Therapeutic AAV Gene Transfer to the Nervous System: A Clinical Reality

Abstract

Gene transfer has long been a compelling yet elusive therapeutic modality. First mainly considered for rare inherited disorders, gene therapy may open treatment opportunities for more challenging and complex diseases such as Alzheimer's or Parkinson's disease. Today, examples of striking clinical proof of concept, the first gene therapy drugs coming onto the market, and the emergence of powerful new molecular tools have broadened the number of avenues to target neurological disorders but have also highlighted safety concerns and technology gaps. The vector of choice for many nervous system targets currently is the adeno-associated viral (AAV) vector due to its desirable safety profile and strong neuronal tropism. In aggregate, the clinical success, the preclinical potential, and the technological innovation have made therapeutic AAV drug development a reality, particularly for nervous system disorders. Here, we discuss the rationale, opportunities, limitations, and progress in clinical AAV gene therapy.

Keywords: AAV; adeno-associated virus; astrocyte; gene therapy; gene transfer; immunity; nervous system; neuron; vector.

Figure 1.. AAVVirus and Vector

(A) Genomic structure of the wild-type adeno associated virus (AAV), indicating the AAV2 genes encoding the replication (Rep), capsid (Cap), and assembly-associated protein (APP) products, flanked by inverted terminal repeat (ITR) secondary DNA hairpin structure. (B) A recombinant AAV2-based vector is generated from the wild-type AAV2 genome by eliminating the open reading frames, solely retaining the flanking ITR sequences and replacing it with a transgene of interest upto a combined maximal sequence length of4.7 kb. These AAV2-based vector genomes (v.g.) are packaged in the protective capsid shell by coexpressing in trans the AAV gene products Rep, Cap, and AAP (in addition to essential gene products from helper virus such as human adenovirus type 5). Any AAV2-based vector genome can be packaged into various different types of AAV variant capsid (CapX) by cross-packaging in which the CapX gene is co-expressed with Rep from AAV2. (C) The genomic structure of self-complementary (sc) AAV has within the same total packaging size of 4.7 kb two complementary copies of the same transgene linked by a third ITR sequence that was modified not to be resolved during viral amplification (black dot. (D) The AAV virion is a multimeric protein composed of 60 subunits in a 20-facetted structure with an outer diameter of approximately 25 nm. All subunits are symmetrically integrated in the particle as dimers (red), trimers (blue), and pentamers (green) according to icosahedral symmetries.

Figure 2.. In Vivo Routes of AAV Delivery to the Nervous System

Local injections of vector to the eye or to the cochlea are preferably chosen for the treatment of neurosensory disorders, as a relatively small area needs to be targeted. Intraparenchymal infusion of AAV remains the most commonly used approach to deliver a therapeutic gene to the brain, even though other delivery strategies to the CSF (intra cerebroventricular, intracisternal, and intrathecal routes) or to the bloodstream could be advantageous for the treatment of multifocal diseases. Intranasal delivery is an alternative and non-invasive option potentially suitable to restore the levels of therapeutic lysosomal enzymes, which are secreted and can diffuse through the CNS. Finally, a few motor neuron diseases can be improved after intramuscular or intra-spinal AAV injection.

Figure 3.. AAV9 Brain Transduction after Intravenous Delivery in C57BL/6

Representative images of eGFP fluorescence signal (and DAPI) across three different coronal sections across the brain after intravenous injection of AAV9 harboring a self-complementary genome (4 × 10¹³ vg/kg) expressing CMV.eGFP (Hudry et al., 2018) at the level of the striatum (upper panel), hippocampus (middle panel), or cerebellum (bottom panel). C57BL/6 animals were injected via tail vein at 6–8 weeks of age. Scale bar, 1,000 µm.