Unique Glycan Signatures Regulate Adeno-Associated Virus Tropism in the Developing Brain

Abstract

Adeno-associated viruses (AAV) are thought to spread through the central nervous system (CNS) by exploiting cerebrospinal fluid (CSF) flux and hijacking axonal transport pathways. The role of host receptors that mediate these processes is not well understood. In the current study, we utilized AAV serotype 4 (AAV4) as a model to evaluate whether ubiquitously expressed 2,3-linked sialic acid and the developmentally regulated marker 2,8-linked polysialic acid (PSA) regulate viral transport and tropism in the neonatal brain. Modulation of the levels of SA and PSA in cell culture studies using specific neuraminidases revealed possibly opposing roles of the two glycans in AAV4 transduction. Interestingly, upon intracranial injection into lateral ventricles of the neonatal mouse brain, a low-affinity AAV4 mutant (AAV4.18) displayed a striking shift in cellular tropism from 2,3-linked SA ؉ ependymal lining to 2,8-linked PSA ؉ migrating progenitors in the rostral migratory stream and olfactory bulb. In addition, this gain-of-function phenotype correlated with robust CNS spread of AAV4.18 through paravascular transport pathways. Consistent with these observations, altering glycan dynamics within the brain by coadministering SA-and PSA-specific neuraminidases resulted in striking changes to the cellular tropisms and transduction efficiencies of both parental and mutant vectors. We postulate that glycan signatures associated with host development can be exploited to redirect novel AAV vectors to specific cell types in the brain. IMPORTANCEViruses invade the CNS through various mechanisms. In the current study, we utilized AAV as a model to study the dynamics of virus-carbohydrate interactions in the developing brain and their impact on viral tropism. Our findings suggest that carbohydrate content can be exploited to regulate viral transport and tropism in the brain.

Systemic delivery of adeno-associated viral vectors

“…The nature and abundance of these extracellular glycans vary dramatically between different species and at different developmental stages. Since AAV attachment is a determining factor in systemic delivery, cautions should be taken to extrapolate re-engineered capsids for different applications [102,115,116,137]. …”

Adeno-Associated Virus Technologies and Methods for Targeted Neuronal Manipulation

“…Beyond neurons, ICV injections also provide access to periventricular cell populations. For example, after ICV injection, AAV4 can be used to transduce the ependymal cells (Liu et al, 2005), and two engineered AAV capsids, SCH9 and AAV4.18, enable transduction of subventricular zone neural progenitors (Murlidharan et al, 2015;Ojala et al, 2018). IT injection can be used to deliver genes to spinal cord motor neurons and dorsal root ganglions (Federici et al, 2012;Foust et al, 2013;Schuster et al, 2014). Retrograde and anterograde transport for circuit studies AAV vectors are also commonly used as part of circuit studies.…”

“…While powerful technologies are being developed, each has technical limits which need to be considered both when designing experiments and when interpreting results. To this end, improved platforms for sharing (Borel et al, 2016;Federici et al, 2012;Gurda et al, 2016;Samaranch et al, 2013) AAV4 enables transduction of ependymal cells (Liu et al, 2005) AAV SCH9 and AAV4.18 enable SVZ progenitor cell transduction (Murlidharan et al, 2015;Ojala et al, 2018) ( Gray et al, 2011;Klein et al, 1998;McCown et al, 1996;Paterna et al, 2000;Tenenbaum et al, 2004;Thomsen et al, 1984;Yaguchi et al, 2013) CAG, CMV enhancer, CBA promoter, globin intron Ubiquitous 1700 (Miyazaki et al, 1989) CAGGS, CMV immediateearly enhancer, CBA promoter, CBA intron1/exon1…”

Engineering AAV receptor footprints for gene therapy

“…Although defective upon systemic injection, one particular mutant AAV4.18, containing K492E, K503E and N585S substitutions displayed a unique transduction profile in the brain. Specifically, increased spread of AAV4.18 throughout the brain parenchyma and selective transduction of migrating progenitors was observed [57]. This acquired tropism and increased CNS spread were attributed to a switch in receptor specificity from MUC to a2,8-linked polysialic acid (PSA), an established marker of neurogenesis [57].…”

“…Specifically, increased spread of AAV4.18 throughout the brain parenchyma and selective transduction of migrating progenitors was observed [57]. This acquired tropism and increased CNS spread were attributed to a switch in receptor specificity from MUC to a2,8-linked polysialic acid (PSA), an established marker of neurogenesis [57]. Based on similar rationale, re-engineering of residues within the AAV5 SIA binding footprint is likely to yield novel AAV5 mutants with altered glycan binding affinity and potentially, altered tropism.…”