Interindividual variability in transgene mRNA and protein production following adeno-associated virus gene therapy for hemophilia A

• Published in: Nature medicine Vol. 28; no. 4; pp. 789 - 797
• Main Authors: Fong, Sylvia, Yates, Bridget, Sihn, Choong-Ryoul, Mattis, Aras N., Mitchell, Nina, Liu, Su, Russell, Chris B., Kim, Benjamin, Lawal, Adebayo, Rangarajan, Savita, Lester, Will, Bunting, Stuart, Pierce, Glenn F., Pasi, K. John, Wong, Wing Yen
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01-04-2022; Nature Publishing Group
• Subjects: 692/308/2779/109/1940; 692/308/2779/109/1941; 692/699/1541; 692/700/565/201; Biomedical and Life Sciences; Biomedicine; Biopsy; Cancer Research; Coagulation factors; Dependovirus - genetics; Dependovirus - metabolism; Dysplasia; Endoplasmic reticulum; Factor VIII - genetics; Factor VIII - therapeutic use; Factor VIII deficiency; Fibrosis; Gene expression; Gene therapy; Genetic Therapy - methods; Genetic Vectors - genetics; Genomes; Hemophilia; Hemophilia A - genetics; Hemophilia A - therapy; Hepatocytes; Histology; Histopathology; Humans; Infectious Diseases; Intravenous administration; Intravenous infusion; Liver; Metabolic Diseases; Molecular Medicine; Molecular modelling; Neurosciences; Proteins; RNA, Messenger; Transcription; Transduction; Transgenes; Transgenes - genetics
• ISSN: 1078-8956; 1546-170X
• DOI: 10.1038/s41591-022-01751-0

Factor VIII gene transfer with a single intravenous infusion of valoctocogene roxaparvovec (AAV5-hFVIII-SQ) has demonstrated clinical benefits lasting 5 years to date in people with severe hemophilia A. Molecular mechanisms underlying sustained AAV5-hFVIII-SQ-derived FVIII expression have not been studied in humans. In a substudy of the phase 1/2 clinical trial (
NCT02576795
), liver biopsy samples were collected 2.6–4.1 years after gene transfer from five participants. Primary objectives were to examine effects on liver histopathology, determine the transduction pattern and percentage of hepatocytes transduced with AAV5-hFVIII-SQ genomes, characterize and quantify episomal forms of vector DNA and quantify transgene expression (hFVIII-SQ RNA and hFVIII-SQ protein). Histopathology revealed no dysplasia, architectural distortion, fibrosis or chronic inflammation, and no endoplasmic reticulum stress was detected in hepatocytes expressing hFVIII-SQ protein. Hepatocytes stained positive for vector genomes, showing a trend for more cells transduced with higher doses. Molecular analysis demonstrated the presence of full-length, inverted terminal repeat-fused, circular episomal genomes, which are associated with long-term expression. Interindividual differences in transgene expression were noted despite similar successful transduction, possibly influenced by host-mediated post-transduction mechanisms of vector transcription, hFVIII-SQ protein translation and secretion. Overall, these results demonstrate persistent episomal vector structures following AAV5-hFVIII-SQ administration and begin to elucidate potential mechanisms mediating interindividual variability.
The analysis of liver biopsy samples after AAV gene therapy for hemophilia A reveals normal histology and long-term persistence of the episomal vector, and identifies potential factors contributing to interindividual variability of transgene expression.