Abstract:
This project is designed to identify a noninvasive and highly effective gene delivery strategy to target
specific neural cells in the CNS and to validate this technology by delivering regenerative molecules to
mammals with CNS injury. We will determine whether systemic delivery of our newly engineered AAV9
vectors can transduce most target CNS cells and whether noninvasive delivery of the genes that target
neuronal intrinsic and extrinsic factors can promote robust axon regeneration and functional recovery
in rodents with spinal cord injury (SCI). A major challenge in neuroscience research is to deliver target
genes to specific types of neural cells widely distributed in CNS. Engineered AAV9 vectors usually
show limited efficacy after intravenous (IV) injection by transducing cells only in some CNS regions.
We thus created new AAV9 vectors that include multiple features of engineered AAV9 capsids, aiming
to develop highly efficient BBB-crossing AAV9 vectors (HEBC-AAV9) that can transduce most target
CNS cells after IV injection. In Aim 1, we will study efficiency of our novel HEBC-AAV9-GFP vectors
for selectively transducing each type of neural cells (neurons, astrocytes, oligodendrocytes, and
microglia) in several strains of adult mice. To solve a crucial issue in neuroscience research with this
technology, in Aim 2 we will develop a regenerative therapy for SCI by systemic delivery of genes to
target neuronal let-7 miRNA. After SCI, severed axons fail to regenerate partly because of reduced
intrinsic growth capacity of mature neurons. Many genes are known to control the growth ability of
mature neurons, but none have been translated to clinical use. The best targets are probably those
with potential to impact multiple genes. Among them, let-7 is important for regulating age-dependent
decline in axon regeneration in worms. In Aim 2, we propose to use unique HEBC-AAV9-synapsin
vectors to target neurons selectively for inducing expression of let-7 inhibitor, lin28, and lin41, aiming
to promote robust regeneration of multiple axon tracts by enhancing growth capacity of mature neurons
in SCI rodents. Chondroitin sulfate proteoglycans (CSPGs) generated by glial scars strongly suppress
axon extension and are major extrinsic molecular targets for treating CNS injury. Our lab designed
small peptides to block functions of CSPG receptor LAR, PTPs, and PTPd by targeting their critical
activity domains and demonstrated their high efficiency for promoting axon regrowth. In Aim 3, we will
induce astrocytic expression of secreted 3 peptides for each of LAR, PTPs, and PTPd with HEBC-
AAV9-GFAP vectors, aiming to promote robust axon regeneration after SCI by targeting extrinsic
CSPGs alone or combined with intrinsic let-7 signals. Our new viral vectors should provide a powerful
tool for gene delivery in CNS and for developing effective regenerative therapies for SCI and other
neurological disorders.