Bejamine Deverman's team at the Broad Institute recently published a research paper titled "An AAV capsid reprogrammed to bind human transferrin receptor mediates brain-wide gene delivery" online in Science. The study designed an AAV capsid, BI-hTFR1, that binds to the human transferrin receptor (TfR1), a protein expressed on the blood-brain barrier (BBB). Compared with AAV9, BI-hTFR1 has a higher active transport capacity across the human brain endothelial cell layer and provides a 40-50-fold enhancement of reporter gene expression in the CNS of mice carrying a human TFRC knock-in. This enhanced tropism is CNS specific and is not present in wild-type mice.
When used to deliver the GBA1 gene, mutations in the gene cause Gaucher disease and are associated with Parkinson's disease. BI-hTFR1 significantly increased glucocerebrosidase activity in the brain and cerebrospinal fluid compared to AAV9. These findings establish BI-hTFR1 as a promising vector for human CNS gene therapy.
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In developing more efficient gene therapy delivery vehicles, a key challenge is designing vectors with a known mechanism of action (MOA) that is relevant to human patients. Traditionally, to enhance adeno-associated virus (AAV) tropism, researchers have relied primarily on capsid library screening in animals. Over the past two decades, many research groups have used this approach because it can be successful without requiring prior knowledge of the MOA. However, most vectors screened in animals fail to achieve the same effects in preclinical models. Despite extensive searches, no AAV capsids with clear transforming potential and known MOA for entry into the CNS have been described.
Here, researchers approach a different angle by first selecting AAV capsids for a specific MOA (i.e., binding to the human transferrin receptor (TfR1)) and showing that one of these capsids can cross the blood-brain barrier (BBB). The researchers chose TfR1 as a target because it is highly expressed on the human BBB and is capable of mediating constitutive, ligand-independent receptor-mediated transcellular transport (RMT) through CNS vessels. And TfR1 is well documented in increasing CNS biologics delivery in mice, non-human primates (NHP), and humans.
The researchers first screened a library of 7-peptide-modified AAV9 capsids for their ability to bind human TfR1 in vitro. The top performing capsid, AAV-BI-hTFR1, showed more efficient gene delivery to human brain endothelial cells and improved active transport through human blood vessel monolayers. When systemically injected into TFRC knock-in (KI) mice carrying a chimeric Tfrc gene with humanized extracellular domain, BI-hTFR1 transduced a large proportion of neurons and astrocytes in multiple brain regions. This enhanced tropism was entirely dependent on the interaction with humanized TfR1, as no enhancing effect was observed in wild-type (WT) mice. This tropism is selectively enhanced in the CNS, consistent with the high expression levels of TFRC in CNS vessels.
Figure 1. BI-hTFR1 efficiently delivers GBA1 and increases GCase activity in the brains of TFRC KI mice. (Huang, Qin, et al. 2024)
To explore their ability to deliver therapeutically relevant cargo, the researchers intravenously injected BI-hTFR1 or AAV9 carrying the human glucocerebrosidase 1 (GBA1) gene into TFRC KI mice. Inactivating mutations in GBA1 are a major cause of Gaucher disease, a lysosomal storage disorder that commonly affects the CNS, and GBA1 mutations are a genetic risk factor for Parkinson's disease and dementia with Lewy bodies. Following systemic delivery of BI-hTFR1:GBA1 in adult TFRC KI mice, expression increased throughout the brain and glucocerebrosidase (GCase) activity increased in the brain and cerebrospinal fluid (CSF), which was not observed when using AAV9. By binding directly to TfR1, the human receptor for natural protein ligands and therapeutic biologics, BI-hTFR1 represents a promising vector for the development of human gene therapies targeting the CNS.
Reference
Huang, Qin, et al. "An AAV capsid reprogrammed to bind human transferrin receptor mediates brain-wide gene delivery." Science (2024): eadm8386.