DNA ejection and AAV capsid degradation under thermal stress: Insights from CDMS and native digestion strategies.
- 1Analytical and Biological Mass Spectrometry, Regeneron Pharmaceutical Inc, 777 Old Saw Mill River Road, Tarrytown, NY 10591, USA.
- 2Drug Product Development and Technology, Regeneron Pharmaceutical Inc, 777 Old Saw Mill River Road, Tarrytown, NY 10591, USA.
- 3Product Biochemistry, Regeneron Pharmaceutical Inc, 777 Old Saw Mill River Road, Tarrytown, NY 10591, USA.
Abstract
Adeno-associated viruses (AAVs) are the predominant vectors for in vivo gene therapy, enabling tissue-specific gene delivery and contributing to a rapidly expanding landscape of approved therapies. As AAV-based products advance toward widespread clinical use, ensuring capsid stability throughout manufacturing, storage, and administration has become a critical quality and regulatory consideration. Despite this importance, the molecular mechanisms governing AAV capsid degradation under moderate, prolonged thermal stress remain insufficiently characterized. Here, we integrate charge detection mass spectrometry (CDMS) and native digestion strategies to elucidate AAV degradation pathways under extended thermal stress. Using AAV1 and AAV8 as model systems, we demonstrate that genome release proceeds through intermediate DNA-extrusion states that retain the mass of full capsids while exhibiting elevated charge states. DNA ejection ultimately drives the conversion of full capsids into empty forms, generating distinct, structurally compromised intermediates influenced by serotype, genome size, and the degree of thermal stress. Furthermore, DNA extrusion/ejection occurs cooperatively with VP1u externalization, facilitating DNA-viral protein-mediated aggregation that can be largely reversed by native enzymatic digestion of either the DNA or protein components. Together, these findings offer new insights into AAV degradation pathways under thermal stress and establish an effective analytical framework for AAV stability assessment.
Related
- Enhancing food digestion and nutrient absorption through pre-digestion: Tailored strategies for special populations
- Full-fat soybean origin and thermal processing affect protein secondary structure, in vitro digestion kinetics, and predicted standardized ileal amino acid digestibility in broiler chickens.
- Cascading fishery collapse associated with thermal habitat degradation, marine heatwaves, and predator influx
- Thermal Stability of Aliphatic and Semiaromatic Polyamides: Tailoring the Choice of Isothermal Aging Temperature by Using Kinetic Models of Degradation
- Analysis of structural modifications and metagenomic sequencing during the ensiling of Caragana korshinskii provides insights into how extrusion pretreatment promotes lignocellulose degradation
BibTeX
@article{du2026ejection,
title = {{DNA ejection and AAV capsid degradation under thermal stress: Insights from CDMS and native digestion strategies.}},
author = {Chen Du and Shuli Tang and Song Nie and Yuetian Yan and Victoria C Cotham and Wen Shi and Yan Zhao and Kuan-Yu Lai and Li Zhi and Lin Luo and Yuan Cao and Serena Wang and Dingjiang Liu and Mohammed Shameem and Glynn O'Grady and Nina Liu and Michael P Rosconi and Erica A Pyles and Shunhai Wang and Ning Li},
journal = {Molecular therapy. Advances},
year = {2026},
month = sep,
doi = {10.1016/j.omta.2026.201855},
url = {https://europepmc.org/article/MED/42910121},
note = {CC BY-NC-ND},
}
RIS
TY - JOUR
TI - DNA ejection and AAV capsid degradation under thermal stress: Insights from CDMS and native digestion strategies.
AU - Chen Du
AU - Shuli Tang
AU - Song Nie
AU - Yuetian Yan
AU - Victoria C Cotham
AU - Wen Shi
AU - Yan Zhao
AU - Kuan-Yu Lai
AU - Li Zhi
AU - Lin Luo
AU - Yuan Cao
AU - Serena Wang
AU - Dingjiang Liu
AU - Mohammed Shameem
AU - Glynn O'Grady
AU - Nina Liu
AU - Michael P Rosconi
AU - Erica A Pyles
AU - Shunhai Wang
AU - Ning Li
PY - 2026
DA - 2026/09/29
JO - Molecular therapy. Advances
DO - 10.1016/j.omta.2026.201855
UR - https://europepmc.org/article/MED/42910121
AB - Adeno-associated viruses (AAVs) are the predominant vectors for in vivo gene therapy, enabling tissue-specific gene delivery and contributing to a rapidly expanding landscape of approved therapies. As AAV-based products advance toward widespread clinical use, ensuring capsid stability throughout manufacturing, storage, and administration has become a critical quality and regulatory consideration. Despite this importance, the molecular mechanisms governing AAV capsid degradation under moderate, prolonged thermal stress remain insufficiently characterized. Here, we integrate charge detection mass spectrometry (CDMS) and native digestion strategies to elucidate AAV degradation pathways under extended thermal stress. Using AAV1 and AAV8 as model systems, we demonstrate that genome release proceeds through intermediate DNA-extrusion states that retain the mass of full capsids while exhibiting elevated charge states. DNA ejection ultimately drives the conversion of full capsids into empty forms, generating distinct, structurally compromised intermediates influenced by serotype, genome size, and the degree of thermal stress. Furthermore, DNA extrusion/ejection occurs cooperatively with VP1u externalization, facilitating DNA-viral protein-mediated aggregation that can be largely reversed by native enzymatic digestion of either the DNA or protein components. Together, these findings offer new insights into AAV degradation pathways under thermal stress and establish an effective analytical framework for AAV stability assessment.
SN - 3117-387X
ER -