Background: It has been reported in preclinical and clinical studies for AAV2 or AAV8 serotypes that pre-existing anti-AAV NABs titers impact negatively transgene expression. Therefore, individuals with pre-existing NABs against AAV above titer of 5 are excluded from most systemic AAV-based clinical trials. We have previously reported that anti-AAV5 NABs titers up to 340 in humans and as high as 1030 in primates did not interfere with the therapeutic efficacy of intravenously administered AAV5 vector. Hence, patients entering the AMT-061 (AAV5-hFIX-Padua) trial are not excluded based on anti-AAV5 NABs levels.

Aims: Overall, those contradictory observations suggest that the neutralization ability of anti-AAV pre-existing NABs in vivo might be different between AAV serotypes.
The aim of this study was to assess the binding characteristics of the pre-existing NABs directed against AAV serotypes 1, 2, 5 and 8.

Methods: Serum samples of 300 healthy donors were analyzed to determine pre-existing anti-AAV NABs titers and avidity (residual binding) of anti-AAV IgGs to specific AAV serotypes.

Results: The results obtained demonstrate that the avidity of pre-existing AAV-specific IgG antibodies in the healthy human population is significantly different between AAV serotypes. Pre-existing anti-AAV2 or anti-AAV8 IgGs form stronger antibody-antigen complexes with respectively AAV2 and AAV8 antigens, while pre-existing anti-AAV5 IgGs generate significantly weaker antibody-antigen complexes with the AAV5 antigen (Figure 1).
Additionally, when analyzing donors that were positive for pre-existing anti-AAV NABs against all three tested serotypes (2, 5 and 8), avidity of pre-existing anti-AAV5 IgGs in 80% of those donors was low. However, avidities of pre-existing anti-AAV2 IgGs or anti-AAV8 IgGs in 91% and 83% of those donors respectively, were high (Figure 2).

Conclusions: These observations support the lack of impact of pre-existing anti-AAV5 NABs measured in vitro on the efficacy of AAV5-based transduction in vivo when compared to pre-existing anti-AAV NABs against serotypes 2 or 8.

[Figure 1. Avidity of AAV-specific IgG antibodies in healthy human population. Significant difference was observed between the different serotypes. ]

[Figure 2. NABs titers against different AAV serotypes. Plotted donors are the ones that tested triple positive, for NABs against AAV2, 5 and 8.]

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ISTH Congress Abstracts - https://abstracts.isth.org/abstract/prevalence-and-affinity-avidity-assessment-of-pre-existing-neutralizing-antibodies-nabs-against-adeno-associated-virus-aav-vector-serotypes-2-5-and-8-analyzed-in-the-serum-of-300-healthy-donors/