Intrathecal sc-AAV9-CB-GFP: Systemic Distribution Predominates Following Single-Dose Administration in Cynomolgus Macaques.

Biodistribution of self-complementary adeno-associated virus-9 (scAAV9)-chicken β-actin promoter-green fluorescent protein (GFP) was assessed in juvenile cynomolgus macaques infused intrathecally via lumbar puncture or the intracisterna magna (1.0×10 or 3.0×10 vg/animal), with necropsy 28 days later. Our results characterized central nervous system biodistribution compared with systemic organs/tissues by droplet digital polymerase chain reaction for DNA and hybridization. Green fluorescent protein expression was characterized by Meso Scale Discovery electrochemiluminescence immunosorbent assay and immunohistochemistry (IHC). Biodistribution was widespread but variable, with vector DNA and GFP expression greatest in the spinal cord, dorsal root ganglia (DRG), and certain systemic tissues (e.g., liver), with low concentrations in many brain regions despite direct cerebrospinal fluid administration. Transduction and expression were observed primarily in perivascular astrocytes in the brain, with a paucity in neurons. Greater GFP expression was observed in hepatocytes, striated myocytes, cardiomyocytes, spinal cord lower motor neurons, and DRG sensory neurons by IHC. These results should be considered when evaluating scAAV9-based intrathecal delivery with the current expression cassette as a modality for neurologic diseases that require widespread brain neuronal expression. This capsid/expression cassette combination may be better suited for diseases that express a secreted protein and/or do not require widespread brain neuronal transduction.