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Title: Development of new gene editing delivery systems that cross the blood brain barrier for the treatment of Krabbe Disease and other lysosomal storage disorders
Supervisor: Sara Benedetti, Alessia Cavazza, Asma Naseem
Project Description:
Infantile Krabbe disease (KD) is a rare neurological lysosomal storage disorder (LSD) caused by a deficiency in the lysosomal enzyme β-galactocerebrosidase (GALC), leading to demyelination of the central and peripheral nervous systems (CNS and PNS) and macrophage-mediated neuroinflammation, resulting in progressive neurodegeneration and early death. There is no definitive treatment for KD, therefore there is an urgent need for a safe and effective therapy capable of correcting the underlying genetic mutations and preventing neuropathy in affected infants1. To address this unmet need, we have developed a perinatal in vivo gene editing platform to correct GALC mutations in the CNS, PNS, and hematopoietic system simultaneously using adenine base editors (ABEs) delivered via Virus Like Particles (VLPs) 2. Intravenous injections of ABE-VLPs in Twitcher mice demonstrated transduction of hematopoietic cells but low targeting of the brain, due to the presence of the Blood Brain Barrier (BBB), a semi-permeable membranous barrier, localized at the interface between the blood and the cerebral tissue, composed of a complex system of endothelial cells, astroglia, and pericytes. A potential smart way to overcome the BBB is represented by the so called “Trojan horse” approach. This strategy takes advantage of specialized receptors expressed by endothelial cells that are capable of shuttling from the luminal to the abluminal side of the BBB while carrying cargoes with them, in a process named receptor-mediated transcytosis (RMT)3. In this project, we aim to leverage predictable antibody-antigen interactions to transiently deliver ABE-VLPs in the CNS via RMT across the BBB. To achieve this, firstly we will study in depth the structure and integrity of the BBB in KD, utilizing patient-specific high-fidelity 3D BBB models alongside our mouse model. Generally, research on the BBB in neurological LSDs, and KD patients in particular, is currently limited; yet structural and functional impairment of BBB has been reported in some mouse models4, with a report of compromised BBB integrity in post-mortem brain tissues of few patients too5. These findings may have implications in delivery of therapeutics to the CNS of KD patients and give invaluable insight for the design of our CNS-targeted ABE-VLPs. Secondly, we will develop targeted gene editing delivery tools by decorating VLP viral envelopes with synthetic antibody-like moieties binding to receptors highly expressed on the BBB and capable of RMT, to shuttle ABE-VLPs in the brain parenchyma and genetically correct brain cell types involved in KD. Through the use of 2D-cultures, organ-on-chip microfluidics and in vivo models, we will assess the therapeutic potential of BBB-targeted ABE delivery vehicles to treat KD, potentially serving as a proof-of-concept for other early-onset neurodegenerative diseases.
1. Heller G, et al 2023 doi: 10.1016/j.ymthe.2022.09.017.
2. Banskota S, et al 2022 doi: 10.1016/j.cell.2021.12.021.
3. Bellettato CM, Scarpa M. 2018 doi: 10.1186/s13052-018-0563-0.
4. Garbuzova-Davis S, et al 2011 doi: 10.1371/journal.pone.0016601.
5. Garbuzova-Davis S, et al 2013 doi: 10.1186/1471-2377-13-174
Contact Information:
Sara Benedetti and Alessia Cavazza