild-type mouse, this level of repair could be reasonably expected to result in significant improvement in Rett syndrome-like phenotypes in treated mice, and, more importantly, programmable editing will never cause overexpression of MeCP2.
While the Oregon Health and Science University research shows that RNA repair holds promise as a proof of concept, Mandel emphasized that more research needs to be done to test whether it reverses Rett-like behaviors in mice, and to improve the efficiency and specificity of the repair.