Researchers systematically analyzed all available approaches to purify and concentrate recombinant adeno-associated viruses (rAAVs), which are used as vectors to deliver different genetic constructs inside cells. They published their findings in a review article in the journal BioEssays.
rAAVs are used to deliver gene therapies to the body to treat genetic diseases; there are currently 2 rAAV-based gene therapy drugs that are approved by the US Food and Drug Administration. One of these is voretigene neparvovec, which is used to treat inherited retinal disease, and the other is onasemnogene abeparvovec, which is used to treat spinal muscular atrophy.
Another rAAV-based gene therapy approved by the European Medicines Agency is called alipogene tiparvovec, which is used to treat familial lipoprotein lipase deficiency. There are also more than 140 clinical trials testing treatments using rAAV for various genetic diseases including Pompe disease.
For example, a phase 1/2 clinical trial is assessing the safety and bioactivity of AAV2/8-LSPhGAA, a gene therapy that aims to enable the expression of a functional copy of the GAA gene, which is mutated in patients’ hepatocytes who have Pompe disease. The trial is currently recruiting participants in North Carolina.
Read more about experimental therapies for Pompe disease
The purification of rAAV vectors plays a crucial role in their concentration, purity, stability, and efficacy, and there are many purification approaches. These can be nonspecific or specific. Nonspecific approaches include density gradient ultracentrifugation, chromatography based on hydrophobic interactions, ion chromatography, ion-exchange matrices, and size-exclusion chromatography.
Specific approaches include affinity chromatography, immunoaffinity chromatography, and metal affinity chromatography. There are also noninstrumental purification methods, which are predominantly based on selective precipitation or ultrafiltration of rAAV particles.
Each of these methods is very different in terms of mechanism, efficacy, labor time, and cost and the purification algorithms should be chosen based on the purpose of the purification, the authors said. “We strive to simplify the choice of the necessary and sufficient technique based on the experimental needs and available resources of the laboratory.”
Belova L, Kochergin-Nikitsky K, Erofeeva A, Lavrov A, Smirnikhina S. Approaches to purification and concentration of rAAV vectors for gene therapy. Bioessays. Published online April 14, 2022. doi:10.1002/bies.202200019
AAV2/8-LSPhGAA in late-onset Pompe disease. US National Library of Medicine. Updated February 21, 2021. Accessed April 21, 2022.