Five Years of Successful Inducible Transgene Expression Following Locoregional Adeno-Associated Virus Delivery in Nonhuman Primates with No Detectable Immunity

Abstract

Anti-transgene immune responses elicited after intramuscular (i.m.) delivery of recombinant adenoassociated virus (rAAV) have been shown to hamper long-term transgene expression in large-animal models of rAAV-mediated gene transfer. To overcome this hurdle, an alternative mode of delivery of rAAV vectors in nonhuman primate muscles has been described: the locoregional (LR) intravenous route of administration. Using this injection mode, persistent inducible transgene expression for at least 1 year under the control of the tetracycline-inducible Tet-On system was previously reported in cynomolgus monkeys, with no immunity against the rtTA transgene product. The present study shows the long-term follow-up of these animals. It is reported that LR delivery of a rAAV2/1 vector allows long-term inducible expression up to at least 5 years post gene transfer, with no any detectable host immune response against the transactivator rtTA, despite its immunogenicity following i.m. gene transfer. This study shows for the first time a long-term regulation of muscle gene expression using a Tet-On-inducible system in a largeanimal model. Moreover, these findings further confirm that the rAAV LR delivery route is efficient and immunologically safe, allowing long-term skeletal muscle gene transfer.

Intramuscular injection of vectorized-scFvMC1 reduces pathological tau in two different tau transgenic models

“…Also, from a clinical perspective, particularly attractive would be the ability to not only express the scFv but to switch it on or off at will, adding a layer of exogenous control to improve safety. Among the currently available inducer/repressor systems permitting control over gene expression, the tetracycline (Tet)-dependent system is by far the most advanced and most widely used, and has been already tested in association with AAV1 and locoregional muscle gene transfer in non-human primates showing no humoral or cellular responses against the transgene [ 94 ]: we will certainly explore this avenue in our system.…”

Gene transfer to skeletal muscle using hydrodynamic limb vein injection: current applications, hurdles and possible optimizations

“…By contrast, GpG‐enriched pDNA has been shown to protect against anti‐transgene immune response . To date, gene therapy pre‐clinical studies (in animal models) have shown no induction of any deleterious immune reaction against the transgene following HLV injection . However, this remains to be confirmed in humans.…”

“…A non‐viral gene therapy protocol based on the HLV delivery of an EPO‐encoding pDNA was evaluated in SM of subtotal nephrectomied Lewis rats and in rhesus monkeys . Long‐term inducible transgene expression (for at least 5 years) of an EPO‐encoding pDNA was also shown in non‐human primates using an AAV vector delivered by HLV injection; no deleterious immune reaction against the transgene could be detected throughout the duration of the study …”

“…HLV gene transfer to SM is a rather quick and simple technique, which may be applied to muscle diseases using either pDNA or viral (mainly AAV) vectors . It has been evaluated in a wide range of animals, including mice, rats, dogs, and non‐human primates . In all of these diverse models, HLV demonstrated some efficiency and it was basically safe.…”

Genetically Engineering the Nervous System with CRISPR-Cas

“…For example, higher cellular concentrations of Cas9 have been shown to decrease specificity, presumably because off-target cleavage is the only possibility after all target sites have been destroyed (Davis et al, 2015). This observation has raised concerns for therapeutic developments that rely on viral gene transfer, which in the case of AAVmediated gene expression, persists for several years after delivery (Nathwani et al, 2011;Wojno et al, 2013;Colella et al, 2018;Guilbaud et al, 2019). Engineered ribonucleoprotein complexes (RNP; Cas9 protein bound to a guide RNA) and Cas9-encapsulating nanoparticles have been developed as non-viral alternatives for local, transient CRISPR expression in the brain.…”