Abstract
Adeno-associated virus (AAV)-based gene therapy is undergoing major expansion into clinical practice, with two treatments currently being granted Food and Drug Administration (FDA) approval. However, the presence of pre-existing neutralizing antibodies (NAB) is one of the significant hurdles for the clinical application of AAV vectors that significantly limits the patient population, which benefits from the treatment. A reliable diagnostic to evaluate the patient's seropositivity is required to ensure the effectiveness of the AAV-mediated therapeutic. Here, we describe a simple method for the determination of AAV NAB activity based on our finding that Compound C makes HEK293 cell highly permissive for infection by 10 commonly used AAV serotypes.
Highlights
We identified a selective inhibitor of AMP-activated protein kinase (AMPK) Dorsomorphin, known as Compound C (CC), as an enhancer of the infection of HEK293 cells by AAV8, and other ASSOCIATED VIRUS (AAV) serotypes, without cytotoxic effects
AAV vectors used in this study were packaged in HEK293 cells by triple transfection with polyethyleneimine (PEI) and isolated by an iodixanol fourstep gradient followed by ion-exchange column purification as described.[33,34]
We showed that the addition of IL-6 and tumor necrosis factor alpha (TNF-a) to CC treatment improved infection efficiency for all serotypes, but not for AAV3 (Fig. 2A)
Summary
Hormel Institute, University of Minnesota, Austin, Minnesota, USA. Adeno-associated virus (AAV)-based gene therapy is undergoing major expansion into clinical practice, with two treatments currently being granted Food and Drug Administration (FDA) approval. The presence of preexisting neutralizing antibodies (NAB) is one of the significant hurdles for the clinical application of AAV vectors that significantly limits the patient population, which benefits from the treatment. A reliable diagnostic to evaluate the patient’s seropositivity is required to ensure the effectiveness of the AAV-mediated therapeutic. We describe a simple method for the determination of AAV NAB activity based on our finding that Compound C makes HEK293 cell highly permissive for infection by 10 commonly used AAV serotypes
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