Capsid Screening Highlights Gene Therapy’s Potential for Rare Brain Diseases

An advanced screening technique helps researchers improve the precision of gene therapy when targeting hard-to-reach areas of the brain.

Conditions such as lysosomal storage disorders (LSDs) and neurodegenerative diseases have historically been difficult to treat with low doses of gene therapy due to their location in the brain and the nature of their progression. Using an advanced screening technique in preclinical models, researchers from Children's Hospital of Philadelphia identified several adeno-associated virus (AAV) capsid variants that could improve the cost, dosage and precision of gene therapy for patients with these disorders. Their findings were published in Science Translational Medicine and Nature Communications.

"Our research has solved challenges in current gene therapies," said Beverly Davidson, PhD, leader of both studies and Director of the Raymond G. Perelman Center for Cellular and Molecular Therapeutics. "Our aim is to establish the potential to create therapies that require lower doses and directly target cells in affected tissues."

AAV capsids are the vehicles that deliver genetic material to affected sections of the brain during gene therapy treatments. They are made up of three proteins in a repeated structure. Scientists insert a small peptide into one of the proteins, synthesizing a novel sequence of amino acids on the surface with a location for the genetic material to settle in with new instructions to fight disease.

However, AAV gene therapy requires a significant amount of genetic material, so delivery to a specific target within the brain is crucial to maintaining high therapeutic impact while controlling costs. With tens of millions of capsid variants available, Dr. Davidson and her colleagues developed a screening method using preclinical models and high throughput sequencing to identify variants that could target difficult-to-treat tissues more effectively.

"The AAV library is essentially a haystack," Dr. Davidson said. "Each capsid is a strand, and with our technology, we can scale down from the haystack, to a haybale, to that eventual golden strand specific to the tiny area of the brain we want to target."

Optimized Protein Production in LSDs

In the first study, published in Science Translational Medicine, Dr. Davidson and the research team sought an alternative approach to treating LSDs, which are genetic diseases that result from deficiencies in enzymes that break down proteins and other cell products for reuse by the cell. They performed an unbiased in vivo screening technique on millions of peptide-modified AAV capsid variants in animal models, expanding on work that suggests delivering gene therapy to cells that line the ventricles of the brain can serve as bio-reactors to produce therapeutic proteins to improve disease phenotypes.

After screening, researchers identified and isolated the capsid variant AAV Ep+ based on its ability to transduce ventricular lining cells and cerebral neurons in preclinical models. The potency of AAV Ep+ was conserved across multiple animal and disease models, including human brain cells.

In a disease model of ceroid lipofuscinosis type 2 (CLN2), an LSD caused by loss-of-function mutations in tripeptidyl peptidase 1 (TPP1), researchers evaluated the AAV Ep+ capsid's ability to package the human TPP1 transgene and deliver it effectively through cerebrospinal fluid. They noted robust and therapeutically relevant TPP1 protein concentrations after low-dose administration of the therapy.

Current therapeutic options for LSDs include enzyme replacement therapy that requires twice monthly transfusions of the enzymes through a chronically implanted device in the brain that could be prone to infection. The study results suggest AAV Ep+ has the potential to deliver the one-and-done gene therapy for LSDs and other neurodegenerative diseases more efficiently, while the low dosage could eventually help reduce harmful side effects and treatment costs for patients.

Luis Tecedor, PhD, and Yong Hong Chen, PhD, Research Scientists in Dr. Davidson's lab, are co-first authors on this study, with additional research contributions from Latus Bio and The Ohio State University.

Targeted Delivery to Deep Brain Structures

In the second study, published in Nature Communications, Dr. Davidson and her colleagues sought to tackle a long-standing challenge in gene therapy: poor accessibility to deep brain structures. Using a similar screening method of millions of capsid variants in animal models, researchers identified AAV-DB-3 as a potential candidate to deliver genetic material effectively and precisely to key deep brain and cortical structures. Additional research for this study came from The Ohio State University and Latus Bio.

For diseases such as Huntington's disease, characterized by progressive degeneration and transcriptional dysregulation in the basal ganglia, gene therapy's efficacy is limited due to how deep in the brain the affected structures are. Their larger area also makes it difficult to target with a single, direct delivery.

Working with animal models and human neurons derived from induced pluripotent stem cells, researchers delivered genetic material via AAV-DB-3 to the globus pallidum, a structure within the subcortical basal ganglia in the brain that plays a crucial role in motor control and other functions. Compared to other variant capsids, AAV-DB-3 demonstrated the highest potency, revealing a high percentage of transduction of the specific neurons impacted by Huntington's and Parkinson's disease.

AAV-DB-3 also consistently transduced a high percentage of neurons in the deep brain and cortical regions despite dose equivalents that were 10 to 100 times lower than what is currently used in the clinic.

The findings in both studies lay relevant groundwork for potential translational research and clinical trials aimed at improving outcomes for neurodegenerative diseases.

"I'm most excited about this work because I've never seen anything so robustly efficient at transducing a large number of neurons throughout the brain," Dr. Davidson said. "Imagine looking at the night sky in New York City, where you'll see only the moon and a handful of stars. That is the current state of gene therapy. What we've created here is like looking at the night sky over the Sahara Desert."

Both studies were funded by the CHOP Research Institute and Latus Bio. Related intellectual property has been licensed by CHOP to Latus.

Dr. Davidson is a paid consultant to Latus, sits on its Scientific Advisory Board, holds equity in the company, and is a named inventor on the intellectual property licensed by Latus.