Abstract
The biodistribution of AAVHSC7, AAVHSC15, and AAVHSC17 following systemic delivery was assessed in cynomolgus macaques (Macaca fascicularis). Animals received a single intravenous (IV) injection of a self-complementary AAVHSC-enhanced green fluorescent protein (eGFP) vector and tissues were harvested at two weeks post-dose for anti-eGFP immunohistochemistry and vector genome analyses. IV delivery of AAVHSC vectors produced widespread distribution of eGFP staining in glial cells throughout the central nervous system, with the highest levels seen in the pons and lateral geniculate nuclei (LGN). eGFP-positive neurons were also observed throughout the central and peripheral nervous systems for all three AAVHSC vectors including brain, spinal cord, and dorsal root ganglia (DRG) with staining evident in neuronal cell bodies, axons and dendritic arborizations. Co-labeling of sections from brain, spinal cord, and DRG with anti-eGFP antibodies and cell-specific markers confirmed eGFP-staining in neurons and glia, including protoplasmic and fibrous astrocytes and oligodendrocytes. For all capsids tested, 50 to 70% of glial cells (S100-β+) and on average 8% of neurons (NeuroTrace+) in the LGN were positive for eGFP expression. In the DRG, 45 to 62% of neurons and 8 to 12% of satellite cells were eGFP-positive for the capsids tested. eGFP staining was also observed in peripheral tissues with abundant staining in hepatocytes, skeletal- and cardio-myocytes and in acinar cells of the pancreas. Biodistribution of AAVHSC vector genomes in the central and peripheral organs generally correlated with eGFP staining and were highest in the liver for all AAVHSC vectors tested. These data demonstrate that AAVHSCs have broad tissue tropism and cross the blood-nerve and blood-brain-barriers following systemic delivery in nonhuman primates, making them suitable gene editing or gene transfer vectors for therapeutic application in human genetic diseases.
Highlights
The introduction of adeno-associated viruses (AAVs) as vectors for transgene delivery has had a significant impact on clinical development of genetic therapies for a range of human diseases [1,2,3,4,5,6,7,8]
To evaluate biodistribution of AAVHSCs after systemic delivery, self-complementary AAVHSC7, AAVHSC15 and AAVHSC17 each packaging an enhanced green fluorescent protein (eGFP) reporter transgene driven by the ubiquitously-expressing chicken beta actin (CBA) promoter were prepared
Following treatment with scAAVHSC17-CBA-eGFP, eGFP IHC detection was characterized across the whole brain in one monkey to evaluate the coronal biodistribution of eGFP across both hemispheres. eGFP detection was observed throughout the brain in both white and grey matter regions including the cerebral cortex, caudate nuclei and putamen (Fig 1A and 1B)
Summary
The introduction of adeno-associated viruses (AAVs) as vectors for transgene delivery has had a significant impact on clinical development of genetic therapies for a range of human diseases [1,2,3,4,5,6,7,8]. A property of Clade F vectors is the ability to traverse the blood-brain-barrier (BBB) following intravascular administration, resulting in transduction of neurons and glia in neonatal and adult mice [15, 16], rats [17], cats [16] and in nonhuman primates [18, 19] These data suggested that systemic delivery of a BBB-penetrating AAV could be used to deliver therapeutic genes to the central nervous system (CNS). Extended to human patients with spinal muscular atrophy, systemic delivery of AAV9-SMN1 increased survival, improved motor milestone achievement and partially restored motor function [5] These data led to the recent approval of Zolgensma for pediatric patients with spinal muscular atrophy [20]
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