Non-Viral Delivery
Work in Progress
While viral vectors are often used to deliver genetic medicine cargos, they have a number of challenges including inability to re-dose, cargo capacity, potential toxicity at higher doses and a complex and expensive manufacturing process.
Non-viral delivery address many of these challenges including the ability to re-dose and to accommodate larger cargos, and ease of manufacturing and therefore lower cost.
Unlike viruses which naturally infiltrate cells, non viral systems rely on physical or chemical mechanisms.
Work is underway in many labs around the world with the objective of making non-viral delivery to the brain a reality.
Delivery Methods
Some non-viral delivery methods for genetic medicine cargos include:
Ribonucleoprotein complexes (RNP) are the simplest components necessary to achieve genetic editing. RNP consist of an editing protein combined with a guide RNA, and are delivered as a liquid formulation to the CNS.
Lipid-based delivery systems encapsulate and protect genetic material and can be designed to target specific cell types. An example of this approach are the covid vaccines which deliver mRNA encapsulated in lipid nanoparticles.
Exosomes and extracellular vesicles are naturally occurring vesicles released by cells that can encapsulate and transport genetic material between cells. They have gained attention as potential non-viral delivery vehicles due to their biocompatibility and ability to target specific cell types.
Physical methods, like Focused Ultrasound, can enhance delivery to the brain.
A Novel Approach
We’re funding two closely-related proposals that will test a new and emerging concept for delivering genetic cargos to the brain using small RNP (RiboNucleo Protein). Note that RNPs could be delivered repeatedly to increase the number of cells that are edited. Having a delivery route amenable to repeat dosing would be a sea change for developing genetic medicines for Rett syndrome.
Jiangbing Zhou, PhD
Yong-Hui Jiang, PhD
Yale University
Jiangbing Zhou, PhD, is professor of neurosurgery and biomedical engineering. He specializes in developing technology for drug delivery to the brain. His lab focuses on developing translational nanomedicine, gene therapy, and stem cell therapy for treatment of neurological disorders through a unique combination of material science, biology, and engineering.
Yong-Hui Jiang, MD, PhD, is professor of genetics, of pediatrics and of neuroscience and chief of medical genetics. As a physician scientist he is active in both basic research and clinical practice. His expertise is on clinical and biochemical genetics of rare and undiagnosed diseases in children and adult.
Guoping Feng, PhD
Massachusetts Institute of Technology (MIT)
Guoping Feng is currently funded by RSRT and a member of our MECP2 Editing Consortium. His lab has already demonstrated efficacy of its base editing cargo delivered using AAV and will provide an excellent basis for comparison to RNP.
The project utilizes a base editing enzyme with an RNA guide that acts as a GPS, coated with small peptides that protect and direct the enzyme into cells. These particles are 2 to 5-fold smaller than AAV and lipid nanoparticles and therefor can diffuse in the brain more effectively, reaching 50 to 90% neuronal delivery in best cases.
Animals will receive the cargo by intrathecal injection, compatible with human clinical delivery protocols. Studies will use repeat dosing and dose ranging to determine long term correction and durability.
Shawn Liu, PhD
Columbia University
Shawn Liu has received RSRT funding for exploratory work in reactivating the silent MECP2. His work has shown success in cell culture and is staged for pre-clinical proof of concept. This proposal will test the combination of new cargo and new delivery platform in disease mice models.
This approach is intended to turn on the inactive MeCP2, which would result in cells expressing a normal MeCP2 gene in addition to the mutant one.
The MECP2 gene is silenced on the inactive X chromosome due to the addition of chemical tags called methyl groups. These tags can be thought of as charms added to a charm bracelet. Removing the methyl groups can reactive the gene. The cargo is an engineered protein that includes a small Cas that acts as a GPS and a protein that can remove the methyl groups. The cargo is coated with small peptides that protect and facilitates its entry into cells.
Animals will receive the cargo either by intrathecal or intracerebroventricular (ICV) injection, compatible with human clinical delivery protocols, and evaluated for gene correction by molecular tools, elevation of MeCP2 levels, and behavioral analytics. Studies will use repeat dosing and dose ranging to determine long term correction and durability.
