CHARM: A Novel Tool for Silencing Pathogenic Proteins in the Brain
Post by Meredith McCarty
The takeaway
Pathogenic protein accumulation in the brain can lead to rapid-onset dementia and death, but no effective treatment options currently exist. In this study, researchers present a novel gene editing tool called CHARM which can reduce >80% of pathogenetic protein in the mouse brain, with exciting promise for translation to human studies.
What's the science?
There are currently no effective treatments for prion disease and other neurodegenerative diseases caused by the accumulation of toxic protein aggregates, and when left untreated, these diseases are fatal. Previous work has shown that either the deletion of the gene that encodes this prion protein (PrP), or the reduction of PrP in the brain reduces symptoms in mouse models of prion disease. As such, the development of technology to reduce prion levels effectively and without side effects has the potential to treat prion disease and other neurodegenerative diseases in humans. While there has been much development in gene editing technologies based on CRISPR, these methods can have adverse consequences and are not easily translatable to human studies.
This week in Science, Neumann and colleagues develop a novel gene editing tool called CHARM, and present evidence of its utility as a compact and effective tool for eliminating pathogenetic protein accumulation in the brain.
How did they do it?
The authors modified and built upon existing epigenetic editing methods which enable the altering of gene expression to avoid cytotoxicity and other side effects. The novel strategy they implement is to fuse the C-terminal of Dnmt3l (an enzyme involved in the regulation of certain cellular pathways) with an H3 tail (fusion with the epigenetic editor), allowing for methylation-mediated silencing of specific DNA sequences. This effector is delivered via an adeno-associated virus (AAV), a tool that can deliver gene therapy to the central nervous system.
They call this new effector CHARM (Coupled Histone tail for Autoinhibition Release of Methyltransferase) and perform multiple tests in mouse models to confirm the effectiveness and safety of this method for reducing prion expression.
They not only expect this design to allow for selective silencing of prion expression using a much smaller construct than contemporary tools, but importantly the turning off of CHARM activity once prion expression has been reduced. This allows for less runaway effects of CHARM activity after the intended pathogenetic protein silencing has been achieved.
What did they find?
After confirming that their mouse models are a viable in vivo option to study prion repression methods, they run several tests in which they modify elements of the CHARM effector’s structure to confirm their design is the most effective design for selective gene silencing.
The results of these tests manipulating different parameters reveal that the CHARM effector is both flexible and specific. This means that CHARM is compatible with different DNA binding domains that could be used in other applications. They also found that CHARM only acts in the intended way on its encoded target without leading to other unintended effects.
They next performed in vivo testing through AAV delivery in the mouse brain and found no adverse effects, a 70-90% decrease in Prnp transcripts, and a 60-80% reduction in PrP protein levels. Finally, they were able to program CHARM for self-silencing, meaning that after the target gene expression is reduced, the CHARM effector is designed to then silence its own promotor, leading to a halt of future activity.
What's the impact?
This study puts forth a novel gene editing tool CHARM, which can permanently silence gene expression with high specificity, low toxicity, and an easy mechanism of delivery across the brain. The authors suggest that the design of CHARM will allow its use on a broad range of genes that are amenable to DNA methylation-mediated silencing.
This technology has incredible implications, not only for prion disease but for numerous other neurodegenerative diseases that currently have limited treatment options.