A long-term study of AAV gene therapy in dogs with hemophilia A identifies clonal expansions of transduced liver cells

Nguyen, Giang N.; Everett, J.K.; Kafle, Samita; Roche, Aoife M.; Raymond, Hayley E.; Leiby, Jacob S.; Wood, Christian M; Assenmacher, Charles Antoine; Merricks, Elizabeth P.; Long, C. Tyler; Kazazian, Haig H.; Nichols, Timothy C.; Bushman, Frederic D.; Sabatino, Denise E.

doi:10.1038/s41587-020-0741-7

Cited by 273 publications

(239 citation statements)

References 58 publications

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“…However, the integrations appeared to be random, and were not associated with HCC. Nguyen et al (2020), reported results from hemophilia A dogs that were treated with an AAV-canine FVIII therapy and followed for up to 10 years. These dogs had numerous integration events and evidence of clonal expansion near genes previously associated with growth control, but examination of the liver at necropsy did not show evidence of neoplasia in any dog.…”

Section: Potential For Insertional Mutagenesis and Tumorgenicitymentioning

confidence: 99%

Development challenges associated with rAAV-based gene therapies

et al. 2021

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The number of gene therapies in development continues to increase, as they represent a novel method to treat, and potentially cure, many diseases. Gene therapies can be conducted with an in vivo or ex vivo approach, to cause gene augmentation, gene suppression, or genomic editing. Adeno-associated viruses are commonly used to deliver gene therapies, but their use is associated with several manufacturing, nonclinical and clinical challenges. As these challenges emerge, regulatory agency expectations continue to evolve. Following administration of rAAV-based gene therapies, nonclinical toxicities may occur, which includes immunogenicity, hepatotoxicity, neurotoxicity, and the potential risks for insertional mutagenesis and subsequent tumorgenicity. The mechanism for these findings and translation into the clinical setting are unclear at this time but have influenced the nonclinical studies that regulatory agencies are increasingly requesting to support clinical trials and marketing authorizations. These evolving regulatory expectations and toxicities, as well as future nonclinical considerations, are discussed herein.

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Section: Potential For Insertional Mutagenesis and Tumorgenicitymentioning

confidence: 99%

Development challenges associated with rAAV-based gene therapies

et al. 2021

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“…3H). This finding had striking similarities with viral integrations when using AAV to deliver Cas9 (Hanlon et al 2019;Nguyen et al 2021), which indicates that ITR or similar fragile sites is a significant cause for vector integrations.…”

Section: Predictable Small Insertions and Deleterious Plasmid Integrationsmentioning

confidence: 77%

Global detection of DNA repair outcomes induced by CRISPR-Cas9

Liu

Zhang

Xin

et al. 2021

Preprint

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CRISPR-Cas9 generates double-stranded DNA breaks (DSBs) to activate cellular DNA repair pathways for genome editing. The repair of DSBs leads to small insertions or deletions (indels) and other complex byproducts, including large deletions and chromosomal translocations. Indels are well understood to disrupt target genes, while the other deleterious byproducts remain elusive. We developed a new in silico analysis pipeline for the previously described primer-extension-mediated sequencing assay to comprehensively characterize CRISPR-Cas9-induced DSB repair outcomes in human or mouse cells. We identified tremendous deleterious DSB repair byproducts of CRISPR-Cas9 editing, including large deletions, plasmid integrations, and chromosomal translocations. We further elucidated the important roles of microhomology, chromosomal interaction, recurrent DSBs, and DSB repair pathways in the generation of these byproducts. Our findings provide an extra dimension for genome editing safety besides off-targets. And caution should be exercised to avoid not only off-target damages but also deleterious DSB repair byproducts during genome editing.

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“…A recent study of dogs treated for hemophilia A with rAAV and followed for 8-10 years showed that the dogs with stable expression of Factor 8 have an accumulation of integrated rAAV in liver cells and have developed what appears to be dominant clones. 34 In order to understand the theoretical scope of the potential utility and limitations of ERT, rAAV as a sole therapy treatment option, and the bimodal approach, we have constructed a table for comparison ( Table 2). Included in the comparison are aspects related to treatment schedule, access, mechanism of action, and potential cost.…”

Section: Discussionmentioning

confidence: 99%

Gene delivery using AAV8 in vivo for disease stabilization in a bimodal gene therapy approach for the treatment of ADA-deficient SCID

Carbonaro-Sarracino

Chun

Clark

et al. 2021

Molecular Therapy - Methods & Clinical Development

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Adenosine deaminase (ADA) deficiency is an inborn error of metabolism affecting multiple systems and causing severe combined immunodeficiency. We tested intravenous administration of recombinant adeno-associated virus (AAV) 2/8-ADA vector in ADA-deficient neonate and adult mice or as part of a bimodal approach comprised of rAAV treatment at birth followed by infusion of lentiviral vector (LV)-modified lineagedepleted bone marrow cells at 8 weeks. ADA À/À mice treated with rAAV and enzyme replacement therapy (ERT) for 30 days were rescued from the lethal pulmonary insufficiency, surviving out to 180 days without further treatment. rAAV vector copy number (VCN) was highest in liver, lung, and heart and was associated with near-normal ADA activity and thymocyte development. In the bimodal approach, rAAV-mediated ADA expression supported survival during the 4 weeks before infusion of the LV-modified bone marrow cells and during the engraftment period. Conditioning prior to infusion may have resulted in the replacement of rAAV marked cells in marrow and liver, with LV VCN 100-to 1,000-fold higher in hematopoietic tissue compared with rAAV VCN, and was associated with immune cell reconstitution. In conclusion, a bimodal approach may be an alternative for patients without reliable access to ERT before receiving a stem cell transplant or gene therapy.

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A long-term study of AAV gene therapy in dogs with hemophilia A identifies clonal expansions of transduced liver cells

Cited by 273 publications

References 58 publications

Development challenges associated with rAAV-based gene therapies

Development challenges associated with rAAV-based gene therapies

Global detection of DNA repair outcomes induced by CRISPR-Cas9

Gene delivery using AAV8 in vivo for disease stabilization in a bimodal gene therapy approach for the treatment of ADA-deficient SCID

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