Year: 2026
Type: article
Abstract: The potency assay is critical to ensure the effectiveness and consistency of recombinant Adeno-associated Virus (AAV) gene therapy vectors, especially clinical-grade products. AAV serotype 9 (AAV9), known for its neurotropic properties and ability to cross the blood-brain barrier, has been a favored vector for targeting neurogenetic diseases. However, assessing AAV9 biopotency has been challenging due to the insusceptibility of the commonly used cell lines to AAV9. To address this, we utilized a cell-based potency assay using the liver-derived human hepatoma (HuH-7) cell line to evaluate infection by self-complementary (sc)-AAV9 vector expressing human N-sulfoglucosamine sulfohydrolase (hSGSH), currently undergoing evaluation as a potential treatment for Mucopolysaccharidosis (MPS) IIIA. The potency of various scAAV9-hSGSH vector batches was tested in HuH-7 cells which reproducibly expressed the transgene, resulting in measurable SGSH production. The SGSH expression and vector genome copies of various vector batches correlated linearly with the viral vector dose (R2 = 0.71-0.95), indicating a generally strong correlation. The reproducibility of the assay was demonstrated by consistent vector copy numbers and SGSH activity in transduced cells across multiple independent runs. Statistical analysis of the results showed high reliability, with relative intra-assay consistency showing a coefficient of variation (CV) of less than 20%) and inter-assay reproducibility with a CV of less than 25%) affirming the precision of the test. Additionally, our data also demonstrate that long-term (>2.5 years) storage at 2-4°C had no impact on the biopotency of rAAV9 vector confirming long-term stability of the vectors. Hence, we have effectively assessed the biopotency of rAAV9 vector in vitro utilizing HuH-7 cells. Overall, this in vitro assay provides a practical and reliable method to assess AAV9 potency, offering a valuable alternative to animal models and supporting the functional quality and consistency of AAV9 gene therapy vector products in general.
Source: PLoS ONE
Institution: University of North Carolina at Chapel Hill
Language: English
Subfield: Genetics
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $2,382
Funders: Deanship of Scientific Research, Imam Mohammed Ibn Saud Islamic University, Ontario SPOR SUPPORT Unit, Foundation for the National Institutes of Health, Bill and Melinda Gates Foundation, United Nations Educational, Scientific and Cultural Organization, University of Southern California, Leakey Foundation, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, Naito Foundation, Japan Agency for Medical Research and Development, Canada-Israel Industrial Research and Development Foundation, Radiation Oncology Institute, Takeda Foundation, Centers for Disease Control and Prevention Foundation, National Institute for Health and Care Research, Københavns Universitet, National Natural Science Foundation of China, King Saud University, Consejo Nacional de Investigaciones Científicas y Técnicas, Lundbeckfonden, Korea Research Institute of Standards and Science, National Research Foundation of Korea, Department of Health and Aged Care, Australian Government, Narodowe Centrum Nauki, Japan International Cooperation Agency, China Scholarship Council, Hamadan University of Medical Sciences, Jimma University, Natural Science Foundation of Chongqing, Secretaría de Ciencia, Tecnología e Innovación del Distrito Federal, Nagase Science Technology Foundation, Novo Nordisk Fonden, Consejo Nacional de Ciencia, Tecnología e Innovación Tecnológica, Education Department of Hainan Province, Fondation Botnar, Alcohol Change UK, Ariel University, State Key Laboratory of Radiation Medicine and Protection, Mito Foundation, National Institutes of Health, Yale School of Medicine, Canadian Institutes of Health Research, Yale School of Public Health, Yale University, Arts and Humanities Research Council, Japan Society for the Promotion of Science, Agencia Nacional de Promoción Científica y Tecnológica, Agencia Española de Cooperación Internacional para el Desarrollo, Beijing Nova Program, Kasetsart University Research and Development Institute, Dirección General de Asuntos del Personal Académico, Universidad Nacional Autónoma de México, National Key Research and Development Program of China, Medical and Health Research Project of Zhejiang Province, National Institute on Aging, National Institute on Drug Abuse, National Institute of General Medical Sciences, National Institute of Allergy and Infectious Diseases, National Institute of Arthritis and Musculoskeletal and Skin Diseases, Natural Science Foundation of Zhejiang Province, Danish Cardiovascular Academy
OpenAlex ID: W7131647748