Engineered recombinant adeno-associated virus (AAV) vectors have advanced the transduction of neurons in the CNS on an expansive, wide-scale basis since the papers first using AAV9 for this purpose. Wide-scale CNS expression is relevant to gene therapy as well as indispensable for basic studies such as disease modeling. For example, the wide-scale gene transfer approach could expedite hypothesis testing in vivo relative to the generation of germ-line transgenic mice for all of the genes of interest. Wide-scale gene transfer is more efficient in neonates than in adults, so improving gene transfer efficiency in adults is an important goal. Here we characterized the relatively novel AAV PHP.EB vector for expansive gene transfer in the CNS of adult rats at three doses. The dose–response data were consistent; expression levels can be controlled in a reproducible manner in the rat from moderate to robust levels. Within the CNS, the AAV PHP.EB-derived expression was neuron-selective to neuron-specific, while outside the CNS, organs such as the liver and heart were transduced by the parenteral gene delivery. Though we demonstrated graded expression levels, only the high dose, 1.2 × 1014 vector genomes/kg, yielded efficient expression in spinal cord motor neurons of the adult rat, so this vector dose would be required for models of spinal cord motor neuron disease. The neuronal expression in the rat CNS was greater with AAV PHP.EB than the previous engineered vector AAV PHP.B. AAV PHP.EB is thus one of the most efficient AAV vectors in the field for CNS gene transfer.
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We thank Thomas Lawson for his charitable donation to amyotrophic lateral sclerosis research and Jean-Charles Paterna (Neuroscience Center Zurich, ZNZ) for critiquing the manuscript. We thank Viviana Gradinaru and Ben Deverman (California Institute of Technology, Pasadena, CA) for the AAV PHP.B and AAV PHP.EB DNAs and discussion. This work was supported by the ALS Association (15-TALS-296) and Meira GTx. Meira GTx had no involvement in the design or execution of the experiments, or in the decision to publish this research. The authors are collaborating with Meira GTx on a pre-clinical gene therapy project for amyotrophic lateral sclerosis.
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Nature Protocols (2019)