Successful Repeated Hepatic Gene Delivery in Mice and Non-human Primates Achieved by Sequential Administration of AAV5 ch and AAV1

Abstract

In the gene therapy field, re-administration of adeno-associated virus (AAV) is an important topic because a decrease in therapeutic protein expression might occur over time. However, an efficient re-administration with the same AAV serotype is impossible due to serotype-specific, anti-AAV neutralizing antibodies (NABs) that are produced after initial AAV treatment. To address this issue, we explored the feasibility of using chimeric AAV serotype 5 (AAV5) and AAV1 for repeated liver-targeted gene delivery. To develop a relevant model, we immunized animals with a high dose of AAV5-human secreted embryonic alkaline phosphatase (hSEAP) that generates high levels of anti-AAV5 NAB. Secondary liver transduction with the same dose of AAV1-human factor IX (hFIX) in the presence of high levels of anti-AAV5 NAB proved to be successful because expression/activity of both reporter transgenes was observed. This is the first time that two different transgenes are shown to be produced by non-human primate (NHP) liver after sequential administration of clinically relevant doses of both AAV5 and AAV1. The levels of transgene proteins achieved after delivery with AAV5 and AAV1 illustrate the possibility of both serotypes for liver targeting. Furthermore, transgene DNA and RNA biodistribution patterns provided insight into the potential cause of decrease or loss of transgene protein expression over time in NHPs.

Immunoadsorption enables successful rAAV5-mediated repeated hepatic gene delivery in nonhuman primates

“…therapy. 5,14,58 Therefore, it cannot be excluded that IA may only be useful for serotypes that allow successful transduction in the presence of some titer of preexisting antibodies and that do not induce effector T cells upon readministration.…”

“…As has been reported by us and others, the AAVmediated hFIX kinetic expression pattern in NHP plasma is characterized by a peak of expression at 7 days (day 56) after administration followed by a decrease and a subsequent stabilization over time, and the expression of hFIX in the NHPs subjected to IA showed similar kinetics. 14,39,40 Following sacrifice, 30 tissue samples were taken throughout the liver for each animal and rAAV5-hSEAP and rAAV5-hFIX-vector DNA copy numbers were determined by qPCR. Similar levels of rAAV-hSEAP-vector DNA copies were measured in the liver tissues of all animals ( Figure 3A-B) and consistent with the hFIX protein data, substantially higher levels (27.86 times) of hFIXvector DNA copies were detected in animals that underwent the IA procedure prior to readministration with rAAV5-hFIX compared with the untreated group ( Figure 3C-D).…”

“…Various approaches have been pursued to overcome the inhibitory effect of NABs such as dose fitting, alternating AAV serotypes, AAV-vector delivery directly to hepatic tissue, capsid engineering of the vector, or use of capsid decoys. 2,[13][14][15][16][17][18] However, those strategies have limitations because they show limited efficacy in the presence of high NABs titers, require the development of another product, or have safety issues. 19,20 Plasmapheresis and immunoadsorption (IA) are extracorporeal blood-purification techniques that have been developed to remove large-molecular-weight substances from the plasma.…”

Gene therapy trials for haemophilia: a step closer to a cure?

“…With AAV antibodies persisting for many years after treatment [14], this might preclude the use of the same vector again or even different serotypes due to cross reactivity. Early pre-clinical data, however, has suggested that it may be possible to redose using alternative serotypes with low cross-reactivity [26,27] or rechallenge with the same serotype in combination with a proteasome inhibitor where antibodies are no longer detectable [28]. Although these finding are encouraging, this is an area that warrants considerable further research.…”

Exploring protein orthogonality in immune space: a case study with AAV and Cas9 orthologs

“…Our analysis suggests that there are no two known AAVs for which exposure to one would guarantee immune naïveté to another across all HLA genotypes. However, immune cross-reaction could be minimized through the use of AAV5 58,59 , the most phylogenetically divergent serotype. Our predictions identify only a single shared highly immunogenic peptide between AAV5 and the commonly used AAV2 and AAV8 in the mouse model (though several other shared peptides of mild MHC affinity exist).…”