Highly targeted approach supports ALS gene therapy development
Technology targets diseased nerve cells, leaves healthy cells alone
Researchers have developed a type of gene therapy that can rescue the function of the TDP-43 protein in diseased nerve cells in amyotrophic lateral sclerosis (ALS) and other neurodegenerative diseases while leaving healthy cells untouched.
The approach uses problematic sequences called cryptic exons that are only present in cells where TDP-43 is not executing its functions properly. When these sequences are not removed from genetic messages as they should be, the therapy becomes active and produces a therapeutic protein that helps to restore the lost functions.
As healthy cells can remove the sequences, no therapeutic protein is produced there, which can substantially reduce off-target effects from the gene therapy and improve its safety profile.
Details of the new development were published in Science, in the study, “Creation of de novo cryptic splicing for ALS and FTD precision medicine.”
“We hope this new technology will enable much bolder therapeutic approaches for [ALS],” lead author Pietro Fratta, MD, PhD, head of the Molecular Neurodegeneration Laboratory at the Francis Crick Institute and a University College London (UCL) professor, said in a university news story.
Avoiding toxicity
“Many potential therapies also alter important cellular processes and this may cause toxicity,” said Fratta. “Therefore, limiting their action to diseased cells, while leaving healthy cells untouched, will increase the safety of gene therapies and allow researchers to pursue many more treatment options.”
ALS, also known as motor neuron disease, is marked by the progressive damage and degeneration of motor neurons, the specialized cells that control voluntary movement.
While the mechanisms that lead to disease are not fully understood, toxic clumps of the TDP-43 protein are evident in about 97% of ALS cases and are known to contribute to nerve cell damage. Such clumps are also observed in many people with the related disorder frontotemporal dementia (FTD) and in those with Alzheimer’s disease.
TDP-43 is a protein that’s normally found in the nucleus, where cells’ genetic material is housed. There, it helps to control how certain genes are read, ensuring that the protein-making machinery receives the correct genetic sequence to create a functioning protein.
When TDP-43 forms clumps, it fails to reach the nucleus to exert its function. As a result, small extra sequences called cryptic exons are erroneously incorporated into the genetic templates that are used to make proteins, ultimately affecting protein production and leading to disease.
“TDP-43 controls many aspects of cellular health, and its dysfunction is a key driver of disease,” said U.S. collaborator and study author Claire Le Pichon, PhD. Thus, “gene therapy holds promise for treating neurodegenerative diseases like ALS and FTD, which are relatively common but for which there are few treatments,” said Le Pichon, an investigator at the Eunice Kennedy Shriver National Institute of Child Health and Human Development.
Targeting cryptic exons
Numerous studies have tried to apply gene therapy methods to block the inclusion of cryptic exons and reduce the cellular toxicity caused by TDP-43. A major barrier is that only a small fraction of motor neurons are diseased at any given moment. Increasing the production of TDP-43 may ameliorate disease in affected cells, but such treatments may also interfere with the vast majority of otherwise healthy cells and worsen patient outcomes.
“While neurodegenerative diseases have devastating effects, we can estimate that less than 0.00001% of cells in a patient’s body are actually diseased,” said Oscar Wilkins, the study’s first author and a research fellow at UCL. “The challenge is finding a way to specifically target treatments to this minuscule fraction of diseased cells, while avoiding unnecessary treatment of the 99.99999% of cells which are healthy.”
Now, with an understanding of TDP-43’s function and AI prediction tools, the team at Francis Crick and UCL designed a gene therapy approach that’s only active in cells containing cryptic exons.
Delivered to cells and mice using a harmless adeno-associated virus (AAV), which is commonly used in other gene therapies, the system, dubbed TDP-REG, successfully targeted and reversed the impact of TDP-43 disease in cells with cryptic exons but not healthy cells.
Then, using stem cell-derived neurons that form synapses, the gaps between nerve cells where nerves communicate, TDP-REG removed cryptic exons from the UNC13A gene. This gene produces a protein that’s essential for regulating neurotransmitter release at synapses, a key process for communicating between nerve cells, and its altered function due to the presence of mutations or cryptic exons has been linked with an increased risk of ALS and FTD.
The therapeutic approach also effectively removed cryptic exons from other genes known to be processed by TDP-43, while no production of TDP-REG was observed in healthy cells.
“There’s huge potential in precision medicine approaches that only target the cells that need treatment,” Wilkins said. “Whether it’s cancer, heart disease, or motor neuron disease, the key challenge is finding something unique about the diseased cells that we can use for our own purposes — in this case, we chose the TDP-43 protein, as it becomes dysfunctional in so many different neurodegenerative disorders.”
Max Chien, a PhD student at UCL who helped with the project, added: “As well as potentially delivering gene therapies, our approach can also be used to detect diseased cells. This could help scientists test whether their treatments perform correctly, before entering into a clinical trial.”
About the Author
