Optimization of adeno-associated viral vector-mediated transduction of the corticospinal tract: comparison of four promoters

Abstract

Adeno-associated viral vectors are widely used as vehicles for gene transfer to the nervous system. The promoter and viral vector serotype are two key factors that determine the expression dynamics of the transgene. A previous comparative study has demonstrated that AAV1 displays efficient transduction of layer V corticospinal neurons, but the optimal promoter for transgene expression in corticospinal neurons has not been determined yet. In this paper, we report a side-by-side comparison between four commonly used promoters: the short CMV early enhancer/chicken β actin (sCAG), human cytomegalovirus (hCMV), mouse phosphoglycerate kinase (mPGK) and human synapsin (hSYN) promoter. Reporter constructs with each of these promoters were packaged in AAV1, and were injected in the sensorimotor cortex of rats and mice in order to transduce the corticospinal tract. Transgene expression levels and the cellular transduction profile were examined after 6 weeks. The AAV1 vectors harbouring the hCMV and sCAG promoters resulted in transgene expression in neurons, astrocytes and oligodendrocytes. The mPGK and hSYN promoters directed the strongest transgene expression. The mPGK promoter did drive expression in cortical neurons and oligodendrocytes, while transduction with AAV harbouring the hSYN promoter resulted in neuron-specific expression, including perineuronal net expressing interneurons and layer V corticospinal neurons. This promoter comparison study contributes to improve transgene delivery into the brain and spinal cord. The optimized transduction of the corticospinal tract will be beneficial for spinal cord injury research.

The Potential of Induced Pluripotent Stem Cells to Test Gene Therapy Approaches for Neuromuscular and Motor Neuron Disorders

“…To date, hundreds of natural AAV serotypes, variants and bio-engineered versions have been described (Hester et al, 2009;Choudhury et al, 2016;Deverman et al, 2016;Chan et al, 2017;Hanlon et al, 2019). Beside serotypes, research efforts are also focusing on the combination of the best serotype with the therapeutic and regulatory sequences-such as promoters or enhancers (Colella et al, 2018;Besse et al, 2020;Nieuwenhuis et al, 2020), for efficient, safe and specific transgene expressions (Guilbaud et al, 2019;Hanlon et al, 2019). This will likely contribute to expedite the translational path from bench to clinic.…”

Rescue of behavioral and electrophysiological phenotypes in a Pitt-Hopkins syndrome mouse model by genetic restoration of Tcf4 expression

“…To this end, we packaged a Cre transgene cassette into a recombinant AAV9-derived PHP.eB vector and bilaterally delivered this viral vector to the cerebral ventricles of neonates (Figure 3A). We expressed the Cre cassette under control of the human synapsin promoter (hSyn) for selective expression in neurons (Nieuwenhuis et al, 2021) To test whether Cre expression coincides with Tcf4 reinstatement in Tcf4 STOP/+ mice, we 195 quantified full-length Tcf4 mRNA transcripts upon delivery of PHP.eB/Cre to Tcf4 STOP/+ 196 neonates. The RT-qPCR results confirmed increased relative expression of Tcf4 transcripts from 197 P10 and P17 Tcf4 STOP/+ brains treated with PHP.eB/Cre (Figure 3G, Tcf4 STOP/+ + Vehicle at P10: 1.0 ± 0.0, n = 2; Tcf4 STOP/+ + PHP.eB/Cre at P10: 1.18 ± 0.03, n = 2, Tcf4 STOP/+ + Vehicle at P19: 1.0 ± 0.04, n = 3; Tcf4 STOP/+ + PHP.eB/Cre at P19: 1.37 ± 0.16, n = 2).…”

“…To this end, we packaged a Cre transgene cassette into a recombinant AAV9-derived PHP.eB vector and bilaterally delivered this viral vector to the cerebral ventricles of neonates (Figure 3A). We expressed the Cre cassette under control of the human synapsin promoter (hSyn) for selective expression in neurons (Nieuwenhuis et al, 2021) (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made…”

Simple RGC: ImageJ Plugins for Counting Retinal Ganglion Cells and Determining the Transduction Efficiency of Viral Vectors in Retinal Wholemounts

“…five years [1][2][3][4]. There are also freely available colocalisation programs [5][6][7][8][9], but none are optimised for determining viral transduction efficiencies of RGCs in retinal wholemounts. Furthermore, only a few of the above mentioned programmes have the option to batch-process a large number of images.…”