Intrathecal Delivery of AAV Vectors in Cynomolgus Macaques for CNS Gene Therapy and Gene Expression Analysis in Microdissected Motor Neurons
This protocol describes a method of delivering adeno-associated viral (AAV) vectors to the intrathecal space of nonhuman primates for CNS-directed gene therapy. It includes the surgical implantation of the catheter, vector infusion, necropsy, laser-capture microdissection of motor neurons, and gene expression analysis. This method allows efficient and reproducible delivery, and would be of interest to test gene therapy vectors for the treatment of disorders of the central nervous system of nonhuman primates. This protocol was tested in cynomolgus macaques and may be adapted for AAV delivery to different species of large animals.
Key wordsAAV Adeno-associated virus Intrathecal Trendelenburg CSF CNS Laser-capture microdissection Motor neurons ddPCR
Florie Borel would like to express her gratitude to Rachel Stock from the Mueller Lab at UMass Medical for generating the images of the laser capture, as well as to Christen Simon and Randy Reed at Northern Biomedical Research for their support.
- 2.Mendell JR, Al-Zaidy S, Shell R, Arnold WD, Rodino-Klapac LR, Prior TW, Lowes L, Alfano L, Berry K, Church K, Kissel JT, Nagendran S, L'Italien J, Sproule DM, Wells C, Cardenas JA, Heitzer MD, Kaspar A, Corcoran S, Braun L, Likhite S, Miranda C, Meyer K, Foust KD, Burghes AHM, Kaspar BK (2017) Single-dose gene-replacement therapy for spinal muscular atrophy. N Engl J Med 377(18):1713–1722. https://doi.org/10.1056/NEJMoa1706198CrossRefPubMedGoogle Scholar
- 3.Gray SJ, Matagne V, Bachaboina L, Yadav S, Ojeda SR, Samulski RJ (2011) Preclinical differences of intravascular AAV9 delivery to neurons and glia: a comparative study of adult mice and nonhuman primates. Mol Ther 19(6):1058–1069. https://doi.org/10.1038/mt.2011.72CrossRefPubMedPubMedCentralGoogle Scholar
- 4.Borel F, Gernoux G, Cardozo B, Metterville JP, Toro Cabreja GC, Song L, Su Q, Gao GP, Elmallah MK, Brown RH Jr, Mueller C (2016) Therapeutic rAAVrh10 mediated SOD1 silencing in adult SOD1(G93A) mice and nonhuman primates. Hum Gene Ther 27(1):19–31. https://doi.org/10.1089/hum.2015.122CrossRefPubMedGoogle Scholar